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Disney-ABC Domestic Television has formally renewed Katie Couric's freshman syndicated talk show Katie for a second season. The show has been renewed on its existing stations in over 95% of the country, including the ABC Owned station group, which already had a two-year deal for Katie. For the 19th straight week since its premiere (through the week of January 14, 2013), Katie continues to rank as the top freshman syndicated talker in Households, total viewers and women 25-54. Among all 18 syndicated talkers, Katie is No. 6 in households (1.9 rating), Total Viewers (2.497 million) and Women 25-54 (1.0 rating). On January 24, Couric's first on-camera interview with Manti Te'o and his parents (2.6/7) jumped 30% over the show's prior 4-week average. Katie joins fellow freshmen Steve Harvey, Trisha Goddard and Dish Nation, which also have been renewed for a second season. The formal greenlight for Katie comes days after original executive producer Jeff Zucker left to run CNN (He retains ownership in the show). Couric now executive produces the show with Michael Morrison, who joined late last month. Co-executive producers are Kathy Samuels and Michael Bass.
'Katie' Officially Renewed For Season 2
15 Comments
At this moment, Katie is gratefully Tebowing for Manti Te'o
Katie lost all relevance in her final years on The Today Show and after leaving. This is another Zucker nought.
Happy for Katie. A well written intelligent show. I watch everyday.
so you're the one.
I agree, Chris, but this is no big news. The multi-million dollar deal was for a two-year commitment. She would have been paid even if the ratings tanked.
She is unwatchable
I prefer Dr Oz Show or Steve Harvey show
Wow. You are now officially the spokesperson for the lowest common denominator.
Cutie-pie Katie gets renewed, Palin gets dumped by Fox News. Yup, it's been a good week.
this despite horrfic ratings, 40% below lead-in, 25% below year ago time period, zero growth, not to mention this is unwatchable.
So do something else. Turn the TV off; take a walk, read a book. Getting ready for work and can't pick a book up to read it? Turn on an audio book. Listen to a Ted Talk on Youtube. If you can't survive without knowing what's happened between the evening news and 7 a.m., there's this cool invention called the radio and it has some very informative news programs and interviews with interesting people.
I apologize for the edgy tone. My daughter stopped by last night to show us her new IPad and the different apps. On Entertainment, she showed us clips from a reality show about a spoiled "Beauty Queen" baby that people watch so they can say how horrible she and her mother are! Edward R. Murrow warned us not to misuse the potential of television. THAT ship has sailed!
When people are surprised I haven't heard of a new show, I explain we ended TV in our home two years ago. My husband was watching four hours of MSNBC, CNN & news "specials" a night. He was driving us all insane so it was either that or foul play.
I plead guilty as well. Turning the TV just to see "what's on" and settling on whatever will "do"; usually a re-run of "Law & Order: "AnyVersionWillDo".
We have Netflix, Amazon Prime and loads of great stuff on Youtube. I belong to a few forums of special interest to me. I am re-learning French, reading The Jewish Daily Forward free on email, checking in on The New York Times (is it just me or are others suspicious they are cheaply getting Sophomores from Columbia who missed school the day they taught grammar and the difference between hard news and editorials?). Anyway, I learn something new every day. My husband and I listen to music again many evenings on these neat things called records. Oh, and we read these paper things called books, too. I had almost forgotten the feel and sound of a page turning.
I'm not being elitist or snotty, really. We decided to try it for a few months and found we'd made a lucky choice. It made us both see how precious time is and how many great things there are to do with it. Also how lowering the noise level in your home will actually make you FEEL healthier.
Sure, I miss a few things. I would have liked to see the Royal Wedding AS it happened, but I knew I'd get to see pics of the most important part; the dress. And I've already invited us to my daughter's house for the Oscars and the Kentucky Derby. People might be very surprised to see just how much they WON'T miss.
So if you find Katie "un-watchable", turn her off, think outside the "Box" and give our way a try for a few months. You might be happily surprised at what you find.
It looks like she's phoning it in. I used to like her. Not any more. It just feels like she's doing it for the money. Just not invested.
I wonder how many watching, also watch the other stupid shows out there….The Chew, The Taste, The Bachelor/Bachelorette, Bachelor Pad, Dogs in the City, The Glass House, and so on and so on…..pitiful….
She had a two-year guaranteed deal with most of the stations that air this show when it was first proposed a year ago, so this wasn't a surprise announcement at the least but hoping next season will be the last season of this trashfest of a show with an irrelevant host at the helm.
ABC has Katie riding the coat tails of General Hospital, GH has great ratings… You see when you don't turn the Channel fast enough and the Witch comes on, she and ABC get big ratings from the GH viewers….. Auto tune your TV so it will go to ANY other channel before this happens! My Tv is auto tuned and ABC never gets a rating from me. I loved my soaps, not more stupid talk shows…. not everyone likes more boring talk shows and the Stupid Reality shows!!! Give me The Walton's and Touched by An Angel reruns any day over what ABC SAYS! "VIEWERS WANT REALITY" HEY ABC HOW DO YOU LIKE THE SOAP VIEWERS REALITY???
The promos are awesome.


