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what is your favorite healthy food?
Mama to 3 amazing boys! I AM BLESSED! Making adoarble clothes for cloth diapered babes. Check out my store: www.etsy.com/shop/FluffyTushClothing
Taco soup or taco salad- it's easy, everyone likes it, and it's pretty healthy if you don't drown it in cheese/ sour cream :)
Angela married to DH Jeremy 9/22/07
DS Jonathan 10/08/00
Rainbow Baby Jonas arrived on 6/3/13!!
Something super easy, filling, and healthy that I eat often are Progresso soups--they have a ton of soups that are lower in calories and high in fiber.
Mama to 3 amazing boys! I AM BLESSED! Making adoarble clothes for cloth diapered babes. Check out my store: www.etsy.com/shop/FluffyTushClothing
Love pomegranates for snacking!
Chili w/ lots of beans, tomatoes, and lean ground turkey; my broth is V8 seasoned with chili spices, garlic, and a little beef buillion.
I do a wrap w/ a whole wheat tortilla w/ black beans, plain yogurt (tastes like light sour cream), salsa and a little shredded cheese and lime juice. nursegina45 has me wanting to add an avocado to that, yummy!
I do a lot of homemade soups and stews w/ lots of veggies. I use Knorr concentrated stock b/c I need to be quicker than making my own, but I don't add butter or any other fat besides what's in the meat.
Chicken cacciatore, esp. if you use breasts (white meat)--lots of tomatoes and sauce with very little olive oil, served w/ salad and whole grain spaghetti.
Usually whatever we are having for dinner I try to make sure that we have at least one veggy. Canned, fresh or frozen. Lately I've been on a fruit kick, so we've been having fruit as a side too. My guys are super sweet & not picky so no one complains when mom makes weird stuff for dinner!
Eggs are quick, cheap & easy. Also quesadillas- just go easy on the cheese. You can load each bite up with salsa for extra flavor. I also like chicken grilled on our Foreman grill- super fast & easy. The freezer/steamer bags of rice & veggys are life savers sometimes. Throw some chicken in the oven or on the Foreman a bag of rice & then veggys in the microwave and you just rocked some healthy dinner with minimal effort.
Hummus.. It's so healthy and low fat. I have even been making my own homemade granola.


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Bayou Bakery, in Arlington, VA, transports its customers to New Orleans.
The Bakeshop, in Arlington, VA, tweaks everyone's classic bakery favorites.
Darryl hits a hip nightspot to sample some seriously boozy cupcakes!
Doughboy Bake Shop gives sweet favorites a bit of an unexpected twist.
Nadia G's No-Bake Cream Cheese PB Pie delivers salty and sweet in one bite.
Buzz Bakery reinvents classic childhood sweets with grownup twists.
Sara Moulton of Sara's Secrets shares a grilled dessert recipe.
Brooklyn Farmacy pours out a funky blend of nostalgic soda-fountain treats.
Kelsey puts more fun into dessert with her fresh Berry Pie Pops on a stick.
The entree and dessert are hits, and the chefs celebrate a job well done.
Get Cooking Channel on your TV.

subset: commoncrawl


text:
|Found 42 Recipes
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Giddy's Cajun Gumboby kendallinn As a true southern woman, I identify with all southern cuisines! Tex-M..
Creole Seafood Gumboby Maggie Perkins I'm a bit of a gypsy. Went to nine schools in 12 years. Three colleges..
Clear onion soupby jpr8682 also might want to add salt to taste it all depends on what you like
Grandma Jenkins' Chicken and Dumplingsby CenturyLink My Grandma Jenkins has been making her dumplings since she was a young..
Tom Kha Gaiby daringgourmet Thai food is amazing! Tom Kha Gai, the highly popular Thai chicken so..
Kale and Roasted Sweet Potato Soup with Chicken Sausageby monal This is a light and healthy soup with a lot of greens! Feel free to om..
My Roasted Turkey Barley Soupby imstuffed My husbands aunt gave me the idea of making turkey soup years ago and ..
Wonton Soup Ala College Inn Chicken Brothby nanalee Seeing the Foodie's homemade wonton soup recipe posted today reminded ..
Easy Homemade Veggie Brothby sproulie Eating right, in large part, is simply just opting for a homemade vers..


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Armed teachers can
help protect children
As we approach the one-year anniversary of the Sandy Hook Elementary School shooting, I believe it is time to re-visit the idea of arming our teachers to help protect the children of this country. How many more tragedies like this will occur until we are willing to take a stand?
Many experts agree that arming our teachers will save lives. Russell Gardner, a local gun expert and hunter education instructor for 22 years, stated, "Teachers should be trained to use firearms to protect themselves and our students. Without it, they are sitting ducks." One good reason to arm our teachers is the time involved in waiting for police to arrive when a school is threatened by a gunman. Steve Siebold of the Washington Times stated, "Every second counts in a school shooting. Calling 911 and waiting for the police to arrive isn't good enough." Milwaukee County Sheriff David Clark Jr. has said "Simply calling 911 and waiting is no longer your best option."
Arming teachers will also serve as a deterrent to school shootings. Siebold mentions that "a gunman is less likely to target a school where it's staff can defend it and fight back." It makes sense that a gunman would select a more vulnerable target and avoid schools where the staff is armed.
In summary, arming teachers is a great idea and will save lives. If we trust our teachers to educate our children, shouldn't we also trust them to protect them?
Nick Seacat Freshman, Las Cruces High School Las Cruces
Soroptimist thanks
for turkey trot
On behalf of Soroptimist International of Las Cruces, we want to thank everyone who donated, sponsored, and participated in our Second Annual Soroptimist International Turkey Trot. Our Turkey Trot would not have been possible without our sponsors.
PLATINUM SPONSORS: Carlos Parra, Dental Associates, PA, The Las Cruces Bulletin.
GOLD Sponsors: Walgreens, Thrivent Financial, Pik Quik, Caliches's Frozen Custard, Hub International Insurance, Sonic, and The Print Guys.
SILVER SPONSORS: Good Samaritan Las Cruces Village, Las Cruces Oral and Facial Surgery, Massage Envy, Sunspot solar Energy, Zia Engineering and Environmental Consultants, and Citizens Bank.
We received dozens of individual donations, and volunteers that helped us make our event a success, and further our mission of helping women and girls in our community. Our club is grateful for the volunteers from Oñate High School football team, and their parents. This group has volunteered at both of our Turkey Trot Runs.
Since the club's inception, Soroptimist International has raised over $260,000 which has helped organizations in Las Cruces. Girls Can, Gospel Rescue Mission, Dress the Child, Coats for Kids, and Casa de Peregrinos, National Child Protection Training Center-Southwest Region, and women returning to school are a few of the causes that Soroptimists has recently supported.
We hope to see you all again at our 3rd Annual Soroptimist Turkey Trot that will take place on Nov. 22, 2014.
Marcia Starich Soroptimist International Las Cruces
Hooks cites study
few have heard of
I did a double take when I saw Mr. Hooks 12/5 column referring to a 98 percent consensus on global warming. This was partly because I no longer feel the need to read this section of the Sun-News because the columnists seem to be so predictable - left or right, and partly because he cites a figure from a study of which I've never heard anyone speak.
Given Mr. Hooks' predictable skepticism of all things climate change, I'm not surprised he questions the methodology used in a study nobody else cites. Though open to interpretation, the questions were nonetheless asked of people whose positions should be taken seriously-earth scientists.
To further show the questionable nature of the 98 percent figure, he refers to a study saying only 93 percent of "us" believe we landed on the moon. The opinions of the masses should be taken as proof opinions of scientists should be taken with a grain of salt?
The number I have most often heard used to evidence support of scientists that the earth is warming and we are responsible for it is 97 percent (Cook, et al). This figure was based on the analysis of 11,944 abstracts from peer reviewed scientific journals from 1991-2011. Key word searches showed "among abstracts expressing a position on AGW, 97.1 percent endorsed the consensus position that humans are causing global warming." According to Cook himself, these abstracts were only from articles by researchers "actively conducting climate research."
Whichever study one chooses to cite, the numbers are impressive. I am not a legal scholar, but if close to 100 percent of legal scholars agree on a particular legal issue, I would consider it so resolved. I am not a scientist, so I defer to the experts who seem to unquestionably agree on this issue.
L.A. Louick Las Cruces


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Threads and Synchronization
How can threads communicate with each other?
Prateek Kumar Singh
Joined: Jan 21, 2011
Posts: 29
I like...
Apr 11, 2013 11:58:17
Hi Guys,
The wait(), notify(), and notifyAll() methods are used to provide an efficient way for threads to communicate with
each other. This communication solves the 'consumer-producer problem'. This problem occurs when the
producer thread is completing work that the other thread (consumer thread) will use.
Example: If you imagine an application in which one thread (the producer) writes data to a file while a second
thread (the consumer) reads data from the same file. In this example the concurrent threads share the same
resource file. Because these threads share the common resource file they should be synchronized. Also these
two threads should communicate with each other because the consumer thread, which reads the file, should wait
until the producer thread, which writes data to the file and notifies the consumer thread that it has completed its
writing operation.
