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A Just for Laughs Gag/Juste pour Rire
Mother Gives Birth to Twin Panda Babies
Kumenkaffi (Thirsty for)
Rescued Horses Lovin' Life at Oregon Sanctuary
For the following, I am providing the link sources beneath each photo. You just need to click on the images to see a larger view.
When feeling overwhelmed by a faraway goal, repeat: I have it within me right now, to get me to where I want to be later.

subset: commoncrawl


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I spy with my little eye… a vital case-cracking clue!
Youngsters at Mereside Primary School swapped their algebra books for magnifying glasses as they tried to solve the mystery of the missing parcel.
But don't worry, this isn't Lancashire Police's newest plan to deal with the latest round of swingeing cuts, merely a classroom exercise designed to get children's creative juices flowing.
The case formed part of the school's writing week, when every class from nursery to year six are encouraged to fall in love with writing.
Teacher Lauren Hollis said: "We build on the children's creativity not only through literacy-based activities but we expand their experiences with art too."


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So I weighed in today and have lost 3 lbs in one week. Realistically, I know that is mostly water. In reading my new fat loss bible…I realize that I have been retaining water and chronically dehydrated. So, this week I started drinking more water, and consequently using the bathroom A LOT! Apparently it has paid off! I have also been watching my calories and tracking with Sparkpeople. It helped that I was off work for 2 days last week, so I could really focus on eating right, with very few temptations. Today, its back to work and lunches out. Let's hope my headstart will help me stay on track.
I will not be posting a picture every week. I will post the jeans picture every 2 weeks, but will post weight weekly. Todays weight, 186 lbs.


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Voted the Best Pilates Studio in Santa Cruz, Agile Monkey is a family business that offers the highest quality instruction in Pilates and CoreAlign to accommodate all needs, fitness levels and budgets. We offer private sessions as well as group classes in Reformer, Mat, Springboard, Chair, Ball, props and more.
At Agile Monkey, we personalize each session to help you overcome any injuries or limitations and strive to help you feel truly fit and healthy. To us, Pilates is more than just exercise - it is a way to find strength and comfort in your body, so that you can focus on living the life you want.
Pilates sessions can focus on fitness, rehabilitation, sport-specific training, prenatal/postnatal and much more. Many of our instructors have decades of experience and tremendous knowledge to help you recover from injury, correct negative movement pathologies, improve posture, alleviate stress, increase muscle tone and flexibility, and help you feel better in your body.
Agile Monkey is also a Balanced Body host site, offering comprehensive Pilates instructor training programs, as well as professional continuing education courses.
Shop Local and find all your workout clothes at Agile Monkey! We carry Beyond Yoga, PrAna, Nux, Toe Sox, as well as props for home use.
"Physical fitness is the first requisite of happiness." - Joseph Pilates
My life is so much better because of Agile Monkey! I have always exercised, but always with pain. Now my back is stronger and more flexible, my balance is improved, I stand taller and feel more alive!
Voted the Best
Pilates Studio
in Santa Cruz!


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Sister in law had a wonderful birthday picnic. Some people know about partys and picnics and the perfect treats. She is like that. Good for us!
I´m wondering, how come the children don´t mind kakofonia at all? They seem to live in it. I mean if the other is on the piano, writing down " the chosen dischords" and the other simultanuesly plays the recorder next to him, and they don´t mind that soundchaos at all. I don´t get that.


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Today is the last day to complete the AJ's online survey looking at how architecture students view both their education and their prospects
The information will be used to gauge how the next generation of architects feel about the schools of architecture, their future and the profession.
The questionnaire is completely anonymous and takes less than two minutes to complete.
Complete the survey by 7 May and you could win a bag full of architectural books including a copy of Kieran Long's Hatch:The New Architectural Generation published by Laurence King.
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When Broker/Owner Tony Williamson opened the RE/MAX Cairns franchise 4 years ago his vision was to create a world class business in Cairns.
The doors opened in 2010 with a staff of just 3 and rapidly grew to a staff of 30 to be the largest Real Estate firm in Cairns.
RE/MAX Cairns covers every real estate need from residential unit and home sales including a prestige division, commercial sales and leasing, property management, new off the plan unit sales and house and land packages.
We even have a Chinese speaking assistant. At RE/MAX Cairns we strive to make your Real Estate experience nothing short of sensational!
Suite 203, 89 Scarborough St, Southport
Queensland, 4215, Australia
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AsaaSoft is a well-known software developer in Iran and even in Middle East. We have several professional teams in software department and out software products range form official and organizational softwares to sepcial purpose and RFID related ones.
Our process to develope high quality software is RUP and we doing the design with Rational Rose tools and finally we do coding with Microsoft .NET IDE.
Our products are felixible and can be customize for right-to-left or left-to-right languages such as Persian, Kurdish, Arabic, English and etc.
We are willing to participate in your outsourced projects and we can prepare some prototypes freely to make preliminary common trust. If you are seeking for a good partner in Iran plz inform us, so we will refer to you ASAP. Our contact email is: info {at} AsaaSoft {dot} net


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Get Involved
Matching Gifts/Workplace Giving
Did you know your contribution could be doubled or tripled with just a few minutes? If your employer offers matching contributions, just obtain the necessary paperwork from your human resources department, complete the donor section, and send it to us along with your gift. If you've already sent in your gift, you can still send us the completed form or online donation information. Please search our matching gifts database below to see if your company participates in matching gifts.
Once we've received your information, we'll take care of the rest!
Serving nearly 90% of people living with ALS in the Chicagoland area
Workplace giving is an easy and effective way to give to the Les Turner ALS Foundation. Whether you work for a private company or a public organization, you can use the ease and convenience of a payroll deduction to make your gift or have your employer match your charitable contributions.
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Ways to Support the Mission:
Serve as a volunteer at either a special event or on an ongoing basis. We are always looking for faces new and old to join us in the fight against ALS. If you're willing to help, there's a job for you! The rewards are well worth a few hours of effort.
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Foundation events are an easy ways to get involved in the fight against ALS and allow you to connect with other members of the community. Together we will raise funds and awareness to support research, patient services and education.
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Blog » Post
The web is a conversational medium of connections
I often hear and see the word "conversation" come up with regards to the web, especially with regards to web 2.0, social media and blogs. Basically, some web pages get described as a "conversation," rather than as communications on a shared topic, or a broadcast.
I am often critical of using the word "conversation" in this way, because there is so little listening going on among the participants who are communicating. In terms of interpersonal communication, what's called the "conversation" is more like a series of overlapping monologues spoken by individuals competing to be heard and taken seriously.
But, there is something significantly conversational about the interplay between our web pages. So, while I don't find it very useful to talk about "the conversation in the blogosphere," etc., I do find it very useful to notice the conversational interplay between websites.
The web is a conversational system in the sense that changes in one part of the system trigger changes in other parts, and the changes go back in forth—they don't just go one way, e.g., out from the center and stop at the end, like TV. Our (human) function in this system is in making changes (e.g., creating new web pages) that become effective when they introduce new links.
This conversational interplay is a kind-of conversation, but it's not a conversation between humans having a dialog in words. Rather, it's a conversation of connections. Our role in this "conversation" is in contributing additional connections in the form of new web links described in our own words, images, sounds, etc.
If we want to avoid the word "conversation" when talking about what's happening online, maybe it would work to instead talk about "clouds of new connections" or (as we do at Juxtaprose) designing websites that are about "connecting" or that "work to make connections."
Ultimately, I think we like to describe all of this as a "conversation" because we perceive so many human characteristics in the web—it's an anthropomorphism. But, it's really still up to us to be human and have actual conversations.
With the web, those actual conversations can be better informed by the great connections we now make online. But those conversations come out of us, not just of our web pages.
Blog Archives
Next post: Making podcasts more web-like


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Author: Robert Jordan and Brandon Sanderson
Genre: Fantasy
Length: 783 Pages
Release: October 2009
My Wheel of Time journey is drawing to a close. I now only have three novels in the series (including the prequel) before I finish. I've read the series a book a month since January 2016. This twelfth installment is the first of the novels completed by Brandon Sanderson after Jordan's death a few years ago. In all honesty, Sanderson is the reason I started the series. But I have appreciated Jordan's writing and the world he built.
Since this was the first Sanderson installment, I think the main thing that needs to be addressed is how it fits with the rest of the series. And it fits pretty well. It has some of Sanderson's style and touches in the humor and characterization, but he kept it very true to Jordan's story. How someone could come in after eleven books and finish it off is beyond me, the world and cast of characters can be daunting as only a reader.
The Gathering Storm ranks among my favorites of the series, but I don't necessarily think it was any better than my other favorite volumes. There are a few things that I think it improved (at least an improvement compared to the last few books, which were kind of dragging at times). Here there aren't any dull moments. The main plot and side stories that have been leading Rand and his Two Rivers friends toward the last battle move at a steady pace, with constant developments.
Many of the characters really grew on me throughout this story. Egwene, who was my least favorite character in the beginning, really started to have a different impact on me here. She has grown a lot and no longer seems as much the know-it-all that she was before. Mat is another of the characters that I was really impressed with. He was probably my favorite perspective to read from this time around (along with the few chapters we got inside Nynaeve's head).
Rand struggled a lot in this one. He is facing inner demons, the approach of the Last Battle, and it's often too much for him to take. He became somewhat more human in this one as he tried to harden himself completely.
The downside to this is really Perrin, who I used to love. Over the series he has really digressed. He did have a few good moments here, but I found myself disappointed every time one of his chapters came up. I wanted to be following someone else.
Siuan and Gareth Bryne are two of the highlights in The Gathering Storm. They really aren't in here much, but I adore their relationship and what they go through together. I found myself loving these moments.


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The chilly fall morning nipped at any exposed skin. My friends and i clambered up a steep hill as our gear threatened to drag us down the dusty slope. Huffing and puffing, we made it to a stretch of relatively flat trail. Then we saw the final steep bit… We could have taken a picture of an apple and still gotten an A for this photo project.
At the top, the rising glow of the sun illuminated the early morning fog that was draped over the quiet town of Cupertino. Colors seemed muted and bland compared to the beauty of the sun peeking over the mountains. Here was a moment that would always be remembered, a step back from the rush and chaos of life. A moment of serenity.