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Jordann and Kayla-La
13,149pages onAdd New Page
this wiki
this wiki
|This character exists primarily or exclusively within the Pre-Super Genesis Wave continuity.
Information in this article may not be canonical to the storyline of the games or any other Sonic continuity.
Jordann and Kayla-La were the great-great-grandparents of Edmund and Dimitri.
|This page was either created or contains content from another article at Mobius Encyclopaedia. When rewriting sections, remember to adhere to our Manual of Style.
Kayla-La was the scientist who discovered that the White Comet would impact Echidnaopolis. Together, the two managed to devise a method to lift Echidnaopolis and part of the surrounding landscape into the sky with twelve Chaos Emeralds, thus making them the founders of the Angel Island.
- Their names appeared in early issues as "Jor-Dann" and "Kayla" before the cultural naming pattern was decided upon, and were retconned afterward.
- The Complete Sonic Comic Encyclopedia mistakenly refers to Kayla-La as Prahb-Jo.
|This article or section about a character is a stub.
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Has Ovechkin picked a company yet?
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Registered User
Join Date: Nov 2010
Posts: 774
vCash: 500
Originally Posted by
Again without adding up all the contracts you can't say anything for certain.
It's not even my main point. They are great players, but who cares what they use. That's not why they're great. Your skill level, height, weight, foot shape, budget etc, etc, has nothing to do with theirs.
This is no different than drinking a Coke or Pepsi because Michael Jackson does etc. etc.
Not trying to argue though. This stuff is all in Marketing 325. Of course it's going to disagreed with in a hockey forum.
Crosby/Ovechkin will make you and me look foolish in a pair of $50 Bauer turbos from 1987 bought off ebay. If he endorsed them people would buy them. The skates are junk. See my point?
Agreed. It's ALL about marketing. The big name companies are paying the stars $3-4 million a year to use their gear because they generate more than $3-4 million dollars of sales from us, the consumers.
From the companies' standpoint, signing a superstar like Ovie will be like striking gold. It's all about increasing their market share from sales.
For most people out there, they will make their purchase because a certain "superstar" wears that brand. It's a psychological effect to think that a certain product is superior than the other because the "pros" use it.
In reality, the gear used by the pros are of totally different quality compared to the consumer products. I.E. pro-stock. The gear used by the pros are completely custom made whereas the consumer level products are mass-produced. We won't get the extra-edge the pros are getting in their equipment in ours. The product could be still better but the margin isn't as big as the companies are making it seem.
Another good example of this are "disguised" sticks. Pros may use older model sticks and even think they may be superior to the new models but are forced to disguise their sticks due to marketing. For the million of hockey viewers out there, newer is better so they will go out and buy the newer version for $50-$100 more because they saw the pros use it.
Ovie will choose the best "equipment" that suits his game, but don't be native in thinking that half the reason wasn't due to endorsement from the company
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Download document:
Already v. Nike: the Supreme Court finds covenant not to sue made competitor's claim for invalidity of trademark moot - .PDF file.
The US Supreme Court issued its opinion yesterday in the closely-watched case, Already, LLC v Nike, Inc. In a decision that is almost certain to affect patent owners as well, the Court unanimously affirmed the Second Circuit's opinion and ended the speculation of commentators that trademark holders might no longer have the ability to dismiss an invalidity challenge by entering into a covenant not to sue. The Court concluded that an appropriately framed, broad covenant not to enforce a trademark against a competitor moots the competitor's action to have the trademark declared invalid.
Click on the link above to download this Morrison & Foerster briefing.