Let's look at a sample code where count is a shared resource. The consumer thread will wait inside the
consume() method on the producer thread, until the producer thread increments the count inside the produce()
method and subsequently notifies the consumer thread. Once it has been notified, the consumer thread waiting
inside the consume() method will give up its waiting state and completes its method by consuming the count (i.e.
decrementing the count).
Class ConsumerProducer { private int count; public synchronized void consume(){ while (count = = 0 ) { try{ wait() } catch (InterruptedException ie ) { / / keeptryin g } } count --; / / co n sum ed } private synchronized void produce(){ count+ + ; notify ( ) ; / / notify theconsumer that count has been incremented . } }
Jayesh A Lalwani
Saloon Keeper
Joined: Jan 17, 2008
Posts: 2744
I like...
Apr 11, 2013 12:33:07
What's your question? Are you asking how wait and notify work internally?
I agree. Here's the link:
subject: How can threads communicate with each other?
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Sports Photos
Thunder 117, Mavs 114
Photo: Louis DeLuca/Staff Photographer
Dallas' Vince Carter (25) dunks as the Thunder's Thabo Sefolosha (2) and Serge Ibaka watch in the first quarter during the Oklahoma City Thunder vs. the Dallas Mavericks NBA basketball game at the American Airlines Center in Dallas on Friday, January 18, 2013.
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Superman costume to appear on SMALLVILLE after all
View Single Post
July 27 2009, 04:20 AM
Dick Whitman
Location: Behind the mask of Donald Draper
Re: Superman costume to appear on SMALLVILLE after all
Someone altered a sketch by Superman artist Gary Frank.
As per normal they do not seem to have much info about the regular cast. Everything is about new characters and guest stars. Whether Zod, Metallo or the JSA. They seem to be bored by the actual premise of the show.
Dick Whitman
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I had never heard my ins and outs, my ups and downs, my days and nights, my sorrows and joys, so opened up before. 1
William Gadsby was for thirty-nine years the remarkable, and highly successful, pastor of Back Lane (later renamed St. George's Road and then Rochdale Road) Baptist Chapel, Manchester. He is a widely known but little researched figure. 2 He was born in the village of Attleborough, Warwickshire. His exact date of birth is not known, but his baptism was registered in Nuneaton Church on 17 January 1773. William's father, John, had 7 children by his first wife, and a further 7 by his second wife, Martha, William being their second child. John Gadsby was employed as a road mender, and the Gadsby family were extremely poor. His large family ran wild about the village in a barefoot and ragged state. 3 The little education William Gadsby received was at Nuneaton Church School, which he attended two or three days a week, but much of what he learned


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good movie
I would preffer that this was from a direct source, rather than a video transfer. Oh well.
Many have whined and complained that this is actually an edited video from footage... Read more
I bought this dvd 3 years ago and I still put it on at least once a week and when we have a party, and love it every time along with everyone else that watches it, young and old... Read morePublished on March 16 2007 by Deborah J. Urtz
The solo that Eddie does is probably worth the price of the video. It features shots from only one of the nights, with a lot of good close-ups and clear views of Eddie's... Read morePublished on Jan. 25 2004
I'm writing this review for 2 reasons.
Reason 1: Pee on all of you, this DVD rocks. Every song, serenade, interview and solo is full of proff that Hagar did NOT ruin Van... Read more
This DVD starts well for about 8 minutes and then slowly puts the watcher to sleep. Bits and pieces cut up to make up a concert. Read morePublished on Dec 26 2003 by Flanger
I personally liked this DVD because it contained interviews with Alex and Michael, solo's with each of the band members, and also great live footage of all of the songs from the... Read morePublished on Dec 23 2003
We want a high quality DVD performance with Dave!!! This DVD is crap.Published on Sept. 29 2003 by M. Kupper
I'll just add to the litany of dissatisfaction with this video. First off, since this is the only live DVD, I have to recommend it, just to see them play and listen to the... Read morePublished on July 15 2003 by James Grote


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Microsoft VP Shane Kim has been anything but a stranger to these pages over the years: we first had a sit-down with the man way back in the halcyon days of early 2006, and as recently as this summer he was discussing the probable-possibility of Xbox games making their way onto mobile devices. Now, after nineteen years with the company and a career spanning the original Xbox through the present day (and Project Natal), our man has officially announced his retirement at the end of the year. Apparently his duties will be split between Dennis Durkin, who's been named chief operating officer of the company's video game unit, and Phil Spencer, who will oversee Microsoft Game Studios. Kim has yet to proclaim any post-Microsoft plans, saying he intends to relax and spend time with friends and family. Now that that's out of the way, Microsoft, when can we expect to see Live Anywhere up and running?
- Key specs
- Prices
- Discussions
- Game format Optical disc, Downloadable
- Online features Multiplayer, Voice chat, Video chat, Store, Browser
- Drive capacity 500 GB
- Controller type Wired, Wireless
- Motion controls Camera / optical
- Video outputs HDMI
- Released 2013-11-22
Microsoft Xbox 360


text: 0 minutes Less than a minute read March 21, 2013 Invisible Jason Russell CBC radio 1's Jian Ghomeshi conducted a "rare" interview with Jason Russell of KONY2012 fame this week. Russell, who is on some kind of media tour (he indulged the most….. Read More 2 minutes read February 2, 2012 Biko's Son The Canadian Broadcasting Corporation's Radio 1's program, The Current, this morning interviewed 40 year old Nkosinathi Biko about South Africa's progress (and his dad's legacy) 22 years after the unbanning of liberation movements (2 February 1990). Read More 3 minutes read January 22, 2012 Mandela and the ANC Nelson Mandela elicits divergent, incorrect and unrealistic reactions among his detractors and supporters. Read More ©Africa is a Country 2015


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T-cell intracellular antigen (TIA) proteins function as regulators of cell homeostasis. These proteins control gene expression globally at multiple levels in response to dynamic regulatory changes and environmental stresses. Herein we identified a micro(mi)RNA signature associated to transiently TIA-depleted HeLa cells and analyzed the potential role of miRNAs combining genome-wide analysis data on mRNA and miRNA profiles.
Using high-throughput miRNA expression profiling, transient depletion of TIA-proteins in HeLa cells was observed to promote significant and reproducible changes affecting to a pool of up-regulated miRNAs involving miR-30b-3p, miR125a-3p, miR-193a-5p, miR-197-3p, miR-203a, miR-210, miR-371-5p, miR-373-5p, miR-483-5p, miR-492, miR-498, miR-503-5p, miR-572, miR-586, miR-612, miR-615-3p, miR-623, miR-625-5p, miR-629-5p, miR-638, miR-658, miR-663a, miR-671-5p, miR-769-3p and miR-744-5p. Some up-regulated and unchanged miRNAs were validated and previous results confirmed by reverse transcription and real time PCR. By target prediction of the miRNAs and combined analysis of the genome-wide expression profiles identified in TIA-depleted HeLa cells, we detected connections between up-regulated miRNAs and potential target genes. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) database analysis suggest that target genes are related with biological processes associated to the regulation of DNA-dependent transcription, signal transduction and multicellular organismal development as well as with the enrichment of pathways involved in cancer, focal adhesion, regulation of actin cytoskeleton, endocytosis and MAPK and Wnt signaling pathways, respectively. When the collection of experimentally defined differentially expressed genes in TIA-depleted HeLa cells was intersected with potential target genes only 7 out of 68 (10%) up- and 71 out of 328 (22%) down-regulated genes were shared. GO and KEGG database analyses showed that the enrichment categories of biological processes and cellular pathways were related with innate immune response, signal transduction, response to interleukin-1, glomerular basement membrane development as well as neuroactive ligand-receptor interaction, endocytosis, lysosomes and apoptosis, respectively.
All this considered, these observations suggest that individual miRNAs could act as potential mediators of the epigenetic switch linking transcriptomic dynamics and cell phenotypes mediated by TIA proteins.
Keywords:TIA1; TIAR; miRNAs; Gene regulatory networks
Nowadays, the central dogma of Molecular Biology —developed from classic research works aimed at determining the biology of prokaryotic organisms— is known to reflect only a part of the agenda containing the genetic information that gives rise to the complexity of eukaryotic organisms. The characterization of post-transcriptional events leading to the generation of multiple RNAs, proteins and functions from only one RNA precursor shows up the existence of multiple overlapping regulatory networks and mechanisms for the control of biological functions beyond transcriptional regulation. There is increasing evidence to support the idea that transcriptome and proteome regulation and heterogeneity are key stages to understand differences in the protein diversity observed in organisms of similar genetic complexity. It's therefore necessary to fully characterize the modulators linking and synchronizing multiple layers of gene expression regulation.
T-cell intracellular antigen 1 (TIA1) and TIA1 related/like (TIAR/TIAL1) proteins are two DNA/RNA binding proteins that regulate many aspects of RNA metabolism at different levels. These multifunctional regulators can modulate: i) DNA-dependent transcription through its interaction with DNA and RNA polymerase II [1-4]; ii) alternative splicing of pre-mRNA through the selection of atypical 5′ spliced sites [5-8], and iii) stability and/or translation of eukaryotic mRNAs through the interaction with 5′ and/or 3′ untranslatable regions [9-16]. Some of these regulatory layers operate in the control of main biological programmes so as to maintain cellular homeostasis; this programmes include apoptosis, inflammation, cell responses to stress or viral infections ([10] and references included). Furthermore, mice lacking either TIA1 [16] or TIAR [17], as well as ectopically over-expressing TIAR [18], show higher rates of embryonic lethality.