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Office Love Birds
Played 124379 times
Office Love Birds game information:
Holding the left click on your mouse, kiss your work collegue without your boss knowing. Are you brave enough not to get caught?
see if you have what it takes, your boss will go nuts if he catches you so make sure you dont get caught, naughty
How to play:
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Events for June 12, 2018
Events Search and Views Navigation
All Day
Techsylvania Conference 2018 Cluj
Techsylvania is a platform designed to empower the local tech community with its new and challenging perspectives brought each year. Therefore, we have prepared for you another round of inspiring keynotes, workshops, hackathon and startup pitches to spark your imaginations. The future is bold. But so are we. These continuous technological advancements are our main […]Find out more »


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About Us
News & Events
Media Center
College of Biological and Physical Sciences has a diverse collaboration with Government organizations, Education Institutions, Research organizations and Business enterprises as well as Non-Profit Organizations.
These include: Research Organizations| Education Institutions| Government Organizations| Science Based Organizations| Professional Bodies| Scientific Equipments/Suppliers| Technology/Innovation| Corporates| Youth Organization| Career Site| Business Enterprises| Company/Directory Listing|
I. Research Organizations:
|Centre de Recherches de Climatologie
|Institut De Recherche Pour Le Developpement (ird)
|Natural Product Research Network for Eastern and Central Africa
|Nile Basin Research Programme
|Research Grants on ICT for Development
II. Education Institutions
|African Mathematics Millennium Science Initiative
|Bioinformatics and Genomics Workshop
|PC Kinyanjui Technical Training Institute
|School of Mathematics and Statistics(TUK)
|School of Pure and Applied Sciences(TUK)
|Training Centre In Communication
|United Nations University (UNU-INWEH)
|University of Games
|Yokohama National University
III. Health Institutions
|LVCT Health
IV. Government Organizations
|Kenya Meteorological Department
|Kenya Power
|Kenya Water Institute
|National Oil Corporation
V. Science Based Organizations
|Biozeq Kenya Ltd
|Geometric Enterprises Ltd
|Geothermal Development Company
|IGAD Climate Prediction and Application Centre
|International Science Programme (ISP) Uppsala Universitet
|International Science Programme (ISP) Uppsala University
|International Science Programme(ISP) [Activities in Kenya]
|Kenya Climate Change Action Plan
|L 'agence Inter-établissements De Recherche Pour Le Développement (aird)
|Pan Africa Chemistry Network
|Somalia Water and Land Information Management
|World Meteorological Organization
VI. Professional Bodies
|Association of Professional Societies in East Africa (APSEA)
|Kenya Chemistry Society
|Kenya Meteorological Society
|Mathematics Association of Kenya (MAK)
VII. Scientific Equipments/Suppliers
|Chem-labs Limited
|Osho Chemical Limited
VIII. Technology/Innovation
|BTI Millman
|Empowerment Resource Technologies Limited
|Innovative Application of ICTs in Addressing Water-related Impacts of Climate Change
|Karibe Technologies
|Mackphilisa Computer Systems Ltd
|School of Information and Communication Technologies(TUK)
|ToshibaMax Ltd
IX. Corporates
|African Barrick Gold
|Centre for Sustainable Urban Development
|East African Educational Publishers Ltd.
|Fast Choice Ltd
|Worldbiz Enterprises Limited
X. Career Site
XI. Business Enterprises
|Ace Auto Centre Limited
|Amazing interiors
|Infoparts Limited
|Jabali Furnitures
|Kitenge Festival
|Natfire comapy Co Ltd
|Novia Trinkets
|Okwea Construction Company
|Olsen East Africa
|Omem Company Ltd.
|Panorama Technical Services Ltd.
|Weee Centre Kenya
|Proscene Systems Ltd
XII. Company/Directory Listing
|Reference Directory of Africa
View College of Biological Partners [PDF format] --updated May 18,2016


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About Us
Whether you're a brand-new, growing startup, or a hi-tech business, we understand you are short on time. So we're going to keep it concise.
We are 8300. We provide innovative tech solutions and services for startups and businesses just like yours, and we do it in the fast lane. We provide all the tools you need to execute your innovative idea the right way and turn it into a winning product.
How? Simple! We provide essential services for developing startups and hi-tech businesses, such as: cloud solutions and services, database optimization, development, QA, open source, training, mentoring, recruiting etc. we'll work with you to upgrade your startup or bring your established business closer to the finish line. Our promise to you is this: we work quickly, we are experts in our field and we offer excellent service.
So, how is matrix involved?
8300 is a special task-force of matrix, Israel's leading IT Integrator. This means you'll benefit from matrix's networking connections, vast experience, capabilities and know-how. You can enjoy mentoring, technologies assimilation and credentials to matrix Eco-system. That said, 8300 also operates as a separate focused unit, specially designed to accommodate growing businesses like yours, giving you the attention you need and deserve.
Our Services
Our Team
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Applicative DBA – SQL Server
This is your chance to join one of the most exciting and fastest growing startup companies.
We provide a cloud-based service designed to guide users through any online experience.
Our development team...
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We are searching for a Senior DevOps Engineer to join our fast-growing tech start up and perform as a valuable team player in our company.
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The Java Team Leader is required to lead and manage a project delivery team throughout the project life cycle, from the SDS period until the project successful completion in terms...
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We are looking for a thoughtful, well-rounded iOS developer to join our team and help build our mobile apps.
You will take a significant part in our mobile engineering practice, implementing...
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The pterosaur to bird theory contains two basic ideas, that differ from current mainstream thinking.
The first idea is that pterosaurs developed into a set of primitive bird lines.
The second is that EACH ONE of those primitive bird lines then developed into a set of corresponding modern birds, in what might be called a set of "parallel lines".
For example:
Seabirds (Ichthyornithes line)
- Pterosaur (Ornithocheiroidea) eg. Pteranodon -->
- An Ichthyornithes subgroup -->
- Gulls, Skimmers (Charadriiformes/Lari)
If any of this is not clear, I invite questions and comments.


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Copyright آ© 2011 Confederation Africaine de Volleyball- All rights reserved
25 June 2013
Cairo, Egypt, June 23, 2013- Tin Hlupic and Gerard Fischer, Namibia's U21 National Beach Volleyball team ended their FIVB World Championships campaign as one of the best African countries at the tournament held in Umag, Croatia. Namibia's duo of Hlupic / Fischer and Tunisian men's team of Arfaoui / Korbi were the only two African teams in the World Champs progressing from the group round to the next stage.
Both African teams were however not able to overcome stronger opponents in the knock-out phase of the competition and failed to make it into the final 16 of the tournament.
Fischer Hlupic were overpowered in the first set by Takvam /Sannarnes from Norway, losing the set 21-8. They then put on a strong fight in the second set, but were not able to turn around the match losing the second set 21-18.
The Tunisian duo lost their match against Kirienko / Uraikin from Russia 2:0 (21-12, 21-9).
The performance of the Tin and Gerard at the World Championships showed a great improvement compared to their level of play at the U19 Beach Volleyball World Championships in which they participated last year in Cyprus. 2013 also being the team's first year in the U21 age category leaves great expectations for years to come.
In the ladies category Namibian Julia Lagger and Saskia Henckert failed to advance from the group phase into the next round. The team however showed good performances against their counterparts from Brazil, Poland and Australia and gained significant experience on the international stage.
"Laggner and Henckert only teamed up at the beginning of this year as new Namibian combination and with the tournament being Saskia's first international competition, significant growth and performance improvements are still expected from this team", commented NVF president, Heiko Diehl.
It is however only with the financial support of Namibian companies, such as Henckert Tourist Centre, African Packaging, Windhoek Consulting Engineers, Rockwater Consulting, Scania Namibia, Absolute Logistics as well as the support of CB Enterprises, Denchi Consulting Engineers, South African Airways and the African Volleyball Confederation (CAVB), that our teams can grow and develop into one of the top teams in world.
After returning from the World Champs, the teams will now sit down and start setting their next goals, together with National team coach, Achim Lück. The Namibian Volleyball Federation is also currently working towards one of their next goals, qualifying one of Namibia's top youth Beach Volleyball teams for next year's Youth Olympic Games.
Gaborone, Botswana, December 11, 2018- Africa Union Sport Council "AUSC Region 5 Volleyba
Durban, South Africa, December 7, 2018- Police 6 of Botswana claimed Zone 6 Club Championship following th
Dakar, Senegal, February 5, 2017
Abidjan, Cote d'Ivoire, Ja