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Adore: Classical and Romantic World Guitar
Nominated for Best Acoustic Guitar Album OWM Music Awards 2016
Classical , World Fusion, Spanish Guitar
Cádiz by Isaac Albéniz, from the Profundo album
"Adore is the much awaited album by Joseph Sullinger, it may have been several years since Earth Voyage, but this has been worth the wait. Adore is a smooth delightful companion of compositions, that is both masterfully played and brilliantly produced. As always anything Joseph Sullinger touches turns to musical gold. A guaranteed winner with lovers of the acoustic instrumental genre and those of us who just adore, really good music, played with style and heart."
Joseph Sullinger
"In 'Adore' Joseph Sullinger has created a unique blend of classical and romantic guitar, which allows each of the slightly ethereal pieces to flow gently into the soul, in a style that is as timeless and pleasurable as the setting sun in the evening, the rhythmic ebb and flow of the ocean, the all-pervasive knowledge that music is the very essence of life."
Janet Mawdesley - Blue Wolf Reviews
New Album
Profundo: Romantic Music for the Spanish Classical Guitar
Coming Summer 2017
"Alluringly romantic and beautiful, "Adore" will appeal greatly to those who are particularly fond of classical and Spanish-style guitar music as well as new age world fusion!"


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Members of the armed forces were honored at Saturday's Colonial Cup with all servicemen and women receiving special recognition as part of "Military Appreciation Day" at the track.
Kershaw County Council received another refresher course in county government Thursday as part of a series of information work sessions scheduled by Kershaw County Administrator Vic Carpenter.
There was a strange sight during last year's Outback Bowl in Orlando as the Florida Gators went up against the Penn State Nittany Lions.
Weylchem, a chemical company with U.S. operations based in Elgin, will open a new production line this month for ammonia borane, a substance used to power fuel cells.
Before an audience at James Leroy Belk American Legion Post No. 17, U.S. Army Col. William "Mark" Kehrer spoke on the importance of not only the nation's veterans, but also their families. Kehrer spoke during the post's Veteran's Day observance Friday in Camden.
Kershaw County Council decided to move forward with two fee-in-lieu-of-taxes (FILOT) agreements at Tuesday's meeting despite various questions and concerns raised over the deals.
To this point in the 2012 election cycle, the Republican contenders have been all bark and no bite when it comes to the nation's energy future. President Obama's approach, however, hasn't been much better.
Kershaw County will soon have greater representation on the Santee-Lynches Council of Governments (COG), a multi-county organization that works toward economic development and collaboration in Clarendon, Kershaw, Lee and Sumter counties.
Q: Where are you from originally?
Kershaw County Council will consider fee-in-lieu-of-taxes (FILOT) agreements between Kershaw County and two companies that could help solidify an industrial park agreement with Sumter County.
The "Little House," a nearly 200-year-old home thought to have been built by Bonds Conway, the first black man on record to have purchased his own freedom in Kershaw County, received a special dedication Tuesday recognizing the site's historic restoration and cultural value to the community.
Andre Bauer must be looking for a second life in S.C. politics. Or more precisely, Andre Bauer must be looking for a second shot at a second life in S.C. politics.
The Kershaw County Recreation Department will hold a public forum at 6 p.m. Thursday on a proposed lighting project at the Kershaw County West Complex ball fields. The forum will take place at the Lugoff Fire Department, 892 U.S. 1 South, Lugoff.
Kershaw County Council decided to take a wait-and-see attitude at its meeting Tuesday and postponed a vote to establish an emergency ban on "bath salts" and synthetic marijuana.