MicroRNAs (miRNAs) are a class of 19-25 nt long non-coding RNAs that regulate post-transcriptionally gene expression by binding with partially complementary sequences on target mRNAs and inhibiting translation or affecting stability of these mRNAs [19]. Multiple lines of evidence indicate that they are key regulators of numerous critical functions in developmental, cell differentiation and disease processes, including tumorigenesis and cancer progression [19]. However, defining the place and function of miRNAs in complex regulatory networks is not straightforward. Systems' approaches such as the inference of a module network from expression data can help to achieve this goal [19].
We have previously described specific changes of transcriptomic dynamics associated to inflammation, angiogenesis, metabolism, and cell proliferation-related genes upon TIA1/TIAR-RNA interference-based silencing in HeLa cells [10]. In the present study, we test the hypothesis whether there are specific changes associated to the pattern of miRNA expression which may interfere/modulate with target genes and, therefore, contribute to the phenotypes described in TIA-depleted HeLa cells. Herein, we identified a miRNA signature that is concomitant and coherent with biological processes and pathways associated to the phenotypes observed in HeLa cells lacking TIA proteins.
Cell culture and RNA interference (RNAi)analysis
HeLa cells were grown and transfected with 20 nM of either a control siRNA (non-silencing siRNA duplex fluorescein labeled 27-6411-02FL from Gene Link) or two siRNAs against TIA1 (5′-AAGCTCTAATTCTGCAACTCTTT-3′; 5′-AACAACTAA TGCGTCAGACTTTT-3′) and TIAR (5′-AAGTCCTTATACTTCAGTTGTTC-3′; 5′-AACCATGGAATCAACAAGGATTT-3′) directed to the positions 59-81/647-669 and 65-87/971-993 to the coding regions of TIA1 and TIAR mRNAs, respectively, as described previously [7,10].
Cell extract preparation, western blot analysis and RNA purification
Whole-cell extracts were prepared by resuspensing the cells in lysis buffer (50 mM Tris–HCl, pH 8.0, 140 mM NaCl, 1.5 mM MgCl2, 0.5% Nonidet P-40 plus a mixture of protease inhibitors), freeze-thawing three times, and centrifugation at 10,000 rpm for 5 min. in a microfuge at 4°C. Resulting supernatants were recovered and stored at -70°C [10]. Immunoblots were carried out loading equal amounts of protein (15 μg) on 10% SDS-PAGE and using the following antibodies: anti-TIA1 and anti-TIAR from Santa Cruz Biotechnology (CA, USA) and anti-α-tubulin from Sigma (UK). RNA was extracted by using a miRVANA kit (Ambion, TX, USA) according to the manufacturer's protocol.
MicroRNA expression profiling analysis
The quality of the total RNA was verified by an Agilent 2100 Bioanalyzer profile. 2000 ng total RNA from each sample was labeled with Hy3™ or Hy5™ fluorescent label, using the miRCURY LNA™ microRNA Labeling Kit Hy3™/Hy5™ (Exiqon, Denmark), following the procedure described by the manufacturer. A Hy3™- and a Hy5™-labeled RNA sample were mixed pair-wise and hybridized to the miRCURY LNA™ microRNA Array (Exiqon, Denmark), which contains capture probes targeting all microRNAs for human registered in the miRBASE version 9.1. The hybridization was performed according to the miRCURY LNA™ microRNA Array Instruction manual using a Tecan HS4800™ hybridization station (Tecan, Austria). After hybridization the microarray slides were scanned and stored in an ozone free environment (ozone level below 2.0 ppb) in order to prevent potential bleaching of the fluorescent dyes. The miRCURY LNA™ microRNA Array slides were scanned using the Agilent G2565BA Microarray Scanner System (Agilent Technologies, Inc., USA) and the image analysis was carried out using the ImaGene® 7.0 (miRCURY LNA™ microRNA Array Analysis Software, Exiqon, Denmark). The quantified signals were background corrected (local background subtraction) and normalized using the global Lowess (LOcally WEighted Scatterplot Smoothing) regression algorithm.
Local background was corrected by normexp method with an offset of 50. Background corrected intensities were transformed to log scale (base 2) and normalized by Lowess for each array [20]. Finally, to have similar intensity distribution across all arrays, Lowess-normalized-intensity values were scaled by adjusting their quantiles [21]. After data processing each probe was tested for changes in expression over replicates using an empirical Bayes moderated t statistic [22]. To control the false discovery rate (FDR), P values were corrected using the method of Benjamini and Hochberg (1995) [23]. FIESTA viewer (http://bioinfogp.cnb.csic.es/tools/ webcite FIESTA) was used to visualize all microarray results and to evaluate the numerical thresholds (-2> fold change >2; FDR < 0.0001) applied for selecting differentially expressed genes [24].
The miRPlus sequences are licensed human sequences (Exiqon, Denmark). Some of them are already annotated in the miRBase database version 18. Microarray data discussed in this publication have been deposited in the NCBI Gene Expression Omnibus database (http://www.ncbi.nlm.nih.gov/geo/info/linking.html webcite) and are accessible through the GEO Series accession number GSE41213.
The method was optimized for microRNA, and reagents, primers, and probes were obtained from Applied Biosystems. Reverse transcriptase (RT) reactions and real-time PCR (PCR) were performed according to manufacturer protocols at the Genomic PCR Core Facility at Universidad Autónoma de Madrid in Madrid Scientific Park. Analyses were performed in two independent samples by triplicate, including no-template and RT-minus controls. U6 RNA and miR-200 expression were used as endogenous reference controls. Relative miRNA expression was calculated using the comparative cycle threshold method.
Generation of miRNA targets dataset
In silico targets predicted for each of the differentially expressed miRNAs by three different algorithms: TargetScan 5.2 ([25] and references included), PicTar-Vert ([26] and references included) and miRDB [27,28] were downloaded using web-app miRBase [29] (http://www.mirbase.org webcite). Given that each algorithm focus on different aspects of miRNA-mRNA pairing, and the lack of experimental validation of most miRNAs targets do not allow a false-positive elimination, we kept the datasets by considering the following score values: TargetScan 5.2 (aggregate PCT > 0.1), PicTar-Vert (PicTar score > 2) and miRBD (target score > 70).
Gene ontology, pathway and network analyses
The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) database analysis were conducted using software programmes provided by GenCodis3 (http://genecodis.cnb.csic.es webcite) [30,31]. Networks and regulatory topologies of functional relationships between gene clusters were created using the CytoScape [32] (http://www.cytoscape.org webcite).
To analyze the putative role of TIA proteins in the control of gene expression on microRNAs, we transfected HeLa cells with double-stranded small interfering RNAs (siRNAs) targeting TIA1 and TIAR mRNAs or with control siRNA (C), as previously reported [10]. The effect of siRNAs on TIA1/TIAR expression was analyzed by Western blotting (Figure 1A). Upon TIA1/TIAR-RNA interference-based silencing, 80-90% and 70-80% depletions of TIA1 and TIAR proteins, respectively (Figure 1A) were achieved 72 h after transfection, thus in agreement with previous observations [10]. By contrast, α-tubulin protein (Figure 1A) was used as control of siRNA specificity, and its expression level was not significantly affected by gene interference approach.
Figure 1. Small interfering RNA (siRNA)-mediated depletion of TIA proteins. (A) Western blot analysis of HeLa cell lysates prepared 72 h after transfection with siRNAs against control (C; lanes 1, 3 and 5) and TIA1 plus TIAR (lanes 2, 4 and 6). The blot was probed with antibodies against TIA1, TIAR, and α-tubulin proteins, as indicated. Molecular weight markers and the identities of protein bands are shown. (B) A schematic representation depicting the miRNA array strategy used in this study. The preparation of the RNA samples from control and TIA-depleted HeLa cells, labeling of above RNA samples with Hy3 and Hy5 dyes, hybridization and analyses of the resulting miRNA profiles are illustrated.
To test the microRNA (miRNA) expression profiles resulting from the reduction of TIA1 and TIAR protein levels, the differences in global miRNA expression patterns in control and TIA1/TIAR-depleted HeLa cells were determined by means of a miRCURY™ LNA Array with specific probes for simultaneous analysis of at least 600 different miRNAs. After having passed sample QC on the Bioanalyzer2100 and RNA measurement on the Nanodrop instrument, the samples were labelled using the miRCURY™ Hy3™/Hy5™ labelling kit and hybridized on the miRCURY™ LNA Array (v.8.1) (Figure 1B). Analysis of the scanned slides showed that the labelling was successful as all capture probes for the control spike-in oligo nucleotides produced signals in the expected range. The quantified signals were normalized using the global Lowess (Locally WEighted Scatterplot Smoothing) regression algorithm, which we have found produces the best within-slide normalization to minimize the intensity-dependent differences between the dyes (Figure 1B). The positive effect of this normalization is illustrated in 3 different plots for each experimental conditions analyzed (Additional file 1). It is interesting to note the high reproducibility of individual miRNA expression levels and their correlation across the different miRNA pools (Additional file 1). On the other hand, an appropriate statistical test analysis was made using a linear model (as implemented in the limma R/Biocounductor package) to compare miRNA expression patterns for three two-color arrays performed from three independent biological replicates (Figure 1 and Additional file 1).