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Zulu Grey
Quarried by hand in small numbers, the Zulu Grey is a unique, indurated sedimentary rock formed over 200 million years ago in the late Permian Period in the province of Kwa Zulu-Natal, South Africa. The stone is quite hard, exceptionally strong and dense. The Zulu Grey whetstone is quarried for its ability to impart a smooth final shaving edge on Straight Razors, however it would be equally effective at finishing a wide variety of fine-bladed tools and instruments. The stone is of course a natural product and as such each hone has an inherently unique appearance. Colours of the whetstones range from Grey-Blue to Grey-Green with a fine speckled pattern on the face of the stone. Some of the hones may also have what appear to be orange swirls or stains – this is created by iron oxide present in the stone and is quite beautiful in appearance.
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Oxygen is essential for eukaryotic life and is inextricably linked to the evolution of multicellular organisms. Proper cellular response to changes in oxygen tension during normal development or pathological processes, such as cardiovascular disease and cancer, is ultimately regulated by the transcription factor, hypoxia‐inducible factor (HIF). Over the past decade, unprecedented molecular insight has been gained into the mammalian oxygen‐sensing pathway involving the canonical oxygen‐dependent prolyl‐hydroxylase domain‐containing enzyme (PHD)–von Hippel‐Lindau tumour suppressor protein (pVHL) axis and its connection to cellular metabolism. Here we review recent notable advances in the field of hypoxia that have shaped a more complex model of HIF regulation and revealed unique roles of HIF in a diverse range of biological processes, including immunity, development and stem cell biology.
The canonical HIF pathway
Oxygen‐dependent regulation of HIF signalling
Hypoxia‐inducible factor (HIF) is a DNA‐binding transcription factor that associates with specific nuclear cofactors under hypoxia to transactivate a myriad of genes to trigger various adaptive responses to compromised oxygen tension. HIF is a heterodimer comprising an oxygen‐labile α‐subunit (HIFα) and a constitutively expressed β‐subunit (HIFβ or ARNT). Both subunits are part of the basic Helix–Loop–Helix PER‐ARNT‐SIM (bHLH‐PAS) family of transcription factors. Three genes encoding distinct HIFα isoforms exist in humans: HIF1A, encoding HIF1α; EPAS1, encoding HIF2α; and HIF3A, which is expressed as multiple HIF3α splice variants (Wang et al, 1995; Makino et al, 2002; Maynard et al, 2003). HIF1α, HIF2α, and HIF3α splice variants 1–3 possess an oxygen‐dependent degradation domain (ODD) and an N‐terminal transactivation domain (NTAD), while HIF1α and HIF2α possess a C‐terminal transactivation domain (CTAD; Jiang et al, 1997; Pugh et al, 1997; Gu et al, 1998; Huang et al, 1998; Ema et al, 1999; O'Rourke et al, 1999; Hara et al, 2001; Maynard et al, 2003). In this review, HIF isoforms will be specified where appropriate, while 'HIFα' will be used to denote either HIF1α or HIF2α.
Under normoxia (normal oxygen tension), HIFα is hydroxylated on at least one of two conserved proline residues within the ODD (Ivan et al, 2001; Jaakkola et al, 2001; Masson et al, 2001) by prolyl‐hydroxylase domain (PHDs)‐containing enzymes (Epstein et al, 2001). Hydroxy‐HIFα is recognized by the β‐domain of von Hippel‐Lindau tumour suppressor protein (pVHL) and is subsequently ubiquitylated by the Elongin BC/Cul2/pVHL ubiquitin–ligase complex assembled via the pVHLα domain, thereby marking HIFα for degradation by the 26S proteasome (Maxwell et al, 1999; Ohh et al, 2000). PHDs require oxygen for catalytic activity; thus, prolyl hydroxylation of HIFα is abrogated under hypoxia, allowing HIFα to escape recognition by the pVHL ubiquitin–ligase complex and accumulate in the nucleus. Similarly, biallelic inactivation of VHL has a profound stabilizing effect on HIFα protein levels, thereby implicating pVHL as a predominant HIFα antagonist. Notably, another hydroxylase‐domain protein termed factor inhibiting HIF (FIH) participates in the negative regulation of HIFα by hydroxylating asparagine‐803 in the CTAD in the presence of oxygen, which sterically inhibits interactions between HIFα and transcriptional coactivators (Lando et al, 2002).
Activation of HIF in hypoxia
Hypoxia is defined in the context of tumours as having an internal partial pressure of oxygen of less than 10–15 mm Hg (Brizel et al, 1999; Khan et al, 2012). In hypoxic conditions or in VHL−/− cells, stabilized HIFα dimerizes with HIFβ before binding to hypoxia‐response elements (HREs) comprising a core 5′‐[A/G]CGTG‐3′ consensus sequence and highly variable flanking sequences in the promoters of HIF‐responsive genes (Wenger et al, 2005). FIH‐mediated asparagine‐803 hydroxylation is also reduced when oxygen is depleted, thereby allowing HIFα to interact with the transcriptional coactivators p300/Creb‐binding protein (CBP; Arany et al, 1996). This transcriptional complex transactivates a specific subset of genes that regulate the cellular adaptive response to hypoxia, including, but not limited to, SLC2A1 (glycolysis), VEGFA (angiogenesis), and EPO (erythropoiesis).
Expanding the canonical HIF pathway
SIRT3 is a novel HIF1α antagonist
Regulation of HIFα stability via ROS. Oxygen tension and the functional status of pVHL are just two of many factors governing HIFα stability. HIFα protein levels are in part a function of HIFα mRNA stability, which can be negatively regulated by miRNAs (Bruning et al, 2011; Taguchi et al, 2008) and mRNA‐destabilizing proteins (Chamboredon et al, 2011). Post‐translational modifications (PTMs) of HIFα, such as small ubiquitin‐like modifier (SUMO)ylation (Carbia‐Nagashima et al, 2007; Cheng et al, 2007) and acetylation (Xenaki et al, 2008; Dioum et al, 2009; Lim et al, 2010), have been reported to affect HIFα stability in a proteasome‐dependent manner. PHDs negatively regulate HIFα at the protein level via oxygen‐dependent prolyl hydroxylation as described above, but the catalytic activity of PHDs is in turn governed by a variety of inhibitors and/or cofactors in the cellular environment (Figure 1A).
In addition to oxygen, PHDs require Fe2+, 2‐oxoglutarate, and ascorbate for prolyl‐hydroxylase activity (Schofield and Ratcliffe, 2004). In contrast, the enzymatic function of PHDs has been reported to be inhibited by nitric oxide, several metabolic intermediates of the tricarboxylic acid (TCA) cycle such as succinate and fumarate, and reactive oxygen species (ROS; Kaelin and Ratcliffe, 2008). The inverse relationship between prolyl‐hydroxylated HIFα and intracellular ROS had been reported by independent groups (Brunelle et al, 2005; Mansfield et al, 2005) prior to the demonstration that peroxide‐derived ROS directly inhibited PHD catalytic activity, presumably by oxidizing PHD‐bound Fe2+ (Pan et al, 2007). However, the relationship between ROS production and HIFα activity has become increasingly complex as FIH was recently demonstrated to have significantly greater sensitivity to oxidative stress than PHDs (Masson et al, 2012), and respiring mitochondria have been reported to produce significantly lower ROS during hypoxia than under normoxia (Hoffman et al, 2007). Thus, the specific role and relative significance of ROS in mediating the hypoxic response remain unclear.
SIRT3 destabilizes HIF1α in a PHD‐dependent manner. The mitochondrial deacetylase sirtuin‐3 (SIRT3) was reported to have a tumour suppressor function when SIRT3−/− cells were observed as having aberrant intracellular metabolism, decreased mitochondrial integrity, and a tumourigenic phenotype relative to wild‐type cells (Kim et al, 2010). However, the specific signalling axis through which these phenomena were occurring remained unclear. Finley et al (2011a) recently reported that SIRT3 destabilizes HIF1α by inhibiting ROS production, thereby promoting maximal prolyl‐hydroxylase activity of PHDs. The net effect of SIRT3‐mediated HIFα destabilization was a shift away from glycolytic metabolism and unchecked proliferation. Specifically, SIRT3‐knockout mouse embryonic fibroblasts (MEFs) demonstrated increased glucose uptake, lactate production, and glucose‐dependent proliferation rates relative to wild‐type cells. Gene set enrichment analysis revealed a hypoxic signature in SIRT3‐knockout MEFs, prompting an investigation into the role of SIRT3 in HIFα‐mediated transactivation of proglycolytic genes. Stable knockdown or knockout of SIRT3 resulted in an intracellular increase in ROS production, which appeared to have a stabilizing effect on HIF1α protein levels under normoxia. This effect was shown to be due at least in part to a decrease in PHD hydroxylase activity, although direct inhibition of PHDs by ROS in this context was not clearly demonstrated. Importantly, stable knockdown of HIF1α in both SIRT3−/− and wild‐type MEFs diminished the metabolic differences previously observed between them. Finally, SIRT3 deletions were found to occur in 20% of all human cancers and 40% of all breast and ovarian cancers. Together, these findings confer a specific tumour suppressor function of SIRT3 that counteracts the switch to anaerobic metabolism under normoxia, referred to as the Warburg effect, promoted by ROS generation and HIF1α stabilization.
An independent study supports the conclusion that loss of SIRT3 stabilizes HIF1α, perhaps through ROS production, and also demonstrates that expression of some non‐metabolic HIF1α‐target genes, such as VEGFA, increases with knockdown of SIRT3 (Bell et al, 2011). This suggests that SIRT3 may oppose additional HIF1α‐driven cancer phenotypes beyond the Warburg effect, such as angiogenesis, apoptotic resistance or cellular migration/invasion.
SIRT3 suppresses ROS production. SIRT3 is a predominant mitochondrial deacetylase and is responsible for regulating the activities of a significant subset of mitochondrial proteins, many of which are strongly implicated in cellular metabolism (Lombard et al, 2007). SIRT3 likely suppresses ROS production through several proposed mechanisms, including but not necessarily limited to deacetylation and activation of the antioxidant enzyme superoxide dismutase (SoD; Qiu et al, 2010; Tao et al, 2010), modulation of certain electron transport chain (ETC) components (Ahn et al, 2008; Kim et al, 2010), and activation of several TCA cycle enzymes (Bell and Guarente, 2011). Specifically, deacetylation and activation of isocitrate dehydrogenase 2 (IDH2) by SIRT3 was reported to promote the production of NADPH, which is requisite for proper function of mitrochodrial ROS detoxifying enzymes (Someya and Yu, 2010). In addition, SIRT3 was shown to deacetylate and activate succinate dehydrogenase (SDH), a component of the ETC, which could potentially affect ROS production (Cimen et al, 2010; Finley et al, 2011b). SDH also catalyses the oxidation of succinate to fumarate in the TCA cycle; thus, SIRT3 may negatively regulate intracellular succinate levels (Finley et al, 2011b). Succinate, like ROS, has been reported to inhibit PHDs (Koivunen et al, 2007), which suggests that intracellular accumulation of succinate in SIRT3−/− MEFs contributes to the stabilization of HIF1α via PHD inhibition (Figure 1A) as observed by Finley et al (2011a), although this possibility has yet to be directly addressed.
Sirtuins and the HIF pathway. SIRT3 is not the first sirtuin to be implicated in HIFα regulation (Figure 1B). Sirtuins are NAD+‐dependent protein deacetylases and/or ADP‐ribosyltransferases known to modulate longevity and metabolism (Imai et al, 2000; Tissenbaum and Guarente, 2001). SIRT1 has been reported to deacetylate both HIF1α (Lim et al, 2010) and HIF2α (Dioum et al, 2009), causing a decrease and, unexpectedly, an increase in transcriptional activity, respectively. SIRT1, in turn, was shown to be upregulated by both HIF1α and HIF2α under hypoxia, which suggests that SIRT1 may play a role in negative‐ and positive‐feedback loops that govern HIFα activity (Chen et al, 2011). In contrast, SIRT6 may negatively regulate HIF1α stability and protein synthesis via yet‐defined mechanism, and may also serve as a HIF1α corepressor to impede the induction of HIF1α‐responsive genes (Zhong et al, 2010). Thus, SIRT3 is currently the only sirtuin family member that has been demonstrated to destabilize HIF1α through a PHD‐dependent mechanism.
PKM2 is a novel HIF1α coactivator
HIFα‐mediated transactivation of target genes is modulated by the relative abundance of transcriptional coactivators and/or corepressors, as well as PTMs that affect the physical accessibility of, or affinity between, HIFα and its cofactors (Figure 1B). For example, de‐SUMOylation of p300 by sentrin‐specific protease (SENP)3 under conditions of oxidative stress enhances binding between p300 and HIF1α, thereby increasing HIF1α transcriptional activity (Huang et al, 2009). Acetylation of lysine‐674 on HIF1α by p300/CBP‐associated factor (PCAF) also enhances binding between p300 and HIF1α, while deacetylation of the same residue by SIRT1 impedes p300 recruitment and transactivation of HIF1α‐target genes (Xenaki et al, 2008; Lim et al, 2010). CBP/p300 interacting transactivator with ED‐rich tail (CITED)2 and CITED4 have been shown to bind p300/CBP at the same domain required for binding to HIFα, effectively blocking the HIF1α/p300 interaction and diminishing HIF1α activity (Bhattacharya et al, 1999; Fox et al, 2004). Recently, the chromatin remodelling factor Pontin was demonstrated to be a HIF1α coactivator by mediating the binding between p300 and HIF1α under hypoxia (Lee et al, 2011). Together, these reports demonstrate that the transcriptional activity of HIFα can be efficiently regulated by modulating recruitment of p300 and other cofactors to the HIFα transcriptional complex.
PKM2 enhances HIF1α transcriptional activity. Recently, Luo et al (2011) identified a novel role for the M2 isoform of pyruvate kinase (PKM2), the enzyme that catalyses the final step in glycolysis, as a coactivator of HIF1α and HIF2α (Figure 1B). PKM2 expression was induced by HIF1α, but not HIF2α, suggesting PKM2 and HIF1α participate in a positive‐feedback loop that increases their respective activities under hypoxia. PKM2 was demonstrated to associate directly with HIF1α and HIF2α, enhance binding of HIF1α and HIF2α to HREs, and promote HIF1α‐ and HIF2α‐mediated transactivation of hypoxia‐inducible genes without affecting total HIFα levels. This effect was purportedly enhanced by prolyl hydroxylation of PKM2 by PHD3, and PHD3 was shown to coprecipitate with exogenous HIF1α/PKM2 complex. While the notion that PHD3, a negative regulator of HIFα protein levels, may be implicated in the positive regulation of HIFα activity is certainly interesting, further study is necessary to more clearly define the function of PHD3 in PKM2‐mediated coactivation of HIF1α. Importantly, PKM2 also appeared to bind p300 (although perhaps indirectly), and stable knockdown of PKM2 reduced p300 occupancy of HREs, suggesting PKM2 enhances recruitment of p300 to the HIFα transcriptional complex at the promoters of HIFα‐responsive genes. These results indicate that PKM2 promotes a shift toward anaerobic metabolism by augmenting HIFα‐mediated transactivation of proglycolytic genes such as LDHA and SLC2A1.
An independent report corroborated the finding that HIFα upregulates PKM2, and that this appears to promote anaerobic metabolism in tumour cells (Sun et al, 2011). It was also suggested that mammalian target of rapamycin (mTOR) upregulates PKM2 via two distinct mechanisms: mTOR‐mediated upregulation of HIFα may lead to transactivation of PKM2, while concomitantly, mTOR‐mediated upregulation of c‐myc may lead to c‐myc‐mediated upregulation of heterogeneous nuclear ribonucleoproteins (hnRNPs), which are reported to favour splicing of PKM2 mRNA over PKM1 mRNA (David et al, 2010; Sun et al, 2011). Thus, in addition to upregulating HIFα, mTOR may modulate HIFα signalling by upregulating the HIFα coactivator PKM2 (Figure 1B).
Role of PKM2 in the Warburg effect. PKM2 has been previously implicated in the Warburg effect observed in cancer cells (Christofk et al, 2008), although its specific role in promoting aerobic glycolysis has remained unclear. Recent work suggests, perhaps counter‐intuitively, that PKM2 promotes glycolysis in tumour cells in a manner that is independent of its biochemical function as a glycolytic enzyme. Phosphorylation of PKM2 at tyrosine‐105 (Y105) inhibits its glycolytic activity, and PKM2 was found to be phosphorylated at Y105 in several cancer cell lines (Hitosugi et al, 2009). Furthermore, cells expressing a PKM2 mutant defective in Y105 phosphorylation—that is, a PKM2 mutant possessing greater glycolytic activity than wild‐type PKM2—exhibited increased oxidative phosphorylation, decreased lactate production, and slowed tumour growth relative to cells expressing wild‐type PKM2 (Hitosugi et al, 2009). These data imply that the enzymatic function of PKM2 actually opposes the Warburg effect. Indeed, Luo et al (2011) demonstrate that a catalytically inactive form of PKM2 promotes anaerobic metabolism via HIFα transactivation as efficiently as wild‐type PKM2. Together, these studies conjure a potential model of metabolic reprogramming in tumour cells whereby PKM2 undergoes a functional shift from glycolytic enzyme to HIFα coactivator.
PKM2 as a transcriptional coactivator. PKM2 has also been reported to act as a transcriptional coactivator for β‐catenin (Yang et al, 2011). Specifically, activation of epidermal growth factor receptor (EGFR) was shown to trigger nuclear translocation of PKM2, where it appeared to bind phosphorylated β‐catenin and subsequently potentiate β‐catenin‐mediated transactivation of cyclin D1 (Yang et al, 2011). Thus, while PKM2 purportedly promotes glycolysis as a HIFα coactivator (Luo et al, 2011), it may also promote cellular proliferation and cell cycle progression via β‐catenin‐mediated upregulation of cyclin D1 (Yang et al, 2011). Therefore, the tumour phenotype to which PKM2 contributes may depend upon the specific transcription factor being activated and its particular subset of target genes. Given the findings reported by Yang et al, (2011) and considering that PKM2 must be cytoplasmic to participate in glycolysis, it would seem likely that a nuclear translocation of PKM2 is necessary and precedes coactivation of HIFα. Future work on the role of PKM2 in HIFα‐mediated transcription might address this possibility.
Expanding roles of HIF in cell biology
The role of HIFα as a transcription factor in the context of oxygen sensing and tumourigenesis has been extensively characterized. While the aforementioned reports serve to extend our current understanding of HIFα regulation in cell signalling, emerging evidence has implicated a larger role for HIFα in cell biology.
Role of HIF in embryonic development
HIF1α drives a metabolic shift to glycolysis during ESC–EpiSC transition. Analogous to the Warburg effect observed in cancer cells, Zhou et al (2012) have recently shown that pluripotent embryonic stem cells (ESCs) also undergo a metabolic shift towards glycolysis during development and this process is regulated by HIF1α. ESCs and epiblast stem cells (EpiSCs) represent two distinct pluripotency stages in embryonic development with unique gene expression profiles and cell morphologies, but a common ability to self‐renew and differentiate into diverse cell types (Tesar et al, 2007). Mouse (m)ESCs isolated from the inner cell mass (ICM) of pre‐implantation blastocysts represent a naive state, while EpiSCs isolated from the post‐implantation epiblast represent a more developmentally advanced or 'primed' state (Brook and Gardner, 1997; Evans and Kaufman, 1981; Tesar et al, 2007; Nichols and Smith, 2009). Notably, unlike mESCs, human (h)ESCs, despite their pre‐implantation origin, are not considered a naive state and are similar to EpiSCs (Tesar et al, 2007). Recent work by Zhou et al (2012) revealed that EpiSCs and hESCs have lower mitochondrial respiration and higher levels of glycolysis in comparison to mESCs, which switch between both oxidative phosphorylation and glycolysis (Figure 2). Strikingly, inhibition of glycolysis with the glucose analogue 2‐deoxy‐glucose (2‐DG) led to cell death in EpiSCs and hESCs, exemplifying their dependence on this metabolic pathway for survival (Zhou et al, 2012). Similarly, haematopoietic stem cells (HSCs) have been recently shown to utilize glycolysis instead of mitochondrial respiration, suggesting that HIFα may regulate the metabolism of multiple stem cell populations (Simsek et al, 2010).
HIF1α has previously been shown to promote a metabolic switch to glycolysis in response to chronic hypoxia through the transcriptional regulation of key metabolic enzymes such as pyruvate dehydrogenase kinase‐1 (PDK1) and lactate dehydrogenase A (LDHA) (Kim et al, 2006; Seagroves et al, 2001). PDK1 phosphorylates and inactivates pyruvate dehydrogenase (PDH), which inhibits the conversion of pyruvate to acetyl coenzyme A (AcCoA) for entry into the citric acid cycle (Semenza, 1996). In addition, HIF1α‐mediated upregulation of LDHA increases the conversion of pyruvate to lactate to promote glycolysis (Semenza et al, 1996). Consistent with the reported role of HIF1α in regulating metabolism, Zhou et al (2012) found that HIF1α expression and several HIF‐regulated glycolytic genes such as PDK1, LDHA, and PYGL were significantly higher in EpiSCs compared with mESC (Zhou et al, 2012). Furthermore, despite their more developed mitochondria and increased mitochondrial content, EpiSC had lower mitochondrial respiration that was attributed to downregulated expression of mitochondrial cytochrome c oxidase (COX) genes and their regulators peroxisome proliferator‐activated receptor‐γ coactivitor‐1β (PGC‐1β) and oestrogen receptor‐related receptor‐β (Esrrb) (Zhou et al, 2012). HIF1α has previously been shown to downregulate PGC‐1β through inhibition of c‐Myc transcriptional activity (Zhang et al, 2007), suggesting that HIF1α may actively suppress mitochondrial respiration while promoting glycolysis through the regulation of key metabolic genes (Figure 2). In support of this notion, Zhou et al (2012) demonstrated that overexpression of HIF1α was sufficient to suppress oxidative phosphorylation, enhance glycolysis, and promote the transition from ESC to EpiSC.
Similarly, independent studies have shown that in comparison to differentiated cells, undifferentiated mESC have decreased mitochondrial respiration concomitant with increased glycolytic metabolism, which is necessary to maintain pluripotency (Mandal et al, 2011; Zhang et al, 2011). While Zhang et al suggest that expression of the inner mitochondrial protein, uncoupling protein 2 (UCP2), is responsible for the glycolytic shift in undifferentiated hESC, several reports have demonstrated a positive correlation between HIFα and UCP2 expression (Zhang et al, 2011; Deng et al, 2012). In addition, putative HIF binding sites have been identified in the UCP2 promoter (Oberkofler et al, 2005), suggesting that HIFα may control the expression of UCP2 and several other glycolytic and mitochondrial proteins to promote a universal metabolic shift.
Significance of glycolysis in EpiSC/hESC. Several theories have been proposed to explain the preference for glycolysis in EpiSC/hESC. Although glycolysis is energetically unfavourable in comparison with oxidative phosphorylation (yielding a mere 2 ATP in comparison with 36 ATP per glucose), this metabolic shift helps to promote cell proliferation through increased production of metabolic intermediates for amino acid, protein, and membrane synthesis. This may allow the developing blastocyst to acquire the necessary building blocks for rapid proliferation and differentiation. Second, oxidative phosphorylation is associated with the production of ROS, a known DNA mutagen. Thus, a shift to glycolytic metabolism may be a protective mechanism for the cell to maintain genomic integrity within pluripotent cells. Third, glycolysis promotes extracellular acidification through increased production of lactic acid, which may aid in invasion and implantation of the blastocyst into the uterine wall.