Page 19 of 25
The Finally Friday Free Concert Series continues at the Fine Arts Center of Kershaw County (FAC) on Friday, May 26 with local favorite Landslide. The ...
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Gold Country Inn
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- Microwave in room
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- Microwave in room
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TripAdvisor Reviews Gold Country Inn Placerville
Travel Blogs from Placerville
OK, we're starting on another one.
We are leaving tomorrow(Thursday) morning. This is a driving trip. We are taking the Infiniti because its much more comfortable on a 4500 mile trip. We are driving to Sedona, Az. for the second night of the ...
I am trying out this app
... to copy and paste from the entries. I can copy them into word or another program so I can use them in other formats later
- Is the iphone app easy to use?
Yep. Super easy (once I figured out how to sync- why was that so hard to figure out??)
- How easy will it be to add photos?
OK it was very easy to add a photo. I can add on my mac or on my phone - just like uploading to Facebook.
- Will ...
Getting ready!
... Thank you Tanya!
What a wonderful dream come true! We are embarking on this fabulous trip to see where our ancestors came from. We are hoping to meet a few. One of our many reasons for picking the month of September is that the seafood is supposed to be spectacular!
We stay at Eden Vale Inn, and catch the gold bug
... got its nickname without the "dead guy" hanging from a noose off the side of one of the buildings. Mark said we couldn't miss the hardware store. I wasn't sure why but after we went inside I can see what he meant. And, happy accident, we needed to buy a flashlight for Roxy. This is gold territory so when we'd had enough window shopping, we headed 1 mile up the road to the Gold Bug Mine, and 1888 hard rock mine that is listed on the National Register ...
Days 253 - 257: Livin' the American Dream
... the car we returned to the Hawkins for pancakes. As we were cooking, we learnt how to master the art of cooking American bacon. I'm unsure that we succeeded, but we tried. An hour later we were out the door for a day on the rapids - the South Fork of the American River.The rapids on the South Fork are "rock garden" style rapids: meaning rocks in the channel create the rapids. Large pools follow each rapid, allowing time to regroup.
We couldn't be in better hands. Nicko, Sami and ...


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We get our pork from suppliers from Indiana and Michigan. As soon as the meat is received at Bob's, we cut everything, vacuum seal it and freeze it. This ensures that everything is the BEST quality you can possibly get.
Incidentally, NONE of the products at Bob's are saline injected, unlike the big box stores, most of which have a 8-20% saline solution added to there meat. So, at Bob's, when you pay per pound, you are not paying for saline solution. Plus, since our meats are vacuum sealed and not on Styrofoam trays sitting in its own blood, it you're not paying for all that extra packaging either.
As a result, we actually are LESS EXPENSIVE and BETTER QUALITY MEATS then at the big box stores like Meijer or Walmart.
LOCAL – HIGH QUALITY – INEXPENSIVE Pork
We keep boneless pork loins on hand to make butterfly chops (a boneless pork chop) and Americas cut chops (a thicker cut boneless chop). We can also prepare a boneless pork loin roast for you either with our without our special seasonings.
Also available are pigs for pig roasts as well as half or whole pigs all processed for you. Processed pork includes: pork chops, pork steak, pork roast, tenderloin, pork cutlets, ham, bacon, Canadian bacon, fresh or smoked picnic spare ribs, back ribs, fresh hams, pulled pork and side pork. Just call us up, tell us if you want a half or whole pig and how you would like it cut. We take care of the rest!
Orders can be picked up at Bob's Processing or at the Sugar Bear in Paw Paw