Additional file 1. Summary of the miRNA array analyses. The following additional data are available in this file: 1) Correlation of Hy3 and Hy5 signals for the spike-in controls between slides. 2) Hy5 vs Hy3 scatter, MA and ratio distribution plots (before and after normalization) and Hy5 vs Hy3 scatter plot for the spike-in controls, for each independent experiments. 3) Statistical test implemented in the limma R.
Format: XLS Size: 1.6MB Download file
This file can be viewed with: Microsoft Excel Viewer
As shown in Table 1, depletion of TIA-proteins resulted in significantly altered miRNA expression profiling. The expression level of 29 out of 600 miRNAs was found to more than double when comparing channels Hy5 and Hy3. The genome-wide profiling analysis identified 17 miRNAs and 12 putative in-silico miRNAs (identified as miRPlus). These were the well-established miRNAs: miR-197-3p, miR-210, miR-373-5p, miR-492, miR-498, miR-503-5p, miR-572, miR-586, miR-612, miR-615-3p, miR-623, miR-625-5p, miR-638, miR-658, miR-663a, miR-671-5p and miR-769-3p, which were differentially up-regulated at least 2-fold (FDR<0.0001). The identified miRPlus sequences were in licensed human sequences; now a great number of these sequences have been annotated in the corresponding miRNA database. This information is available in Table 1. Regarding the differentially expressed miRPlus in our experimental conditions, an update of these miRPlus sequences in the miRBase 18 indicates that miRPlus-17836 is miR-30b-3p, miRPlus-17864 is miR-744-5p, miRPlus-17867 is miR-203a, miRPlus-17877 and miRPlus-17960 are miR-483-5p, miRPlus-17878 is miR-193a-5p, miRPlus-17942 is miR-125a-3p, miRPlus-17950 is miR-371-5p and miRPlus-17961 is miR-629-5p (Table 1). The remaining miRPlus sequences have not been yet assigned to the putative specific miRNAs (Additional file 2).
Additional file 2. Update of miRNAs sequences previously identified as miRPlus. The miRPlus sequences are licensed human sequences (Exiqon, Denmark). Many of them are already annotated in the miRBase database version 18.
Format: XLS Size: 104KB Download file
This file can be viewed with: Microsoft Excel Viewer
Table 1. MicroRNA expression profiling in TIA-depleted HeLa cells
To validate previous results on identified miRNAs and their relative expression levels, four up-regulated and two unchanged miRNAs were confirmed by TaqMan reverse transcription and polymerase chain reaction (QPCR) analyses. As shown in Figure 2, the results obtained by quantitative amplification were fully comparable and the relative fold changes in miRNA expression were consistent with the data detected by hybridization in the corresponding microarrays (compare Table 1 and Figure 2).
Figure 2. Validation of miRNA array-predicted changes by quantitative RT-PCR (QPCR). Quantitative miRNA expression analysis was carried out using TaqMan probes by QPCR. The represented values were normalized and expressed relative to control values (whose value was fixed arbitrarily to 1) as ratios and are means ± SD (n = 2).
As a first attempt to understand the relevance of 29 differentially up-regulated miRNAs, in silico methods for predicting human miRNA target genes were used (Figure 3). The potential target genes were identified by searching the TargetScan, PicTar and miRBD databases [25-28]. By comparing and selecting alone non-repeated target genes, 2683 miRNA target genes were identified (Additional file 3). All these potential target genes were tested using computer tool GeneCodis3 [20,21] to elucidate the biological processes and cellular pathways assigned using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) database analyses (Figure 3 and Additional file 3). A total of 253 biological processes from GO database and 50 cellular pathways from KEGG categories were estimated with a significance hypergeometric test (corrected hypergeometric p value < 0.01). Target genes corresponding to the biological processes from GO categories were mainly involved in regulation of DNA-dependent transcription, signal transduction, multicellular organismal development, positive/negative regulation of transcription from RNA polymerase II promoter, cell adhesion, transmembrane transport, apoptotic process and nervous system development (Table 2 and Additional file 3). In addition, target genes associated to the main biological pathways were also identified using KEGG database involving pathways in cancer, MAPK signalling pathway, focal adhesion, endocytosis, regulation of actin cytoskeleton, Wnt signalling pathway, neuroactive ligand-receptor interaction, glutamatergic synapse, ubiquitin mediated proteolysis and tuberculosis (Table 2 and Additional file 3). Collectively, these results suggest that TIA-protein depletion promotes the induction of a miRNA signature which may directly and indirectly contribute to the establishment of the observed cellular phenotypes in TIA-deficient HeLa cells [10].
Additional file 3. List of predicted potential target genes associated to up-regulated miRNAs in transiently TIA-depleted HeLa cells. Potential target genes of up-regulated miRNAs identified in TIA-deficient HeLa cells were predicted by TargetScan 5.2, PicTar-Vert and miRDB database downloaded using web-app miRBase (http://www.mirbase.org webcite). The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) database analyses were conducted using software programmes provided by GenCodis3 (http://genecodis.cnb.csic.es webcite).
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Figure 3. Prediction of potential and experimental target genes associated with up-regulated miRNAs in TIA-depleted HeLa cells. Integrative analyses of potential target genes and miRNAs regulated in TIA-depleted HeLa cells using TargetScan 5.2, PicTar-Vert and miRDB software tools (see corresponding section in Methods). Diagrams depicting the number of potential target genes and up-regulated miRNAs (Additional file 3) by TIA silencing are shown. Venn diagram depicting the numbers of genes that were intersected among putative target genes (highlighted in gray) associated to miRNAs and experimentally defined differentially expressed genes (see additional data files in [10]) in HeLa cells lacking TIA proteins. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) database analyses were conducted using software programmes provided by GenCodis3.
Table 2. Top ten biological processes and pathways associated to up-regulated miRNAs in TIA-depleted HeLa cells
On the other hand, the collection of differentially expressed genes previously identified by expression microarray Human Genome U133 Plus 2.0 (Affymetrix) in TIA-depleted HeLa cells [10] were intersected with potential target genes identified using in silico target prediction tools. The results show that only 7 out of 68 (10%) up- and 71 out of 328 (22%) down-regulated genes were shared (Figure 3 and Table 3). GO and KEGG database analyses were independently performed for up- and down-regulated genes with GeneCodis3 software. The results suggest that the enrichment functional categories (p < 0.05) are related with signal transduction, innate immune response and response to interleukin-1 for up-regulated genes and glomerular basement membrane development for down-regulated genes. Further, cellular pathways of neuroactive lingand-receptor interaction and apoptosis were associated to up-regulated genes, whereas endocytosis and lysosomes were linked to down-regulated genes (Table 4). Taken together, these results suggest that at least a fraction (40%) of the up-regulated miRNAs (12 out of 30) could be contributing to the establishment of differential expression profiles associated to the HeLa cells lacking TIA proteins.
Table 3. Intersection between potential and experimentally defined target genes related to miRNAs in TIA-depleted HeLa cells
Table 4. Biological processes and pathways linked to experimentally defined and differentially expressed genes in TIA-depleted HeLa cells
Based on above observations an interesting question emerges: why should many specific genes be up- or down-regulated by induced miRNAs in TIA-depleted HeLa cells? A simple answer to this question might be that these mRNAs or their precursors (pre-mRNAs) are targeted by TIA proteins through one or multiple layers to exert the control of their gene expression; thus, these regulators can act as multifunctional RNA binding proteins (see references included in Background). To approach this issue, the experimental profiles of the binding patterns of TIA proteins (i.e., the RNA map corresponding to TIA proteins) at the up- or down-regulated pre-mRNAs (Figure 4 and Table 3) were examined using the iCLIP database of TIA proteins kindly provided by Jernej Ule's laboratory [8]. For example, as shown in Figure 4 and in Table 3, the pre-mRNAs analyzed show multiple sequence sites located across the full-length pre-mRNAs, both exons and introns, and which we have classified as genes with either low (+), medium (++) or high (+++) density of TIA binding sites. The greatest relevance of this observation is the fact that the density of binding sites on these pre-mRNAs is found in both up- and down-regulated genes and located with frequency on the last exons of these pre-mRNAs and particularly on the sequences located at the 3′ untranslated regions of the mature mRNAs. In this regard, it is reasonable to think that the existence of a feedback loop that represses the expression of many genes, which could be activated in the absence of TIA proteins, for example at the post-transcriptional levels (i.e. mRNA stability or translational activation), to dampen its expression in order to promote or counteract the cellular phenotypes associated to the absence of the TIA proteins.
Figure 4. RNA map of TIA proteins in targeted pre-mRNAs related with miRNAs in TIA-deficient HeLa cells. (A and B) Profiles of experimental crosslink sites of TIA1 and TIAR proteins (adapted from TIA-iCLIP analysis [8]) on up- (A) and down-regulated (B) pre-mRNAs in TIA-depleted HeLa cells. Representative examples of up- (A) and down-regulated (B) genes with low (+), medium (++) and high (+++) density of TIA binding sites are shown. The bar graph in each panel indicates the number of cDNAs that identified each crosslink site.