HIF maintains pluripotency in ESC through TGFβ/Activin/Nodal signalling. Analogous to undifferentiated HSCs, which have recently been shown to regulate quiescence and long‐term maintenance through the regulation of HIFα expression levels, ESC may maintain pluripotency through a HIF‐dependent mechanism (Takubo et al, 2010). TGFβ/Activin/Nodal signalling has previously been shown to drive the ESC to EpiSC transition and maintain pluripotency in EpiSC and hESC by preventing spontaneous differentiation into neuroendoderm (Vallier et al, 2004; James et al, 2005; Camus et al, 2006). Activin B is a direct transcriptional target of HIFα, and hypoxia has recently been shown to promote an ESC to EpiSc transition, suggesting a role for HIFα in not only the metabolic changes, but also the phenotypic changes that occur during ESC development (Wacker et al, 2009; Takehara et al, 2011). In support of these previous findings, Zhou et al (2012) found that inhibition of Activin signalling was sufficient to block the HIF‐induced transition from ESC to EpiSC. Furthermore, Activin signalling has been shown to decrease PHD2 and stabilize HIF1α, suggesting that a positive‐feedback loop may exist to further amplify signalling (Wiley et al, 2010; Figure 2). In support of this notion, Zhou et al (2012) found that activation of Activin signalling increased both HIF1α expression and glycolytic genes, while downregulating mitochondrial genes. Interestingly, PGC‐1β has been shown to be directly downregulated by Activin/Nodal signalling, providing additional evidence to support a role for Activin/Nodal signalling in the active suppression of mitochondrial metabolism (Li et al, 2009). These results infer a novel mechanism by which HIF regulates Activin/Nodal signalling to alter the expression of key metabolic genes, promoting a highly glycolytic metabolism to promote the ESC to EpiSC transition (Figure 2).
Regulation of HIF during early embryonic development. Both the pre‐implantation and post‐implantation blastocysts exist in the hypoxic environment of the uterus (Fischer and Bavister, 1993). However, Zhou et al (2012) observed considerable differences in the expression level of HIF between these two pluripotent states. Despite the ability of the post‐implantation blastocyst to access the vasculature of the surrounding tissue, the EpiSC had higher HIF1α expression. This apparent contradiction warrants further research into the mechanisms underlying HIF expression in pre‐ and post‐implantation ESCs. Does the post‐implantation blastocyst experience lower oxygen tension in comparison to the pre‐implantation ESC or are there other unique or non‐canonical mechanisms of HIF regulation at play during early embryonic development? Earlier work by Fischer and Bavister (1993) demonstrated that hESC utilize glycolysis for metabolism even when removed from the hypoxic environment of the blastocyst. These results indicate that factors other than oxygen tension may regulate HIFα levels (discussed previously) to alter glucose metabolism and promote ESC to EpiSC transition.
The metabolism of HSCs was recently shown to be controlled by the haematopoietic transcription factor, Meis1, which directly activates HIF1α expression (Simsek et al, 2010). These findings suggest that the high expression of HIF1α and shift to glycolytic metabolism is not just a result of the hypoxic environment of the bone marrow, but is tightly regulated by haematopoietic stem‐cell–associated transcription factors. Whether or not unique ESC‐associated transcription factors regulate HIFα in ESC remains to be determined.
Role of HIF in immunity
Inflamed tissues are often hypoxic as a result of decreased perfusion, oedema, vascular insult and/or influx of oxygen‐consuming immune cells or pathogens (Nizet and Johnson, 2009). Thus, the role of HIFα in host immune responses and inflammation has become an important research focus. HIFα expression is stabilized through a variety of mechanisms in a range of immune cell types (Nizet and Johnson, 2009). For instance, recognition of pathogen‐associated molecular patterns (PAMPs) such as lipopolysaccharide (LPS) by toll‐like receptors (TLRs) in human monocytes was reported to activate the mitogen‐activated protein kinase (MAPK) pathway and ultimately NF‐κB, thereby transactivating HIF1A and producing an abundance of HIF1α mRNA and protein under normal oxygen tension (Frede et al, 2006). In addition to canonical stabilization of HIFα protein levels, hypoxia was demonstrated to prohibit PHD‐mediated inhibition of IκB kinase‐β (IKK‐β) (Cummins et al, 2006), thereby leading to activation of NF‐κB and subsequent upregulation of HIF1A as demonstrated in murine macrophages (Rius et al, 2008). The mechanism of HIFα stabilization in T cells appears to be analogous to that in myeloid cells, as engagement of T cell receptors (TCRs) by foreign antigens upregulates HIF1α via phosphoinositol‐3‐kinase (PI3K)/mTOR signalling pathway (Nakamura et al, 2005). Together, these reports demonstrate that stabilization of HIFα in the context of inflammation and immunity can occur through multiple signalling cascades in both normoxic and hypoxic environments.
Following stabilization, HIF1α plays an important role in executing the immune response. Metabolic reprogramming in myeloid cells by HIF1α was shown to increase the production of glycolysis‐derived ATP, a critical energy source for macrophage aggregation, invasion, and motility (Cramer et al, 2003). In addition, HIF1α‐mediated upregulation of β‐integrin was reported to enhance neutrophil adhesion to activated endothelium (Kong et al, 2004). Neutrophil apoptosis was shown to be significantly attenuated in hypoxia by HIF1α (Walmsley et al, 2005) and more recently, PHD3 (Walmsley et al, 2011), although whether this latter effect occurs in a HIF1α‐dependent or ‐independent manner remains unclear. HIFα also contributes to the production of a number of proinflammatory cytokines such as TNFα, interleukin (IL)‐1α, IL‐1β, IL‐6, and IL‐12, which are necessary for macrophage activation and phagocytic function, as well as lymphocyte proliferation (Peyssonnaux et al, 2005, 2007; Jantsch et al, 2008; Nizet and Johnson, 2009). In some cases, such as TNFα, HIFα‐mediated upregulation occurs as a result of direct transactivation. While the aforementioned studies might suggest that HIFα acts predominantly as a proinflammatory factor, it is worth noting that HIF1α has been demonstrated to have a robust anti‐inflammatory effect in hypoxic gastrointestinal mucosa and epithelium as a result of transactivation of specific genes that have a 'protective‐barrier' effect, such as mucins (Furuta et al, 2001; Comerford et al, 2002; Louis et al, 2006). This effect has also been successfully demonstrated in relevant animal models (Karhausen et al, 2004; Cummins et al, 2008; Robinson et al, 2008). Together, these roles indicate that HIFα depends upon its transcriptional activity in directing the immune response in a wide range of cell types.
HIF1α promotes differentiation of TH17 cells by several distinct mechanisms. Interestingly, HIFα also appears to regulate the differentiation of multiple leukocyte lineages. Hif1a−/− mice exhibit impaired B cell maturation, and HIF1α has been shown to modulate this process by initiating glycolysis at specific stages of B cell differentiation (Kojima et al, 2002, 2010). Similarly, mTOR‐mediated upregulation of HIF1α was reported to induce differentiation of naive T cells to proinflammatory TH17 cells, as opposed to anti‐inflammatory Treg cells, by promoting glycolysis (Shi et al, 2011). These reports further support the notion that HIF1α contributes to B and T cell differentiation through its role as a transcription factor.
A recent report from Dang et al (2011) demonstrates that the effect of HIF1α on T cell differentiation extends beyond its transcriptional activity. In TH17‐skewing conditions (IL‐6 with low levels of TGF‐β), signal transducer and activator of transcription 3 (STAT3) was shown to transactivate HIF1A. STAT3 was previously reported to positively influence TH17 cell differentiation, presumably by upregulating the key TH17 transcriptional regulator RORγt (Harris et al, 2007), but HIF1α had not been previously implicated in this mechanism. Dang et al (2011) observed decreased expression of TH17 signature genes such as IL‐17 in murine Hif1a−/− T cells relative to wild‐type T cells, even after culturing in TH17‐skewing conditions. In addition, the proportion of Hif1a−/− T cells that stained positive for Foxp3, the predominant transcriptional regulator of Treg cells, was significantly greater than in wild‐type T cells. Furthermore, incubation of TH17‐skewed wild‐type T cells in hypoxia increased the IL‐17+ population, but hypoxic treatment had no effect on IL‐17 expression in TH17‐skewed Hif1a−/− T cells. These results imply that HIF1α supports TH17 cell differentiation by promoting expression of TH17 signature genes.
Dang et al (2011) elucidated the precise mechanism by which HIF1α upregulates IL‐17 by demonstrating direct transactivation of RORγt by HIF1α. RORγt then directly induces expression of IL‐17A and IL‐17R in response to TGF‐β and IL‐6, thereby promoting differentiation of naive CD4+ T cells to TH17 cells (Ivanov et al, 2006). Thus, these results place HIF1α at a central point in the IL‐6/TGF‐β–STAT3–RORγt signalling cascade that drives TH17 cell differentiation.
Upregulation of RORγt by HIF1α is dependent upon the transcriptional activity of HIF1α (Figure 3A), a mechanism that is analogous to the aforementioned modes of HIF1α‐mediated inflammation and differentiation of lymphocyte lineages. However, one key observation—that RORγt overexpression in Hif1a−/− T cells did not significantly increase the IL‐17+ population—suggested that HIF1α might have additional roles in TH17 cell differentiation downstream of RORγt transactivation. Intriguingly, HIF1α and RORγt were shown to directly interact with one another, and induction of IL‐17 by HIF1α did not require a functional DNA‐binding domain (Dang et al, 2011). In fact, the binding site for RORγt was mapped to amino acids 1–80 on the N terminus of HIF1α, a site that is normally required for HIFβ dimerization and binding to HREs (Jiang et al, 1996; Dang et al, 2011). Dang et al (2011) propose a model (Figure 3B) in which HIF1α transactivates the RORγt gene prior to physically associating with its protein product at the promoters of TH17 signature genes. The purpose of HIF1α in this transcriptional complex appears to be recruitment of p300 through its CTAD, which improves the accessibility of RORγt target gene promoters through p300‐mediated acetylation of histones H3 and H4 (Dang et al, 2011).
Finally, it is proposed that HIF1α actively suppresses Treg differentiation by targeting Foxp3 for ubiquitin‐mediated degradation (Figure 3C). While the identification of a possible degradation complex remains to be elucidated, downregulation of Foxp3 by HIF1α appears to be a post‐translational, proteasome‐dependent phenomenon requiring direct interaction between Foxp3 and the N terminus of HIF1α. Expression of hydroxylation‐defective HIF1α mutants, as well as siRNA‐mediated knockdown of PHD2, abrogated Foxp3 degradation (Dang et al, 2011), suggesting prolyl hydroxlation of HIF1α necessarily precedes Foxp3 degradation. However, ubiquitylation and degradation of Foxp3 was also shown to significantly increase under hypoxia, a state in which HIF1α levels increase but prolyl hydroxylation of HIF1α by PHD2 is impeded. Therefore, further studies are required to clarify the role of prolyl‐hydroxylated HIF1α in Foxp3 degradation.
This work is particularly interesting in that it demonstrates HIF1α promoting TH17 cell differentiation via three unique and functionally distinct roles: a DNA‐binding transcription factor, a transcriptional coactivator for RORγt, and a direct mediator of Foxp3 degradation (Figure 3). Notably, all three roles require the same region of HIF1α: the HLH domain, which is necessary for HIF heterodimerization and DNA binding (Jiang et al, 1996), and was shown in this report to bind RORγt and Foxp3. Therefore, it is formally possible that these three functions are mutually exclusive. To better understand the dynamic shifts in HIF1α function in T cell differentiation, it may be informative to elucidate the relative binding affinities of HIFβ, RORγt, and Foxp3 to the HLH domain of HIF1α.
This report also implicates HIF1α in the T cell–mediated inflammatory response. Hif1a−/− T cells failed to develop experimental autoimmune encephalitis in response to myelin oligodendrocyte glycoprotein peptide, whereas wild‐type T cells exhibited a robust inflammatory reaction. This observation provides a rationale for investigating the use and efficacy of HIFα inhibitors in the treatment of T cell mediated autoimmune disorders.
Together, these findings outline a positive‐feedback mechanism by which TGF‐β and IL‐6 upregulate HIF1A in a STAT3‐dependent manner, leading to transactivation of RORγt and subsequently HIF1α/RORγt/p300‐mediated transactivation of TH17 signature genes. Upregulation of IL‐17, in addition to HIF1α‐mediated degradation of Foxp3, appears to favour TH17 cell differentiation and impede the development of Treg cells, thereby enhancing the inflammatory response. Ultimately, an increase in local inflammation brings about a decrease in oxygen tension, and the resulting stabilization of HIF1α would presumably act to amplify the entire cascade.
Role of HIF in Notch‐dependent developmental pathways
HIFα modulates human leukocyte differentiation via canonical and non‐canonical pathways, as described above. However, HIFα can induce or suppress the differentiation of a multitude of cell types, including but not limited to adipocytes, medulloblastoma precursor cells, neuroblastoma cells, glioblastoma stem cells, and neural crest progenitors (Jogi et al, 2002; Yun et al, 2002; Sainson and Harris, 2006; Pistollato et al, 2010; Qiang et al, 2012). A number of these reports have implicated Notch, a well‐established mediator of cellular differentiation processes, as an important effector.
The Notch receptor is activated upon engagement by a membrane‐bound ligand, such as Serrate or Delta, on an adjacent cell (Rebay et al, 1991). Ligand binding triggers proteolytic cleavage of Notch, which is mediated in part by γ‐secretase (De Strooper et al, 1999). The cleaved intracellular domain (ICD) of Notch translocates to the nucleus, and upon binding to cofactors such as p300/CBP, Suppressor of Hairless (Su(H)), and/or mastermind (MAML1), transactivates Notch‐responsive genes such as HEY and HES (Fortini, 2009).
Collaboration between HIFα and Notch occurs at the intersection between the oxygen‐sensing and developmental pathways. Notch has been reported to potentiate HIFα‐mediated transactivation of hypoxia‐inducible genes in the context of both tumourigenesis and cellular differentiation (Sahlgren et al, 2008; Zheng et al, 2008). Conversely, HIFα has been demonstrated to enhance Notch signalling through several distinct mechanisms. HIF1α has been shown to stabilize Notch ICD in both normoxia and hypoxia in a manner that is independent of its transcriptional activity, although the precise mechanism by which this occurs remains unclear (Gustafsson et al, 2005). Additionally, HIF1α transactivation of anterior pharynx‐defective 1A (APH‐1A), a component of the γ‐secretase complex, has been reported to lead to an increase in intracellular levels of Notch ICD (Wang et al, 2006). HIF1α interaction with Notch at the HEY2 and HES1 promoters has also been reported, and this association augments Notch‐mediated transactivation under hypoxia in a manner that appears to depend upon a functional CTAD but does not require DNA binding by HIF1α (Gustafsson et al, 2005; Chen et al, 2010). Interestingly, Notch that is impaired in its ability to bind p300/CBP still retains the ability to transactivate target promoters under hypoxia, suggesting that the function of HIF1α in the Notch transcriptional complex is to recruit p300/CBP (Gustafsson et al, 2005). This is reminiscent of the aforementioned TH17 cell differentiation model, whereby HIF1α recruits p300 to the RORγt transcriptional complex, but does not bind the IL‐17 promoter directly to induce transcription. In addition, FIH, another important component of the oxygen‐sensing pathway, was reported to negatively regulate the transcriptional activity of Notch, although this effect appeared to be independent of FIH catalytic activity (Coleman et al, 2007; Zheng et al, 2008). Interestingly, FIH has a significantly higher affinity for Notch than for HIF1α (Wilkins et al, 2009); thus, sequestration of intracellular FIH by Notch may mediate transcriptional de‐repression of HIF1α (Zheng et al, 2008), thereby delineating another mechanism by which Notch might augment the adaptive response to hypoxia.
Crystal cell development requires direct stabilization of endosomal Notch by HIFα. A recent report from Mukherjee et al (2011) proffers an entirely novel context for HIFα and Notch interaction, with each factor functioning in a starkly non‐canonical manner (Figure 3D). Development and differentiation of crystal cells, a Drosophila blood cell subtype that is involved in wound healing, was shown to depend upon stabilization of Notch by HIFα within early endocytic vesicles. The Drosophila HIFα orthologue, Sima, promotes crystal cell expansion in the lymph gland, and was demonstrated to be crucial for the survival of committed crystal cell precursors. Similarly, Notch was shown to be required for initial commitment to the crystal cell lineage as well as expansion, maintenance and survival of committed crystal cell precursors. In addition, cleavage of the Notch ICD was vital for crystal cell survival, indicating an important role for Notch signalling in crystal cell development.
Sima overexpression stabilized Notch protein levels, a finding that is in keeping with previous observations regarding positive regulation of Notch signalling by HIFα. However, rather than observing an increase in nuclear levels of Notch ICD, Mukherjee et al (2011) found that the majority of intracellular Notch colocalized with Sima in early endocytic vesicles within crystal cells. Live endocytic trafficking assays performed on wild‐type Drosophila lymph glands revealed that internalization of full‐length Notch (Notchfl) from the plasma membrane preceded its rapid degradation in all non‐crystal cell types. Intriguingly, activation of Notch in committed crystal cell precursors did not appear to require ligand engagement by either Serrate or Delta, suggesting that Sima‐mediated activation of Notch was occurring in a non‐canonical, ligand‐independent manner. This mechanism seems sensible from an evolutionary standpoint, as Notch signalling in crystal cells appears to be vital for their differentiation and survival, but given that they circulate freely in the bloodstream it is not feasible for them to continuously engage the Notch ligands of neighbouring cells.
The function of Sima in crystal cells did not appear to include transactivation of hypoxia‐responsive genes. Overexpression of the Drosophila HIFβ orthologue Tango failed to induce expression of a Sima/Tango‐responsive reporter plasmid, and hypoxic treatment only augmented crystal cell expansion. It should be noted, however, that following Notch cleavage and nuclear translocation of ICD there is a possible transcriptional role for Sima in enhancing Notch‐mediated transactivation of Notch‐target genes through recruitment of, and association with, other cofactors in the Notch transcriptional complex as described previously (Gustafsson et al, 2005; Chen et al, 2010).
Another non‐canonical aspect of Sima function in this report involves its unusually high degree of activity in normoxic conditions. Mukherjee et al (2011) demonstrate elevated synthesis of nitric oxide synthase‐1 (NOS1) in crystal cells, which appeared necessary for Sima stabilization under normal oxygen tension. This unique feature of crystal cells likely explains why internalized Notch is rapidly degraded in other cell types—intracellular Sima levels may not be sufficiently high under normoxia to stabilize Notch in early endosomes.
Augmentation of Notch signalling by HIF1α was previously shown to require both hypoxic conditions and ligand (Serrate) engagement (Gustafsson et al, 2005). The report by Mukherjee et al (2011) suggests that the sole function of HIFα in crystal cells is augmentation of Notch signalling, and HIFα appears to accomplish this in large part by stabilizing Notch in a ligand‐independent manner in a normoxic environment (Mukherjee et al, 2011). Indeed, the proposed role of HIFα in this context represents an entirely new mechanism by which it regulates differentiation and development. Future studies aimed at further characterizing this mechanism should attempt to do so in other species. Drosophila crystal cells are reportedly most similar to mammalian myeloid cells, thus it would be interesting to see if stabilization of Notch by HIFα occurs in the endosomes of myeloid precursors. Additional roles for HIFα in the stabilization of endosomal Notchfl are also worth investigating: Mukherjee et al (2011) report that overexpression of Rab5 suppresses Sima/Notch‐mediated crystal cell expansion by increasing endocytic turnover of Notchfl. HIFα has been previously demonstrated to downregulate the Rab5 effector rapabtin‐5 (Wang et al, 2009), thereby prolonging the endocytic pathway. However, it is unknown whether HIFα‐mediated attenuation of early endosome fusion serves as a secondary mode of Notchfl stabilization.
One important question left in the wake of this study pertains to the apparent lack of HIFα transcriptional activity at HIF‐target genes in crystal cells, particularly under hypoxia. Crystal cells are involved in wound healing and mediating the humoral immune response (Meister, 2004; Bidla et al, 2007), and are therefore likely to be presented with significant hypoxic challenge in necrotic or inflamed tissues. Given that HIFα is the key transcriptional regulator of the cellular adaptive response to hypoxia, how do crystal cells overcome the metabolic stresses that inevitably occur in low oxygen tension? One possibility is that they are not meant to. Crystal cells rupture in response to injury or inflammation and release significant amounts of prophenoloxidase, which is crucial for clotting and defence against pathogens (Meister, 2004; Tang et al, 2006; Bidla et al, 2007). Perhaps the lack of an adaptive response to hypoxic challenge is an evolutionarily‐developed strategy to ensure vulnerable crystal cells rupture in inflammatory or necrotic conditions. If this is indeed the case, then the mechanism(s) responsible for attenuating the transactivation of HIFα‐responsive genes in Drosophila crystal cells under hypoxia warrant future study.
Concluding remarks
Recent work regarding the oxygen‐sensing pathway and its predominant effector, HIFα, has expanded our current understanding of HIFα regulation and transactivation, the role of HIFα and underlying genetic aberrations in health and disease, and the multifaceted nature of HIFα in a variety of physiological and pathophysiological contexts. Unexpected new players have entered into the 'canonical' model of HIFα regulation and transactivation, and have shown demonstrable influence on the hypoxic response by modulating PHD function or enhancing HIFα transcriptional activity. Furthermore, the traditional dogma regarding the function of HIFα as a hypoxia‐inducible, DNA‐binding transcription factor is challenged by new reports that demonstrate its capacity as a transcriptional coactivator in developmental and inflammatory contexts, a director for proteasome‐mediated degradation, and a stabilizing force for internalized receptors within the endosomes. These notable studies not only provide additional research avenues, but also questions, the answers to which will undoubtedly enhance our appreciation for the increasing complexity and biological relevance of the HIF pathway in humans and other organisms.
Conflict of Interest
The authors declare that they have no conflict of interest.
This work was supported by funds from the Canadian Institutes of Health Research and the Canadian Cancer Society.
- Copyright © 2012 European Molecular Biology Organization