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Toronto police have recovered two types of pills that may be linked to the death of a 22-year-old man and a 22-year-old woman at Toronto's VELD Music Festival.
Last weekend, a man and woman both died after ingesting what police are calling "party drugs" at the two-day music festival. Thirteen other people who attended the festival were taken to hospital after feeling ill from ingesting the drugs.
According to Detective Sergeant Peter Trimble, one of the drugs is described as a small brown pill, while the other is said to be a small clear capsule with a white substance. Tests on the pills are pending.
"There have been two postmortems completed on our two victims," he said. "Neither victim had any anatomical cause of death and the cause of death right now is pending toxicology."
The youngest of the 15 people who fell ill as a result of taking the drugs was 16 years old. According to police, the average age of the patients was 19 and most of the distress calls were placed late Saturday night leading into early Sunday morning.
The two pills were given to police after they issued a public appeal for attendees to turn over any drugs they purchased at the festival.
Det. Sgt. Trimble said it is unknown whether or not the victims ingested the same type of drug because in many cases "they didn't know what they were taking."
"Some [of the 13 people] remain in hospital. I understand they will all survive and make full recoveries and some have already been released," Det. Sgt. Trimble said, adding that some of the victims suffered seizures. "Some people were taking upwards of 10 pills, some people were picking up pills on the ground," he said.
Toronto police's homicide squad was called in to investigate the deaths because they were a result of criminal activity that caused harm to more than a dozen people.
During a tour of a Toronto Community Housing building on Monday, Mayor Rob Ford told CP24 that the deaths are "traumatic," but said he would continue to support concerts and festivals in the city.
"It's up to the homeowner, or the permit owner, or whoever," he said. "If you're having a party at your house and someone dies at your house, it's your responsibility. It's the permit-holder's responsibility."
Charles Khabouth, the founder of INK Entertainment and one of the festival's organizers, extended his condolences to the families of the victims in a statement and said that "safety and security have always been the number one priorities of the VELD Music Festival."
Police say that upward of 30,000 tickets were sold for the event, which took place Aug. 2 and Aug. 3 at Toronto's Downsview Park, and are asking anyone with pictures or videos from the festival to come forward or upload them to tps.on.ca/veld.
With reports from Sahar Fatima and Ann Hui
Report Typo/Error
Follow us on Twitter: @GlobeToronto


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Cases of new onset psoriasis in patients receiving TNF inhibitors for the treatment of rheumatoid arthritis (RA) have been reported; however, case reports do not permit the calculation of drug-associated incidence rates. Here, Harrison et al (Ann Rheum Dis 2009; 68: 209) compare incident psoriasis in TNF inhibitor treated vs. untreated patients enrolled in the […]
Mechanisms of the Atheroprotective Effect of Methotrexate Studied
RA patients who use methotrexate have been shown to have lower cardiovascular mortality compared to non-users; however, the mechanism underlying this protection is unclear. Here, Reiss et al (Arthritis Rheum 2008; 58(12): 3675) explore the atheroprotective effects of methotrexate in an experimental model of atherogenesis. Methods THP-1 monocytes/macrophages and human donor peripheral blood mononuclear cells […]
Immune Tolerance Induced by Chicken Type II Collagen as Treatment for RA
Type II collagen is abundant in the joint and is a major target of immune-mediated damage in RA. As suggested in studies of experimental animals, induction of immune tolerance at the level of the cartilage may be a way of reducing the signs and symptoms of RA and prevent joint destruction. However, whether oral administration of Type II collagen is an efficacious treatment for humans with established active RA has not been established.
Denosumab suppresses erosions but has no effect on articular signs and symptoms in rheumatoid arthritis
Bone erosions in response to inflammation in RA are driven by osteoclasts which are in turn activated by RANKL (Receptor Activator for Nuclear Factor κ B Ligand) binding to its receptor RANK. Inhibition of RANKL has the potential to retard bone erosions in RA patients, thereby limiting progressive joint damage and destruction.
Does Postmenopausal Hormone Replacement Affect Rheumatoid Arthritis Outcomes
Estrogens are known to modify immunologic responses and the modulation of rheumatoid arthritis (RA) disease activity during pregnancy is well documented. However, whether physiologic replacement of estrogen and other hormones after menopause adversely affects RA outcomes is controversial.