During the last decade, much progress has been made in the understanding of network topology and the relevance and properties of its basic modules. In this study, we analyze and assess module networks inferred from both miRNAs and gene expression data using a bioinformatic tool as CytoScape [32]. By matching expressed miRNAs and experimentally defined up- and down-regulated genes in TIA-depleted HeLa cells, a putative regulatory network of TIA-associated genes and miRNAs was constructed. Based on the number of potential gene interactions with single miRNAs, both up- and down-expressed genes regulating TIA-mediated differential expression were connected (Figure 5). This network draws a cellular scenario where the reduction of TIA proteins could lead to molecular responses mediated by individual miRNAs. Such a computational approach, starting from expression data alone, can be helpful in the future process of identification of the function of these miRNAs by suggesting modules of co-expressed genes in which they could play a regulatory role.
Figure 5. Regulatory network of up-regulated miRNAs and intersection with target genes associated with TIA-depleted HeLa cells. Elongated circles represent differentially expressed miRNAs and genes (red and brown- increased, green- decreased). Lines represent regulatory relations between differentially expressed miRNAs and genes.
Current data suggest that a significant portion of the information containing the human genome is regulated by miRNAs. These small RNAs recognize and regulate target genes. In this regard, more than 60% of protein-coding genes are predicted in silico as targets, based on conserved base-pairing between the 3′- and 5′-untranslated regions (UTR) of the mRNA and the seed sequence of the miRNAs [33] without considering putative target on coding sequences. At present, near to 2,000 miRNAs have been identified in the human genome and about 20-30% of human genes are controlled by one or more miRNAs [33-36]. Multiple lines of evidence indicate that they are key regulators of numerous critical functions in development and disease, including cancer. Many of them have been reported to have molecular features and act either as tumour suppressors or oncogenes [33-35]. The changes in miRNA signature identified in this study might directly and indirectly function as encouraging/counteracting mechanism of biological processes and cellular pathways to promote/attenuate the inflammatory, angiogenic and proliferative responses linked to TIA-depleted HeLa cells [10].
Given that many target genes associated with the identified up-regulated miRNAs are down-regulated (Table 3) in TIA-depleted HeLa cells [10], from a mechanistic viewpoint our results indicate that mRNA abundance in most targeted genes was somewhat affected by miRNAs, thus suggesting that, for a substantial number of genes regulated by TIA-protein absence, destabilization of mRNA may be the main mechanism of protein repression by these miRNA-mediated regulators. This observation agrees with a recent study, suggesting that mammalian miRNAs predominantly reduce target mRNA levels [34]. However, some miRNAs such as miR-744 enhances cyclin B1 mRNA expression through a novel mechanism. miRNA positively-regulates gene expression by targeting promoter elements; this phenomenon is known as RNA activation [37]. In this regard, it is reasonable to think that this mechanism could be operating in the up-regulation observed for seven target genes associated with the up-regulated miRNAs (Table 3). Furthermore, miRNAs do not only regulate the expression of protein-encoding genes but also other miRNAs: for instance, let-7a controls the expression of important epigenetic regulators, including epigenetic miRNA regulatory circuits, and organizes the whole gene expression profile [38]. On the other hand, there are miRNA-target interactions that involve multiple sites for a given target and confer much stronger repression. More often, different miRNAs work together to co-target a given mRNA, therefore their combined repressive effect greatly exceeds the individual contributions [34-36]. Both regulatory situations are observed in the down-regulated genes associated to up-regulated miRNAs in TIA-depleted HeLa cells (Table 3). This suggests that clusters of miRNAs can play a more prominent role than only reinforcing the expression patterns dictated by transcriptome dynamics. The existence of interactions among these regulators and the interactions between their regulatees suggests that these miRNAs generate networks that modulate antagonistic cellular responses, such as apoptosis or cell proliferation induction and/or repression. Our set of data indicates that the action of miRNAs could potentially be another important mechanism in the regulation of gene expression and some gene regulatory networks mediated by TIA proteins. These observations thus suggest the existence of feedback mechanisms that promote miRNA expression, which might therefore contribute to dampen the phenotypes observed in TIA-lacking HeLa cells. A type 2 incoherent feedforward loop [39] may contribute to the repression model between TIA proteins, target genes and associated miRNAs. This feedforward loop could reinforce the functional role of TIA proteins as repressors of inflammatory, angiogenic, and proliferative responses (Figure 6).
Figure 6. An overview summarizing the cellular and molecular events associated to the reduction of TIA proteins. The availability of TIA proteins could be contributing to a regulatory loop mediated by miRNAs to modulate target gene expression related to inflammatory, angiogenic and/or proliferative responses/phenotypes in HeLa cells.
Regarding some miRNAs identified in this study, these could be related to miRNA-mediated gene expression fluctuations and, more faithfully, to signal outcomes in the context of specific regulatory networks. This suggest that miRNAs can strenghten transcriptome patterns by buffering the deleterious effects of some network states linked to random fluctuations in gene expression program, in agreement with previous findings [25]. Since little experimental information is available on potential target genes and miRNA cluster identified in our study and we have not experimentally carried out analysis of gain- and loss-of function, we used experimental data on miRNA function to understand the potential implication and relevance of these miRNAs in the development and progression of (patho)-physiological conditions. For example, miRNA-30b has been implicated in angiogenesis [40], TRAIL-induced apoptosis in glioma cells [41] and oral squamous cell cancers [42]. miR-193a-5p targets YY1-APC regulatory axis in human endometrioid endometrial adenocarcinoma [43]. miR-203 participates in a regulatory network that modulates epithelial to mesenchymal transition [44] as well as to promote suppression immune [45]. miR-210 is induced under hypoxia and works as a iron sensor to stimulate cell proliferation and promote cell survival in the hypoxic region within tumor [46-48]. Further, this miRNA modulates the mitochondrial functioning and metabolism [49,50], represses FGFRL1 and E2F3 expression, which inhibits cell apoptosis in hypoxia response [51,52], predicts poor survival in patients with breast cancer [53] and activates notch signalling pathway in angiogenesis [54]. In addition, miR-371-5p is increased in gastric cancer [55] and miR-373 functions as an oncogene in hepatocellular carcinoma given that is a new regulator of protein phosphatase 6 [56], an repressor of the large tumor suppressor homolog 2 [57] and promotes tumor invasion and metastasis in testis cancer [58,59]. Further, both miRNA-210 and miRNA-373 participate in the gene expression control of DNA repair in hypoxic stress [60]. miRNA-492 and miRNA-498 are highly expressed and regulate metastatic functions in hepatoblastoma, rectal cancer, adenocarcinoma and retinoblastoma [61-64]. In addition, miR-503 is induced in angiogenesis, down-regulates CUGBP1 and modulates metastatic function in hepatocellular cancer cell [65-68]. miRNA-572 has been involved in chronic lymphocytic leukaemia via targeting anti-apoptotic genes [69,70]. miRNA-586 is up-regulated in colorectal cancer [71]. miRNA-615-3p dysregulates CDKN2A, NF2 and JUN in malignant mesothelioma [72] and enhances the phagocytic capacity of splenic macrophages [73]. miRNA-625 promotes invasion and metastasis of gastric cancer by targeting ILK [74]. miR-629 is associated to lung cancer by targeting NBS1 [75]. miRNA-638 and miR-658 are up-regulated in cell transformation and human gastric cancer [76,77]. In addition, other miRNAs show a great potential as activators of cell proliferation/transformation phenotypes; for example miRNA-663 functions as an oncogene promoting tumorigenesis by targeting p21(WAF1/CIP1), VEGF, JunB, JunD and TGFB1 genes [78-80] and plays also a role in inflammatory response of endothelial cells [81]. miRNA 671 regulates CD44 inducing metastasis by regulating extracellular matrix functions [82] as well as miRNA-dependent gene silencing involving Ago2-mediated cleavage of a circular antisense RNA [83]. miRNA-744 up-regulates Cyclin B1 expression [37] and down-regulates Transforming Growth Factor Beta-1 expression [84]. On the other hand, other miRNAs identified in our study have bipolar features and putatively antagonistic behaviours as opposite regulators acting either positively or negatively under specific biological programmes. In this regards, there are at least 2 miRNAs showing functions to repress proliferation, angiogenesis and transformation phenotypes. This is the case of miRNA-483-5p, which suppresses the proliferation of glioma cells via directly targeting ERK1 [85] and angiogenesis in vitro by targeting serum response factor [86]. However, microRNA-483-5p and miRNA-195 have been identified as predictors of poor prognosis in adrenocortical cancer [87]. miR-125a-3p is down-regulated in non-small cell lung cancer, having inverse effects on invasion and migration of lung cancer cells [88]. This miRNA is a potent prognostic marker in gastric cancer [89]. Collectively, the miRNA expression profiling identified in this study has important aftermath in tumorigenesis and could contribute/repress to the development of cell proliferation, angiogenesis, transformation phenotypes, because individual miRNAs are associated with diagnosis, prognosis and treatment efficacy linked to the biology of human tumors. Therefore, these miRNAs can target to oncogenes or tumor suppressor genes (also they can even function themselves as such) and take part in the promotion or inhibition of tumorigenesis and cancer progression, thus being able to confer robustness to these pathological phenotypes.