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Google just announced a pretty major update to Gmail, and you should now be able to give it a go in the browser, if you so desire. As our pal Paul O'Brien from MoDaCopoints out, all you have to do to enable the new tabbed organization is go to the gear icon on the right of the screen and choose "Configure inbox." (If you're using a wrapper like Mailplane, you might need to do this from a traditional browser first, then restart the app.)
Next you'll be asked which tabs you want to use -- primary, social and promotions are checked by default; updates and forums are other options. Once enabled, you'll find e-mails sorted among the tabs. To move e-mails between tabs, just click and drag. (You'll also be asked if you want to continue to get that particular kind of e-mail in the new tab.)
Still no sign of the new Gmail app for Android, which also will bring a new sorting experience.


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Chemically modified inhibitors of the COX-2 enzyme relieve anxiety behaviors in mice by activating natural "endocannabinoids" without gastrointestinal side effects, Vanderbilt University scientists will report next week.
Endocannabinoids are natural signaling molecules that activate cannabinoid receptors in the brain, the same receptors turned on by the active ingredient in marijuana.
These receptors are also found in the gastrointestinal system and elsewhere in the body, and there is evidence that they play a role in wide range of physiological and pathological processes, in addition to modulating stress and anxiety.
If the "substrate-selective" COX-2 inhibitors developed at Vanderbilt also work in humans without side effects, they could represent a new approach to treating mood and anxiety disorders, the researchers conclude in a paper to be posted online Sunday in the journal Nature Neuroscience.
Clinical trials of some of these potential drugs could begin in the next several years, said Lawrence Marnett, Ph.D., director of the Vanderbilt Institute of Chemical Biology and the paper's co-senior author with Sachin Patel, M.D., Ph.D.
The Vanderbilt scientists are pursuing other potential applications of activating endocannabinoids by substrate-selective COX-2 inhibition, including relieving pain, treating movement disorders, and possibly preventing colon cancer.
"The door is really wide open," said Patel, assistant professor of Psychiatry and of Molecular Physiology & Biophysics. "We've just scratched the surface."