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Shrimp with Whole Wheat Couscous and Yogurt Hummus sauce
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Mountain Mike's Deer Positioner Skull Hanger
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Love em or Hate em...The #IceBucketChallenges Are Worth It
Photo Credit: YouTube
There are a lot of people hating on the #IceBucketChallenge videos. Even people in the WPOC office have been whining about them. One person in particular has complained non-stop but I know for a fact they have NEVER watched a single one.
But I digress
For all of you that are sick of watching people dump buckets of ice on their heads, this is for you.
Since the #IceBucketChallenge started, over $20 million has been raised for ALS research. You read that correctly. That's an amazing accomplishment and it will provide so much for those who are currently struggling with the disease.
So before you judge and make snarky comments on people's facebook pages, know that people like Anthony have hope for a better future because of all the money raised. Watch his story below.


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Kraus GVR-204-RE_Base Irruption Blue Rectangular Glass Vessel Sink with PU
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text: 11/22/16 12/06/12 POPSUGAR Living Holiday Etsy Holiday Gifts 10 Etsy Gifts That Lend a Handmade Touch December 6, 2012 by Emily Bibb 181 Shares Chat with us on Facebook Messenger. Learn what's trending across POPSUGAR. These handmade Recycled Newspaper Ornaments ($49 for three) are a modern twist on the Sunday "funnies." This ornament makes a unique gift that you can hang on a tree or fashion into garland. These handmade Recycled Newspaper Ornaments ($49 for three) are a modern twist on the Sunday "funnies." This ornament makes a unique gift that you can hang on a tree or fashion into garland. Recycled Newspaper Ornaments $49 from etsy.com Buy Now Give this Handmade Soap Gift Box ($50) as a set, or wrap the soaps individually for a thoughtful hostess gift or warm welcome for holiday guests. Handmade Soap Gift Box $50 from etsy.com Buy Now This 2013 Letterpress Calendar ($26) is a stylish desk accessory and an affordable gift that any coworker will appreciate. 2013 Letterpress Calendar $26 from etsy.com Buy Now A handcrafted Dark Wooden Tray ($35) is a versatile gift because of its many uses. Add a candle or a few gourmet cheeses for an added touch. Dark Wooden Tray $35 from etsy.com Buy Now A fun take on the mason jar trend, this set of hand-poured Beeswax Candles ($150) will make a tabletop statement and the perfect decor gift. Beeswax Candles $150 from etsy.com Buy Now Fill this Personalized Birch Bark Wood Vase ($20) with fresh flowers for a thoughtful housewarming or hostess gift. Etsy Vases Etsy Personalized Birch Bark Wood Vase Rustic Home Decor (item E10290) $19.99 from Etsy Buy Now See more Etsy Vases Everyone's favorite, a Red Berry Holiday Wreath ($59) is sure to impress and be cherished well into the holidays. Red Berry Holiday Wreath $59 from etsy.com Buy Now These Hand-Printed Cotton Napkins ($20) are a great gift for friends who love to entertain or host holiday get-togethers. Hand-Printed Cotton Napkins $20 from etsy.com Buy Now Fill this Hand-Painted Ceramic Bowl ($35) with homemade treats or pinecones for a rustic Christmas gift! Hand-Painted Ceramic Bowl $35 from etsy.com Buy Now Printed by hand on natural cork, these Mojave Letterpress Coasters ($20 for four) make a fun, quirky gift for drink enthusiasts. Mojave Letterpress Coasters $20 from etsy.com Buy Now Share this post Decor GiftsEtsyHoliday GiftsHolidayHostess Gifts


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If that top one disappears, don't come looking for it at my house! It won't be here! So . . . red beard is truly only two generations away from Immortality. Good to know.
Oh . . . found a touch of pollen on Imm. a couple of days ago. Didn't have a chance to use it, but it was there.
Betty W.
-----Original Message-----
From: Linda Mann <email@example.com>
To: iris-photos <firstname.lastname@example.com>
Sent: Sun, May 8, 2011 7:58 pm
Subject: [iris-photos] HYB: red bearded whites
Couple of sibs from (IMM x little orange seedling) X DECADENCE
DECADENCE hasn't mixed as well as I hoped with the IMM etc lines, but
getting some nice colors.
Linda Mann east TN USA zone 7
where bloom is finally winding down and I'm horrified by how many pods
are out there!