In summary, our findings suggest that expression changes in individual miRNAs in TIA-depleted HeLa cells could directly or indirectly impact on biological processes and signaling pathways to favour cell phenotypes associated to the down-regulation of TIA proteins (Figure 6). Further, our observations suggest that some of the identified miRNAs are consistent with an adaptive response aimed at attenuating the inflammatory, angiogenic, and proliferative responses developed in TIA-protein absence. Therefore, cross talk between regulatory mechanisms that promote increased/reduced expression of miRNA-mediated genes may contribute to modify the relative expression levels of the mRNAs associated with TIA-lacking HeLa cells. Based on this integrative analysis, our results provide a prominent stand for future approaches aimed at characterizing the role of specific miRNAs in TIA-mediated gene expression regulation.
Gene expression profiling approaches have improved our understanding on how the human transcriptome dynamics orchestrates global responses at molecular level to answer to changing environmental challenges. Our study identifies a human miRNA collection that displays significant changes in the transient absence of TIA proteins in HeLa cells. The most prominent changes are linked to the up-regulation of miR-30b-3p, miR125a-3p, miR-193a-5p, miR-197-3p, miR-203a, miR-210, miR-371-5p, miR-373-5p, miR-483-5p, miR-492, miR-498, miR-503-5p, miR-572, miR-586, miR-612, miR-615-3p, miR-623, miR-625-5p, miR-629-5p, miR-638, miR-658, miR-663a, miR-671-5p, miR-769-3p and miR-744-5p. The integration of the identified miRNAs with the potential target genes and previous gene expression data under the same experimental conditions revealed enrichment of biological processes and signaling pathways controlling relevant members of the pathways associated to the oncogenesis. Down-regulation of these cellular components may contribute to establish the inflammatory, angiogenic, and proliferative responses previously described in TIA protein-lacking HeLa cells. Therefore, our results are consistent with the existence of regulatory networks that generate correlated expressions, commonly involving miRNAs [33,90]. This regulatory architecture may increase the fidelity of inhibition of the downstream components by acting on them redundantly. In other words, a transient loss of TIA proteins can be partially compensated for by the adaptive presence of specific miRNAs. Collectively, these findings suggest that TIA proteins can act as multifunctional regulators to provide a novel meeting point between the mechanisms for cross-talking among concerted post-transcriptional regulatory layers that coordinate complex cellular responses.
iCLIP: In vivo ultraviolet (UV)-crosslinking and immunoprecipitation to identify the RNA crosslinking sites of TIA1 and TIAR proteins; GO: Gene ontology; KEGG: Kyoto and encyclopedia of genes and genomes; siRNA: Small interfering RNA; TIA1: T-cell intracellular antigen 1; TIAR/TIAL1: TIA1 related/like protein; UTR: Untranslated region of the eukaryotic mRNAs.
Competing interests
The authors declare that they have no competing interests.
Authors' contributions
JMI conceived the research and designed all experiments. CSJ, IC and JB carried out the experiments and analyzed the data presented in this paper. CSJ, IC, JB and JMI wrote the paper. All authors read and approved the final manuscript.
We are grateful to Jernej Ule for providing iCLIP data analysis of TIA proteins and Juan Carlos Oliveros for helping us with bioinformatics tools. This work was supported by grants from the Spanish Ministry of Innovation and Science through FEDER funds (BFU2008-00354 and BFU2011-29653). The CBMSO receives an institutional grant from Fundación Ramón Areces.
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Zhu DX, Zhu W, Fang C, Fan L, Zou ZJ, Wang YH, Liu P, Hong M, Miao KR, Liu P, Xu W, Li JY: miR-181a/b significantly enhances drug sensitivity in chronic lymphocytic leukemia cells via targeting multiple anti-apoptosis genes.
Xiao W, Bao ZX, Zhang CY, Zhang XY, Shi LJ, Zhou ZT, Jiang WW: Upregulation of miR-31* is negatively associated with recurrent/newly formed oral leukoplakia.
Cummins JM, He Y, Leary RJ, Pagliarini R, Diaz LA Jr, Sjoblom T, Barad O, Bentwich Z, Szafranska AE, Labourier E, Raymond CK, Roberts BS, Juhl H, Kinzler KW, Vogelstein B, Velculescu VE: The colorectal microRNAome.
Guled M, Lahti L, Lindholm PM, Salmenkivi K, Bagwan I, Nicholson AG, Knuutila S: CDKN2A, NF2, And JUN are dysregulated among other genes by miRNAs in malignant mesothelioma. A miRNA microarray analysis.
Jiang A, Zhang S, Li Z, Liang R, Ren S, Li J, Pu Y, Yang J: miR-615-3p promotes the phagocytic capacity of splenic macrophages by targeting ligand-dependent nuclear receptor corepressor in cirrhosis-related portal hypertension.
Wang M, Li C, Nie H, Lv X, Qu Y, Yu B, Su L, Li J, Chen X, Ju J, Yu Y, Yan M, Gu Q, Zhu Z, Liu B: Down-regulated miR-625 suppresses invasion and metastasis of gastric cancer by targeting ILK.
Yang L, Li Y, Cheng M, Huang D, Zheng J, Liu B, Ling X, Li Q, Zhang X, Ji W, Zhou Y, Lu J: A functional polymorphism at microRNA-629-binding site in the 3′-untranslated region of NBS1 gene confers an increased risk of lung cancer in Southern and Eastern Chinese population.
Li D, Wang Q, Liu C, Duan H, Zeng X, Zhang B, Li X, Zhao J, Tang S, Li Z, Xing X, Yang P, Chen L, Zeng J, Zhu X, Zhang S, Zhang Z, Ma L, He Z, Wang E, Xiao Y, Zheng Y, Chen W: Aberrant expression of miR-638 contributes to benzo(a)pyrene-induced human cell transformation.
Guo J, Miao Y, Xiao B, Huan R, Jiang Z, Meng D, Wang Y: Differential expression of microRNA species in human gastric cancer versus non-tumorous tissues.
Liu ZY, Zhang GL, Wang MM, Xiong YN, Cui HQ: MicroRNA-663 targets TGFB1 and regulates lung cancer proliferation.
Tili E, Michaille JJ, Adair B, Alder H, Limagne E, Taccioli C, Ferracin M, Delmas D, Latruffe N, Croce CM: Resveratrol decreases the levels of miR-155 by upregulating miR-663, a microRNA targeting JunB and JunD.
Yi C, Wang Q, Wang L, Huang Y, Li L, Liu L, Zhou X, Xie G, Kang T, Wang H, Zeng M, Ma J, Zeng Y, Yun JP: MiR-663, a microRNA targeting p21(WAF1/CIP1), promotes the proliferation and tumorigenesis of nasopharyngeal carcinoma.
Ni CW, Qiu H, Jo H: MicroRNA-663 upregulated by oscillatory shear stress plays a role in inflammatory response of endothelial cells.
Rutnam ZJ, Yang BB: The non-coding 3′ UTR of CD44 induces metastasis by regulating extracellular matrix functions.
Hansen TB, Wiklund ED, Bramsen JB, Villadsen SB, Statham AL, Clark SJ, Kjems J: miRNA-dependent gene silencing involving Ago2-mediated cleavage of a circular antisense RNA.
Martin J, Jenkins RH, Bennagi R, Krupa A, Phillips AO, Bowen T, Fraser DJ: Post-transcriptional regulation of transforming growth factor beta-1 by microRNA-744.
Wang L, Shi M, Hou S, Ding B, Liu L, Ji X, Zhang J, Deng Y: MiR-483-5p suppresses the proliferation of glioma cells via directly targeting ERK1.
Qiao Y, Ma N, Wang X, Hui Y, Li F, Xiang Y, Zhou J, Zou C, Jin J, Lv G, Jin H, Gao X: MiR-483-5p controls angiogenesis in vitro and targets serum response factor.
Soon PS, Tacon LJ, Gill AJ, Bambach CP, Sywak MS, Campbell PR, Yeh MW, Wong SG, Clifton-Bligh RJ, Robinson BG, Sidhu SB: miR-195 and miR-483-5p identified as predictors of poor prognosis in adrenocortical cancer.
Jiang L, Huang Q, Zhang S, Zhang Q, Chang J, Qiu X, Wang E: Hsa-miR-125a-3p and hsa-miR-125a-5p are downregulated in non-small cell lung cancer and have inverse effects on invasion and migration of lung cancer cells.
Hashiguchi Y, Nishida N, Mimori K, Sudo T, Tanaka F, Shibata K, Ishii H, Mochizuki H, Hase K, Doki Y, Mori M: Down-regulation of miR-125a-3p in human gastric cancer and its clinicopathological significance.
Friard O, Re A, Taverna D, De Bortoli M, Corá D, Circuits DB: a database of mixed microRNA/transcription factor feed-forward regulatory circuits in human and mouse.


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I find the 100mm length ideal for use with a butterfly bracket and a flash. I've had the faster Pentax SMC-A 100mm f/2.8 Macro for 20 years, the image is much brighter than the f/4 K version. Still it can be a challenge to focus with the helix racked out at full extension. Mine has the reputation of being better corrected due to the FREE focusing (which stands for Fixed Rear Element Extension, the rear element group moves near maximum extension). It goes to 1:1 without additional extension tubes and IIRC the f/4 PK Macro goes only to 1:2 (half life sized).