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Improving genetics | International collaborators sharing information
Canada plays a major role in the international effort to catalogue and share the DNA sequences of cattle, says a researcher.
A database of 234 dairy cattle genomes was recently used to identify genetic mutations that hamper dairy production. Similar collaborations are ongoing in the beef sector. A recently published paper highlighted the effort.
The work was part of the 1,000 Bulls Project in which researchers are sharing the DNA sequences of key ancestor animals from around the globe.
The results will help with genomic predictions, potentially providing a test that could identify an undesirable genetic trait in a young animal.
Producers could then select against the trait and increase the speed of genetic herd improvements.
"If every country was having to sequence these same animals it would be extremely expensive. The cost of sequencing has come down significantly in the last five to 10 years, but it's still an expensive thing to do," said Mary De Pauw, project manager for the Canadian Cattle Genome Project. "It's a wonderful opportunity."
In the study published in Nature Genetics, 234 sequences from Holstein-Fresian, Jersey and Fleckvieh dairy cattle were used to identify mutations that lead to embryonic death and chondrodysplasia, which hinders the development of long bones, causing a form of dwarfism.
"They've been able to find the area in the genome where this mutation exists and so this will allow us in the future so we can do a quick test… then we can select against animals who have this mutation so that we don't continue to breed this into our populations," said De Pauw.
The Canadian Cattle Genome Project is focused on prolific bulls that have influenced the herds of all of the major Canadian beef breeds.
"We've found bulls that when you look at the number of progeny they have in our current herds, they have like over 100,000 progeny, because they were used so significantly for breeding because of their positive attributes," she said.
"We've done a huge analysis of the pedigree for all of our breeds and that's how we've chosen which animals to sequence."
More than 300 animals have been sequenced because of the Canadian effort, using DNA samples that go back to 1900. In return, De Pauw said researchers can access sequences of more than 1,100 animals.
"A lot of the early data was primarily Holstein, but the beef breeds are going to be a major component of that," she said.
Genomics is more advanced and widely employed in the dairy industry because of its focus on a single breed, Holstein, and single trait, milk production, said De Pauw.
"Genotyping is a huge thing in the dairy industry. Everybody does it and they've had significant gains by using it," she said.
"But the beef breeds, because the beef industry is a little bit more segmented, it's taken a little bit longer to get them up and going and it's also a little bit more challenging because you're dealing with several breeds. You're not just dealing with one breed."
De Pauw said genomics could also be used to select animals for carcass traits and feed efficiency.
"I think the gains will be a little bit harder in the beef industry, but we're quite confident we can make significant strides with genomics," she said.