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The men's restroom did not have enough shit tickets to clean my ass!.
by dronp October 30, 2010
by Napleo June 09, 2009
Cleveland Steamer Plush
The vengeful act of crapping on a lover's chest while they sleep.Buy the plush
A term used when you run out of toilet paper or, halfway through a shit realize that there is none there.
DAMMIT!!! I'm out of SHIT TICKETS again!
by Slashedace April 04, 2011
Another name for toilet paper
by jonhboy August 02, 2014
"That guy keeps pissing on the floor in our bathroom and leaving his shit tickets in our trash can. Who the fuck does that; takes a shit and throws their disgusting shit tickets in the trash after? That's pretty fucked up."
by Department of Behavioral Hell August 23, 2015
Cleveland Steamer Plush
The vengeful act of crapping on a lover's chest while they sleep.Buy the plush
Another word for Toilet Paper. Also known as "Asswipe" (pronounced As-weep-eh)
Mark (from inside the bathroom): HONEY! GET ME MORE SHIT TICKETS!
by Shabangshwa95<3 February 13, 2009
Anything that is good in a homeless man's imagination
by Greg stegs February 02, 2010


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Falana, whose testimonial is contained in the current edition of ALERT, a monthly pull-out by the EFCC, pleaded with his colleagues to be wary of briefs from politically-exposed persons (PEPs).
July 30, 2016
How A Governor Tried To Use Femi Falana To Launder N405million
Falana, whose testimonial is contained in the current edition of ALERT, a monthly pull-out by the EFCC, pleaded with his colleagues to be wary of briefs from politically-exposed persons (PEPs).
12 comments:
- Jeff Kingsley Jr said...
We need more lawyers like dis trust me
- 30 July, 2016 19:44
- samaila sulaiman said...
Nice one
- 30 July, 2016 19:48
- maryam sadisu said...
Mr falana good moves there! Your reputation is already speaking for you.
- 30 July, 2016 20:14
- Harbolarkale Niyi said...
Chief Femi Falana is a well respected and known personality. I respect the man alot
- 30 July, 2016 21:05
Some people with pocket size brain would have fell for such chicken change that would have soil their reputation, if you stand for nothing you will fall for everything.
- 30 July, 2016 21:52
True talk
- 30 July, 2016 22:13
- Adeniyi Adedotun said...
Sir can you please get us the little clue of this past governor it might be one those we're giving kudo's to outside there, at least a little link that a reasonable person could know this might be the governor in question.
Once again Sir, thanks for the love you have to this grate country of ours may God continues to streinghtning you.
- 31 July, 2016 06:39
I will always respect you but I don't know while PMB did not appointed you
- 31 July, 2016 09:31
Sincerely we need more men like falana, he stands true 2 his words, wuh he believes in n he stands firm, majority of d lawyers out dere waver all d time cos of greed, selfishness, lack of integrity n lust 4 moni..
- 31 July, 2016 13:55
Lovely! We need more trustworthy personalities to make this Nation what it is meant to be.
- 31 July, 2016 17:06
thumbs up baba!
#Olu, can you see?
- 01 August, 2016 05:47
- SPACO said...
Thank God we have a no-nonsense lawyer.
- 01 August, 2016 09:06


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Jaguar XF Police Cruiser is on the Beat
It says something about our society when we get all excited about a new police car. Jaguar wades into this hotly-contested arena in the UK — sorry Alabama! — as one of the sort-of cool police cars.
You know, an atypical police vehicle like the Hyundai Genesis, Lexus IS-F, Lamborghini Gallardo, and so on. In the UK, police aren't as infatuated with high-speed pursuits, so this Jag is a diesel.
Seven are on the beat right this moment, in Staffordshire, Mercia, and the West Midlands — and other than the day glo stickers, light bar, and "Home Office" electronics, they're standard.
Just for fun, click on to see a clip of a UK police show — things really are different over there.
GALLERY: Jaguar XF Police Cruiser
[Source: Autocar]
Get the Flash Player to see this player.