The reputation of the SMC-A f/2.8 lens being legendarily sharp is well deserved, I can't speak to the f/4 PK version. But I can tell you any K version won't work as well with all exposure modes using Pentax bodies more recent than about the LX of 1980, without the lens information contacts or the "A" setting.
You're rather unlikely to find one for the $100 price mentioned, though. The PK 100 f/4 Macros typically go for twice that, and the f/2.8's for $400-800 depending on condition. Either is rarer than the 50mm f/2.8, also wickedly sharp and contrasty, and which seem to sell for about $250. The 200 f/4' ED's are another matter entirely, there's one at auction right now that'll likely go for close to $2k (It was only about $700 when available new--they were very, very scarce even in 1990).


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Associations Focus: CEDIA Awards Deadline ApproachesEnter in one of the following Manufacturers' Excellence Award categories: Best New Product Best New Technology Product Hall of Fame All companies entering 3/01/2009 7:00 AM Eastern
Associations Focus: CEDIA Awards Deadline Approaches
Mar 1, 2009 12:00 PM
Enter in one of the following Manufacturers' Excellence Award categories:
- Best New Product
- Best New Technology
- Product Hall of Fame
All companies entering the Manufacturers' Excellence Awards must be active CEDIA Manufacturer Members. If you are entering the Best New Product or Best New Technology categories, you must also be officially exhibiting at CEDIA Expo 2009.
The deadline for Manufacturers' Excellence Award submissions is May 8, 2009, at 5 p.m. PST. In honor of CEDIA's 20th Anniversary, completed awards submissions received on or before April 1, 2009, will receive a 20 percent discount toward the cost of the first awards submission.
Award winners will be selected from the pool of finalists and announced at the CEDIA Electronic Lifestyles Awards banquet, Saturday, Sept. 12, 2009. Best New Product and Best New Technology finalists will be announced and promoted starting in early July.
For information on the guidelines and entry forms of the CEDIA Manufacturers' Excellence Awards, please visit www.cedia.org/awards, or email firstname.lastname@example.com with any questions.
The CEDIA Electronic Lifestyles Awards are the most prestigious form of recognition in the residential-electronic-systems industry. These highly sought after awards are presented each year to recognize excellence and hard work in the industry.
Get recognized for your hard work and submit an awards entry today. The 2009 Electronic Lifestyles Designer Awards entry forms and guidelines are available at www.cedia.org/awards. The deadline for Designer Award entries is May 1, 2009 at 5 p.m. PST.
In honor of CEDIA's 20th anniversary, members who submit a completed entry online and mail in all required materials by April 1, 2009, will receive 20 percent off the cost of their first entry.
For detailed information on how to submit an entry for CEDIA's Electronic Lifestyles Designer Awards, please visit www.cedia.org/awards. If you have any questions about the CEDIA Awards program, please email email@example.com.
For more information about CEDIA, visit www.cedia.org or call (800) 669-5329.
CEDIA University is making it easier than ever to gain the knowledge and skills to be successful in the residential-electronic-systems industry. Check out these great educational opportunities.
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Associations Focus: CEDIA Awards Deadline Approaches
Mar 1, 2009 12:00 PM
March 26
Intermediate EST Workshop: Taking Your Skills to the Next Level
Orange, Calif.
March 26-28
EST Advanced Residential Boot Camp
April 1
ESPA-certified EST Review and Exam
Associated Lighting Representatives Oakland, Calif.
April 19-21
CU On the Road @ Capitol Sales Learning Institute Regional Outreach
April 21-23
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EST Basic Residential Boot Camp
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The style of delivery was wildly different for each person. Some of us were very matter of fact and detailed. Some of us were more big picture about it. When we each shared our statements, it gave everyone else on the team an opportunity to fill in some gaps of perception, like… Read More
A What's Hot / What's Not list for your business or career
If you want to get awesome results, you've got to get real about what's currently not-so-awesome. In the Desire Map process you begin by going through a gratitude exercise, followed by a dissatisfaction review. It's the whole picture, with a ton of love and no-sugar coating… Read More
How we run meetings. Get personal. Get down to business. Get it done.
Being a virtual company is mostly all great — the freedom, working in your yoga pants with a puppy on your lap — or from a hotel room, solitude to create… Read More
Cheap easy. Quality easy. And The Myth of Endurance.
What would your life be like if you only did what was easy? It's almost unsettling to go there, isn't it? When I use to try to answer that question for myself I'd squirm a bit. Lazy dilettante. As if. What would I do with all that extra time I'd have if I just did the easy stuff? Hmmm . . . maybe I'd have more time to enjoy what I've got and to get more of what I want. Maybe things would be . . . easier… Read More
Champagne & happy tears. It's your turn to lead. The Desire Map Licensing Program launches TODAY. For real. For everyone.
There have been plenty of times in the last few months where I sat at my desk and let the tears roll. Happy tears of "Holy, what, really?!" Like when I saw the pictures of the first Desire Map workshop that was being taught — without me. And the women were laughing, posting core desired feelings on sticky notes, and building a "desire altar." I just whispered to myself, "Wow, it's happening." I was a puddle that day. And I'll probably cry writing this post. Because it's a very, very big day… Read More
You can earn some real money — with real meaning. Easy breezy.
Team D and I are so thrilled that our work can create abundance for others. It's a very big bonus that I didn't see coming in my work. I was just going to write books and paint… and then this happened — a whole new level of affiliate love and cash for a lot of people… Read More
Destroy before you create. So scary. Sooo effective.
Half truths piled on top of half truths do not add up to the whole truth. tweet It's a just a heap of "not-quite-there-ness." Grey. Cluttery. If you want satisfaction you have to… Read More
What it takes to make good stuff in the world. Creativity, cadence, core desired feelings.
CREATIVITY. CADENCE. CORE DESIRED FEELINGS. What it takes to make good stuff in the world. Notice when your dream has come true. If you're addicted to accomplishing things, you might miss the fact that you've actually accomplished something pretty awesome… Read More
Desire Map Licensing is open for YOU. A business model for conscious prosperity.
The Desire Map book has been transformed into a teachable curriculum that can be delivered as live or virtual workshops, destination retreats, or even combined with other events and courses. It's called The Desire Map Licensing Program… Read More
I implore you: Do not dumb down.
I've been asked to dumb down. More than once. By bosses. By advisors. By magazine editors. And I have one line for that… Read More


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We encourage all prospective students to visit and tour Temple's law school campus and participate in one or more of our group information sessions. We have a variety of options available and you can choose the one which best fits your schedule and interests.
Open Tours
Tours of the Law School facilities conducted by our Student Ambassadors are available every weekday while classes are in session. Tours will leave the Law School Admissions & Financial Aid Office, Barrack Hall Suite 101, at 12 noon and you should plan to arrive at least 15 minutes ahead of the scheduled departure time. Tours will last until approximately 1:00 pm. Open tours for the fall 2015 semester run from September 21st to December 4th.
Prospective students may also take self-guided tours of the law school buildings. All visitors must come to the Admissions Office in Barrack Hall, Suite 101 to pick up a visitor's pass.
Group Information Sessions
Prospective student group sessions are available throughout the year. These include an informal information session with an admissions professional, a tour with a current law student, and during the academic year, a visit to a first-year class. These sessions are an excellent opportunity for students to learn more about the law school and the admissions process. If you are unable to attend a group session, individual appointments can be arranged in some circumstances.
Please contact the Admissions Office to reserve a space in an information session by calling 800-560-1428 or via email at email@example.com. All sessions begin on the first floor of Barrack Hall in the Admissions Office. We ask that you arrive at least 15 minutes before, if possible, so as not to delay the start of the program.
Fall 2015 Group Information Sessions
- Monday, September 21st 4:30 pm – 7:50 pm
- Wednesday, September 23rd 10:30 am – 1:30 pm
- Thursday, October 8th 4:30 pm – 7:50 pm
- Monday, October 12th 10:30 am – 1:00 pm
- Thursday, October 22nd 11:15 am – 2:25 pm
- Thursday, November 5th 11:15 am – 2:25 pm
- Tuesday, November 10th 5:30 pm – 8:50 pm
- Monday, November 23rd 10:30 am – 1:30 pm


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Black Friday deals are huge this year.
Below is a list of when stores and malls will kick off the shopping extravaganza:
Sears: 8 p.m. Thanksgiving Day
Toys R Us: 8 p.m. Thanksgiving Day
Wal-Mart: 8 p.m. Thanksgiving Day
Target: 9 p.m. Thanksgiving Day
Best Buy: midnight
Kohl's: midnight
Macy's: midnight
JCPenney: 6 a.m. Friday
Radio Shack: 6 a.m. Friday
If you want coupons click here: http://www.theblackfriday.com/
Tell us how you make out. We want to know what your deals are.
The Ultimate Black Friday Shopping Guide (2012)
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Multiple monitors? The Console has you covered! A full array of displays fits easily across its cockpit design, and wraps around you for comfortable viewing. Enjoy the health benefits of a standing desk: The keyboard shelf raises, lowers, and tilts with the squeeze of a paddle. Add Extension Tubes and an Additional Shelf for even more equipment above. Customize this cart to make it your own!
The wrap-around design of the Console is perfect for lining up multiple monitors for easy viewing. With an overall depth of 39", you can spread out your equipment with plenty of elbow room left over. The adjustable shelf holds more than just your keyboard. There's space for your tablet too!