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TeamViewer 13.2.5287 Crack Download Free Latest Here
TeamViewer 13 is a simple and fast solution for remote control, desktop sharing and file transfer that works behind any firewall and NAT proxy. To connect to another computer just run TeamViewer on both machines without the need of an installation procedure. With the


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I loved the "Disappearing Act" challenge from last week. I think this sketch is going to help me use lots and lots of paper from my stash.
This is what I made yesterday
One 12x12 inch piece of paper and I have 6 cards.
I used a new embossing folder from The Paper Studio "Curl Ribbon"
The sentiment stamp is from Stampabilities.
The digital stamp came from Michelle Perkett Studio and can be purchased here. This is the first time I've used it and I just love it.


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Customizing the Rating Scale Element
The rating scale question type is very similar to a normal rating (single-select) question, but adds in some animation and images that can be more appealing than the usual radio buttons in a default grid question.
1: Underlying Question Architecture
Underneath a rating scale question is... you guessed it, a single-select question.
<radio label="q1"> <title>What do you think of these actors' portrayal of James Bond?</title> <row label="r1">Sean Connery</row> <row label="r2">George Lazenby</row> <row label="r3">Roger Moore</row> <row label="r4">Timothy Dalton</row> <row label="r5">Pierce Brosnan</row> <row label="r6">Daniel Craig</row> <col label="c1">Great as Bond</col> <col label="c2">4</col> <col label="c3">3</col> <col label="c4">2</col> <col label="c5">Not So Bond</col> <col label="c6">Who?</col> </radio>
2: Converting to Rating Scale
To convert the above question to a rating scale question, simply add
uses="ratingscale.5" to the question tag, like so:
<radio label="q1" uses="ratingscale.5">
Only the question tag is shown here because no other changes were made. The rows and cols are all the same.
3: Customization
From here onward, we'll be customizing the question away from the default look shown above. To do this, we'll be working in the XML with customizable variables that are unique to the rating scale question type. In the examples, only the question tag will be shown as we're not going to be doing anything to the
<row> or
<col> tags. If you add a rating scale question using Builder, you'll notice that there are several new XML variables put into the question tag. You only need to include the ones you want to customize.
Here's a brief description of each, but first some terminology when referencing these variables:
Bubble: We call the draggable/animated image that moves across into the answer location a bubble
Radio: On a default single-select question, the form that would appear in the answer grid is called a radio button; to signify it's location/replacement, the word radio gets used frequently in these configurable variables to represent either the radio button form or the grid cell that usually contains it
- Configure styling of the draggable icon before a selection has been made
- Type: CSS
- Example: ratingscale:bubble_inner_starting_css="color: pink; font-size: 32px;" changes the color and size of the bubble
- Configure styling of the draggable icon after a selection has been made
- Type: CSS
- Example: ratingscale:bubble_inner_answered_css="color: rgba(0,0,0,0.5)" changes the color of the bubble post-selection
- Specify the pixel width and height of the draggable image (bubble)
- Expects image to be square
- Type: integer
- Example:
ratingscale:bubble_size="40"sets the image to 40px by 40px
- Specify an image to use as the bubble
- Type: URL
- Example:
- Configure styling of bubble
- This will be overridden by the more specific rules below, but allows you to set some global bubble properties (like maybe a border radius)
- Useful when you don't want to use images and would rather use a styled CSS div instead
- Recommend NOT using in combination with images
- Type: CSS
- Example:
ratingscale:bubble_css="background: #aaa; border: 1px solid #777"would set the bubble to have an unsightly grey circle with a darker grey border
- Configure styling of bubble when bubble is sitting in the starting zone
- Overrides the
- Recommend NOT using in combination with images
- Type: CSS
- Example:
ratingscale:bubble_starting_css="border-color: red"would change the bubbles border to red when it's in the starting position
- Configure CSS styling of bubble when bubble is on the answer grid
- Overrides the
- Recommend NOT using in combination with images
- Type: CSS
- Example:
ratingscale:bubble_answered_css="border-color: green"would change the bubbles border to green once it's moved onto the grid
- Configure styling of bubble when bubble is disabled
- Overrides the
- Recommend NOT using in combination with images
- Type: CSS
- Example:
ratingscale:bubble_disabled_css="background: #ddd;"would set the background to light grey when the bubble is disabled
- Hides the radio buttons in the grid
- Only used if
- Recommend NEVER showing the radio forms (kinda defeats the purpose, right?)
- Type: Bool (1 = HIDE, 0 = SHOW); default = SHOW (only when ratingscale:radio_img="")
- Example:
ratingscale:hide_radio="1"would hide an otherwise shown radio button
- Configure styling for the radio image (dotted circle)
- Type: CSS
- Example:
ratingscale:radio_inner_css="display: none;"hides the dotted circle
- Specify the pixel width and height of the radio button image
- Expects image to be square
- If using an image sprite then the states should be squares, for example a 20x40 image sprite where each state is a 20x20, size would be 20
- Type: integer
- Example:
ratingscale:radio_img_size="40"would set the image size to be 40px by 40px
- Replaces the radio button in the grid with an image when the radio button is unselected
- Type: URL
- Example:
- Replaces the radio button with an image when the radio button is selected
- Set this value to empty if you do not want an image behind the "bubble" image/CSS
- Type: URL
- Example:
- CSS styling of the grid cell
- Note that this applies to the interior table cells and does not affect the actual table/grid; to do that, you need to configure the survey theme
- Mostly useful if you simply want to change the background color on selected/unselected cells or use a background image
- Type: CSS
- Example:
ratingscale:radio_css="background: black"to set the grid cells to black
- CSS styling of the grid cell when that answer is selected
- As with ratingscale:radio_css, this applies to the interior table cells and does not affect the actual table/grid; to do that, you need to configure the survey theme
- Mostly useful if you simply want to change the background color on selected/unselected cells or use a background image
- Type: CSS
- Example:
ratingscale:radio_selected_css="background: #f80"to set the grid cells to orange when selected
- Toggles a transition when using an image for the radio buttons, assuming that image has 2 states: selected and unselected
- Only applies if using a replacement image for the radio buttons; indicates that the image has two states: selected and unselected; adds CSS to transition between the two states within the single image
- Type: Bool (1 = ON, 0 = OFF)
- Example:
ratingscale:radio_img_transition="0"disables any transition
- CSS transition effect used when shifting between states in the replacement radio button images
- Assumes
- CSS3 transitions are worthy of much documentation on their own. If you're into it, here's an article you can check out about these.
- Type: String
4: Customization Examples
The rating scale question type allows a lot of customization and there are any number of directions you can take it. Let us play.
Example 1:
<radio optional="1" label="q3" uses="ratingscale.5" ratingscale:bubble_size="40" ratingscale:bubble_img="pushpin.png" ratingscale:radio_img="bg-dot.png" ratingscale:radio_selected_img="">
In this example, we're replacing the default button/marker with a pushpin. The pushpin graphic is an image sprite in that it's a single image that contains both the normal and selected states in a single image and we only show the applicable portion. For this one, the graphic is 80px wide and 40 pixels tall; each state is 40px wide. Here's what it looks like:
We use
ratingscale:bubble_size="40" to set the square dimensions of the image, and
ratingscale:bubble_img="pushpin.png" to set the image. For the cells, we want a placeholder image (orange dot, in this case) to appear when that answer is not selected and no image to appear when it is (because it would interfere with our pushpin graphic), so we do that with
ratingscale:radio_img="bg-dot.png" and
ratingscale:radio_selected_img="" respectively.
Example 2:
<radio label="q4" uses="ratingscale.5" ratingscale:bubble_size="30" ratingscale:bubble_css="background: #87e0fd; background: -moz-radial-gradient(center, ellipse cover, #87e0fd 1%, #047ea3 71%, #046887 100%); background: -webkit-gradient(radial, center center, 0px, center center, 100%, color-stop(1%,#87e0fd), color-stop(71%,#047ea3), color-stop(100%,#046887)); background: -webkit-radial-gradient(center, ellipse cover, #87e0fd 1%,#047ea3 71%,#046887 100%); background: -o-radial-gradient(center, ellipse cover, #87e0fd 1%,#047ea3 71%,#046887 100%); background: -ms-radial-gradient(center, ellipse cover, #87e0fd 1%,#047ea3 71%,#046887 100%); background: radial-gradient(ellipse at center, #87e0fd 1%,#047ea3 71%,#046887 100%); filter: progid:DXImageTransform.Microsoft.gradient( startColorstr='#87e0fd', endColorstr='#046887',GradientType=1 ); border-radius: 14px; box-shadow: 0 1px 2px 0 rgba(0,0,0,0.4)" ratingscale:bubble_inner_starting_css="display: none;" ratingscale:bubble_inner_answered_css="display: none;" ratingscale:radio_inner_css="display: none;" ratingscale:radio_css="background: #efefef;" ratingscale:radio_selected_css="background: #fff;" ratingscale:hide_radio="1">
This example doesn't use any images, instead it uses CSS to create the bubble. Let's run through the customization, since it's quite a bit more intense than the previous example.
ratingscale:bubble_size="30"Sets the size of the bubble to 30px
ratingscale:bubble_css="background:This line sets the gradient effect on the bubble. There's a lot of CSS happening here, but this is actually pretty easy to do with help from this excellent online CSS gradient tool: http://www.colorzilla.com/gradient-editor/
ratingscale:bubble_inner_starting_css="display: none;"Removes the default arrow icon so that the
bubble_cssattribute can properly be shown.
ratingscale:bubble_inner_answered_css="display: none;"Removes the default arrow icon so that the
bubble_cssattribute can properly be shown when an answer has been made.
ratingscale:radio_inner_css="display: none;"Removes the default image that appears in the answer grid
ratingscale:radio_css="background: #efefef;"Sets a light grey background on non-answered cells
ratingscale:radio_selected_css="background: #fff;"Sets the background to white for selected answer cells
ratingscale:hide_radio="1"Hides the default radio form; this is necessary when setting
5: Some Considerations
5.1: Mobile Support
This question type will work on mobile devices, but please be aware that more than a few columns will cause horizontal scrolling which may not be obvious on a mobile device. If that happens, you may see an increased number of answers in the first couple columns as respondents may not realize there are more columns that appear off-screen.
5.2: CSS Styling
There are many cool things that can be done with CSS, but not all browsers will apply it equally. A good example is the rule that set the background gradient above. If you dissect that rule, you'll see that each browser needs its own customization, and even then, support may be limited to modern browsers. A key principle is to use advanced CSS to enhance the experience, but don't rely on it for core functionality.
6: Reference
- Demo Survey: Check out the examples above, live, in action, in a real survey: http://v2.decipherinc.com/survey/selfserve/9d3/121195?list=0
- CSS Help: Here are 3 really useful CSS resources
- CSS Cheat Sheet: http://www.cheatography.com/davechild/cheat-sheets/css2/
- CSS3 Generator: http://css3generator.com
- CSS Gradients: http://www.colorzilla.com/gradient-editor/