With a gentle squeeze of the paddle, the manually-adjustable, spring-assisted keyboard shelf rises 7" above and 6" below the rear stationary surface it attaches to, for a variety of sitting or standing positions. (The rear stationary surface can be installed at any height in 1" intervals along the legs.) The front surface also tilts 15° negative (away from you) or 9° positive (towards you) to ease wrist strain. Extremely sturdy, the front keyboard shelf holds up to 40 lbs.
Double or even triple the space of your workstation with Additional Shelves and Extension Tubes (available separately). Position (and later reposition) the shelves during assembly by installing in holes at 1" increments along the vertical legs. Attach a CPU SideRack on the inside or outside legs of your Console for quick accessibility. See all of our accessories to configure it just the way you want.
With an expansive 5' wide surface and an overall weight capacity of 150 pounds, the sturdy Console easily holds many monitors and lots of heavy equipment. Durable shelves are made with 1" thick, industrial-grade particle board and scratch-resistant laminate. And note the large casters!
Six 4" rubber casters make the Console easy to move, whether to access cords at the back of the cart or to roll it around in a room. With the front two casters locking, the workstation stays put. Need more maneuverability? Upgrade to robust 5" casters.
Organize and protect your cords and cables with the included cable management kit: 3" cable alley, 9" cable alley, cord handler, and fasteners to attach alleys to cart legs. (Another nice little touch: We even supply the tools to put your Cart together!)
Download Detail Dimension Drawing (pdf)
By Chuck@CSUSanMarcos
from San Marcos, CA
Comments about Console Cart:
This has been the main desk in all of my offices since 1996. Yup, twenty years next year. Still going strong. Take off the keyboard shelf and fits thru a standard door. Great product that I have recommended to lots of folks in IT around here. Getting ready to buy another.
By Grumpy Ole Dude
from Martinsburg, WV
I convinced my boss I needed a desktop that would hold two to three computers and I got a black Console Cart. Finally I had everything I needed to get my job done. I used this cart for five years and when I retired I tried to get them to let me take it home. No suck luck. I bought a grey Console Cart for my home over 4 years ago and I wouldn't sell it for any price. Better than sliced bread! I use it daily and it is so comfortable my wife has to pull me away. Try it and you will love it too. Trust me on this one.
By SCAPsinger
from Dacula, GA
Purchased this desk on the recommendation and review of a fellow video producer who had outfitted his entire facility with the previous generation of Fit Console workstations. Was looking for a new desk that would accommodate multiple monitors, a multitude of external hard drives, video playback decks and similar equipment. I was also intrigued by the many sit/stand desk options out there in the market, since sitting down for hours on an editing job can be exhausting.
I received the shipment a month or so before I was ready to build out, and so the rather large bundle of desk plus add-ons took up a good bit of space (I was anticipating a longer fulfillment time and earlier installation, so it's my fault it sat for so many weeks). The first thing I noticed were the thorough instructions and the included tools - everything that would be needed to put the desk together.
All told, installation was very simple and only required a 2nd person to help flip the desk over after assembly. The single biggest hassle, however, was the keyboard desk mechanism, as the desk includes the sit/stand lift mechanism but the included instructions only had installation guidelines for a tilting keyboard desk. The other instructions were not included in my shipment. After searching around the Anthro website I found some instructions for another cart Anthro sells that had my mechanism listed, and it got me about 90% of the way there. I still ended up with some extra cable restraint pieces that appear to not go anywhere and weren't listed in the other instructions, but it seems unnecessary to me at this time.
I added the standard monitor mounts for a 2 monitor setup, and they clamped on fairly easily. In all, I'm very satisfied with the desk itself, and only wish that it hadn't taken me almost 2 more hours to find and figure out instructions on installing the keyboard shelf.
HIGHLY recommend, it's absolutely worth the price, this desk will probably be in use for 10-20 years.
By Darcy
from New York, NY
For the past 10 years I've used this in a variety of environments and configurations and in a variety of different roles. As design director in a video broadcast firm with multiple computers with multiple monitors, with a designer by my side and clients over my shoulder (i had two 24" Sony CRTs and two sony video monitors on this thing! Oops. I just "aged" myself.) ... As an artist in a mixed media studio doing video projection and animation... As a designer picking up work at a home office in my extra hours while raising a couple of kids... I've added shelves, I've changed the height of the work area to better accommodate variety of working postures - standing and wrapping myself around my cintiq 24hd while i doodle... I have high shelves on my fit console unit - don't put a large format photo ink jet printer on the top shelf - as the head goes back and forth this would send a slight tremor through the unit. Not disconcerting - just noticeable. The large printer was moved to a cart. I love this thing so much, I build my studios/offices around it.
This product rates a 2 out of 3!
Manual Adjustability
Easy partial adjustability – squeeze the paddle to quickly move or tilt the spring-loaded keyboard shelf when you want to change positions. Perfect for shared workstations!
See our full line of height-adjustable workstations and carts »
Read our blog to learn
informative product facts, user
stories, and featured staff biographies. »


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other popular MBA colleges in India include Symbiosis Institute of Business Management (Pune), Narsee..., skills and colleges offering education in management courses in India. Management is all... resources and financial resources are required to be managed in a way that optimizes the profits... of the organization in an efficient manner. In India, the growing demand for skilled managers has increased the scope... specialization in different areas like Humana Resource, Finance, Marketing, and International Business. It opens Read More...
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-taught person; I have studied business as well. The Journey so far. I launched my label `Anupamaa` in 2004...The festive seasons is just picking up in India. Wasting no time designer duo Shyamal and Bhumika... from Indian crafts and hand embroideries. Silhouettes give a true feel of free flowing Ghaghras... of an Indian touch- clutches inspired from Indian Gods like Lord Krishna and Shree Balajee`s eyes. Every... amalgamation of Indian art and tradition." Anupama Dayal with her extraordinary ability Read More...
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present days opportunities especially for women are more found in the financial services industry... insurance companies. and since then there are 2.5 lakh women employed in this industry. as a financial... advisor the women can become their own boss, start their business without any investment, flexiblity Read More...
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for underprivileged kids. This time Khushii brought together India`s leading style gurus at its first fashion inspired... Creations by 21 Indian designers, including Ranna Gill, Nikhil and Shatanu, Varun Bahl, Rohit Bal, Rajesh... are in need of socio-economic development. Samsaara, the luxurious abode of fashion, showcased the latest... brand Famous talks about the fusion of the traditional with the modern. Payal expressed, "I have... to the ensembles. Borrowing inspiration from the Indian sari drapes, the silhouettes are underlined with full body Read More...
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the person at the royapettah road office, we successfully located the place... First thing to rant about... of the day we just needed ten mins of his precious timeIf the person holds a post does it mean that he... can just waste others time like nobodys businessIf he is really busy and really cant spare Read More...
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of parliament is Meira Kumar. Furthermore, Delhi , the capital of India, sports a woman chief minister, Sheila... independent . No other country has as many women in politics at the senior level as India does.Women are given... right of property and right of inheritance in India. There are many women doctors , as a percentage..., in India than in many developed countries. We have another set of women excelling in writing though... to the common perception a large percentage of women in India work. In rural India, agriculture and allied Read More...
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I am a freelance creative writer, content business owner and i alsocreate online creative...,TIMES OF INDIA and many more providing them content for theirwebsites, being a consultant for many... onlinewriting workshop students are house wives from all over India who wantto enhance...oneself and the ability to break out of the mould without fear. My jobgives me more financial stability, more....So, if there is a home business you want to pursue, get the rightguidance and let nothing, absolutely nothing deter Read More...
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I am a student living abroad. I visited India last week(just for a week) for the first time since i... left India. While on my way to India,I had a mixture of feelings like how i would feel when i reach... India, meet my family and so on and so forth. But I was perfectly fine in India, did not miss any of my... friends,did not feel overwhelmed too by the family love and affection. The day i left India, my feelings... were neutral too :)After reaching here, I do not miss India or my family. Am fully fixed Read More...
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Do you know that India proudly ranks numero uno in road accidents in the world? Do you know... a child dies due to some domestic accidents?Every individual in India is well aware... consequences that may crop up as a bolt out of the blue. It is the basic nature of Indian minds to choose... awareness, alertness and practice. The most obvious proof for this is the fact that our India proudly ranks... creatures on the Indian roads... with these mad share autos and the government buses they feel Read More...
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that she would be coming to India next week on a holiday with her husband and children and as usual would...,in various stages of study.I was always a simple person,I never cared much about wearing different... look as if I had a huge wardrobe.Iwas a seedha sadha person.But Suseela was different.She liked... as a reply,Rama?Susee voice broke.She said,Bharathi turned to me and said,Mind your own business,Rama.You can....Then I said,Well,Susee,There is not much difference between Mind your own business and Do not interfere Read More...
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Which shampoo brand you prefer the most for your hair?
|Matrix Biolage
|3% | 4 votes
|Dove Nutritive Therapy
|22% | 29 votes
|L'Oreal Paris Total Repair
|26% | 34 votes
|Sunsilk Co-Creations
|40% | 52 votes
|Fiama Di Wills
|6% | 8 votes
|Total Votes: 127
|View all Polls