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So I just received the "official" Fulbright Grant Packet which looks like a contract (I guess it IS, actually). Once copies of this material travel to the head-honchos at the Administration Building of Northwest Missouri State University we've cleared another hurdle.
Now we gather all the paperwork like birth certificates, marriage license, Grant Packet, etc. for notarization followed by sending to Jefferson City, MO, for apostille. After that we've got to: 1) continue to learn Latvian, 2) find an apartment, 3) look into overseas insurance, 4) figure out what & how much/little we should take, 5) work on travel arrangements, 6) practice trumpet for a February trumpet recital, and much, much more.
Anyway, we're happy for the opportunity and I know that this will be an experience that "the ladies" and I will remember for the rest of our lives...kind of like those canoe trips down the Missouri River with my old man, except MUCH safer!


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Rooting your Android phone is a term that you are bound to across at some point or another while searching on how to optimize your Android device. With millions of new Android owners each month we decided to give a basic introduction into the world of rooting and to let you decide if it's something you'd like to do.
What is Rooting?
Rooting is the process by which you regain administrative access to your phone. Even though Android is an open source operating system you still don't have full "root access" to do what you please. Back when the iPhone launched in 2007 the hardcore techies quickly realized the true potential of the device, and the cruel software limitations that Apple had sealed it with. What became 'Jailbreaking' on iPhone was quickly translated to other platforms as well, and when the world saw the first Android back in 2008, the term "Rooting" was born.
Why Root your Android Phone/Tablet?
The main reason people root their Android device for freedom and control, when you root your Android phone or tablet you gain full control over your system and can tweak it to your liking.
Improved Performance: You can speed up your Android device by relocating your phones cache thus allowing you to save phone memory and have a faster phone. There are applications available in the Android Market that will allow you to overclock your device to make it go as fast as you dare.
Alter System Files: You can replace many parts of the "Android Core" which include the ability to add new themes, edit the core apps (maps, calendar, clock, etc), change the recovery & boot images, add linux binaries.
More Application Choics: You will be able to install apps that are only compatible with rooted phones, some of these apps include an app that will allow you to take a screenshot on your phone, overclock your device and tether apps.
Install applications to your SD CARD: One of the most talked-about feature (or disadvantage) of any Android device is the limitation where you can install applications only in the phone's internal memory and not the SD card. While Google may reason that SD cards are slower in general and cannot run apps as effectively as internal memory, fact of the matter is that most Android devices do not come with massive internal storage spaces, and hence greatly limit the number of applications that can be installed at a time. With rooted devices, you can use Apps2SD, which will copy ALL your applications to a ext2/3/4 formatted SD card an will also store future builds in card. Freedom to choose!
Latest Android OS (Operating System): With many carriers holding back the updates to the latest Android operating system, rooting your device will give you the option to install any current and future OS's by installing custom-tailored ROMs.
WI-FI and Bluetooth Tethering: After having rooted your device, you can also use WiFi or Bluetooth tether to share your cellular data connection with your laptop or PC. The application works with ad hoc connections and will get you up and running online on your laptop in no time. Similarly, tethering can also be achieved over a Bluetooth connection.
How to Root Your Phone/Tablet?
To Root your phone you will have to download an application from the internet, the most popular apps areSuperOneClick, Z4Root and Universal AndRoot. The procedure for rooting an Android device varies from device to device as such there are many different device specific guides. Website AddictiveTips has put together the largest guide for rooting phones that we found. Some other good resources for finding rooting guides include the XDA Developers Forums & this topic at AndroidForums . In most cases the Rooting procedure is as easy as a couple of clicks, of course there are nightmare stories and people having difficulties, so read on about the risks of rooting.
What are the risks of Rooting?
Rooting your phone does come with some risks, the most notable risk is that you will void any warranty that you have on your device. However you may be able to find the stock rom for your device in which case you can reverse the rooting and make your phone stock again. You may have difficulty finding the stock ROM for your device, it all depends on your device, but it's something worth finding before you do root.
Other then the voiding your warranty, there isn't that much risk involved. There have been some users onAndroid forums that have run into problems, bricking your device is a possibility if a freak accident occurs while flashing archives. You should however not run into any problems, most users report the process as being a painless easy process.
Rooting your device can be a fun process, giving you the freedom and control to take your phone to a whole new level of function. We do recommend that you do your due diligence to ensure the rooting process goes as smoothly as possible. What are your thoughts on rooting? Was it an easy experience? Do you recommend it to others? We'd love to hear your thoughts.
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- Need for speed shift full for Android Snapdragon Devices


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Customer Reviews
All reviews are from people who have redeemed deals with this merchant.
What You'll Get
All the coolest people from history have ridden horses: knights, cowboys, the top portions of centaurs. Witness history with this Groupon.
Choose Between Two Options
- $10 for two tickets to the Scottsdale Arabian Horse Show (a $20 value)
- $19 for four tickets to the Scottsdale Arabian Horse Show (a $40 value)
The 58-year-old show draws spectators with equine competitions featuring more than 2,500 horses, along with educational seminars, gala events, international cuisine, and more than 300 vendors of jewelry, art, and apparel. The event runs from Thursday, February 14, to Sunday, February 24, at the WestWorld grounds.
The Fine Print
Promotional value expires Feb 25, 2013. Amount paid never expires. Limit 10 per person. Valid only for option purchased. Merchant is solely responsible to purchasers for the care and quality of the advertised goods and services.
About Arabian Horse Association of Arizona
In 1955, 50 horses and their handlers gathered on the grounds of the Arizona Biltmore Hotel for the first Scottsdale Arabian Horse Show. Over the next 58 years, the event moved twice—first to Paradise Park, then to WestWorld—and grew to nearly 2,500 participants. Today, the 450 members of the Arabian Horse Association of Arizona, which was incorporated the same year as the horse show, help to oversee the annual flagship event. Many of the members also serve as competition judges, work to foster youth participation in the equestrian arts, and raise awareness about which tiny hats look best on thoroughbreds.
The annual Scottsdale Arabian Horse Show draws competitors from local Arizona farms and across the country. The showcase gathers thousands of horse enthusiasts for seminars, competitions, galas, international cuisine, and more than 300 vendors. Though the Arabian Horse Association of Arizona holds the show as its main event, it also oversees the Arabian Breeder Finals, a halter and breeding showcase; the annual qualifying America Cup Championship; and yearly endurance rides and stallion auctions.