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It only takes a minute to get started. Fill out our simple sign up form and login to the website.
This step is discretionary but will seriously raise your chances of getting noticed by other people on the site. Introduce yourself, your pasttimes and upload your best photograph.
Use our website's advanced search to locate members who suit your criteria. Or just scan through listings and locate daters that you like.
Located someone who could be a good match? Drop them a line or just a wink and see what they say. With a little luck you'll be on your way to locating that special someone for you.
It only takes a few minutes


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Are you a webdesigner or developer? You might want to download and take a look at the Internet Explorer 8 Beta 1 preview. You'll be able to see the great new features that IE8 brings to the table and take advantage of them to enhance your websites, with the benefit of making them ready for when IE 8 launches!!
My teammate, Dan Stolts, has just completed a three-city road show where he did presented on IE8. I thought his blog post on his favorite IE 8 features was pretty interesting.
I'll be installing the beta tonight. J


text:
Tired of useless tips?
Automated exception search integrated into your IDE
no such entry: "PowerPoint Document", had: [Object7, SummaryInformation, Object3, Object4, PP40, Object5, Object6, Object1, Object2]
This site uses cookies, as explained in our cookie policy. If you agree to our use of cookies, please close this message and continue to use this site.


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Cancer of the breast is the most well-known disease with women worldwide. Certain danger variables expand the likelihood of a lady getting this sort of disease. This incorporates age, hereditary qualities, being overweight, and way of life. Taking contraception pills is additionally connected with having bosom tumor, however, this is not demonstrated yet.
How is bosom tumor treated? Patients with bosom tumor have a considerable measure of treatment alternatives. These medications are balanced depending to the sort of growth and the forecast of the patient. Treatment modalities ought to be talked about with your specialist and it could be radiation, surgery, chemotherapeutic medications, and hormone treatment. Normally a blend of this treatment modalities works best. Continue reading


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|Message view
|« Date » · « Thread »
|« Date » · « Thread »
|"Daniel Kulp (JIRA)" <j...@apache.org>
|[jira] [Updated] (AMQ-2471) Add fine-grained authorization to the web console
|Tue, 04 Feb 2014 16:10:11 GMT
[ https://issues.apache.org/jira/browse/AMQ-2471?page=com.atlassian.jira.plugin.system.issuetabpanels:all-tabpanel ] Daniel Kulp updated AMQ-2471: ----------------------------- Component/s: webconsole > Add fine-grained authorization to the web console > ------------------------------------------------- > > Key: AMQ-2471 > URL: https://issues.apache.org/jira/browse/AMQ-2471 > Project: ActiveMQ > Issue Type: New Feature > Components: Broker, webconsole > Affects Versions: 5.4.0 > Environment: For all environments > Reporter: Mark Gellings > Priority: Minor > Fix For: 5.x > > Original Estimate: 240h > Remaining Estimate: 240h > > The web console doesn't support fine-grained authorization at the moment. > http://old.nabble.com/Dynamically-setting-activemq-username-password-when-logging-into-web-console-to26118677.html#a26126782 > Scenario with a guest and admin user: I'd like guest to have read privs (see messages on queues, etc.), and admin to have read/write privs (see messages on queues, delete messages, delete queues, etc.). In our scenario guest is producing a message and just wants to verify the message has been created successfully on the queue. Admin owns the queue and the broker as they are on a separate development team than user guest. They do not want guest to be able to delete messages/queues etc. Right now we have no way to let guest see for themselves that the message is on the queue unless we give them the admin user/password for the basic authentication prompt when using the web console. If we give that out, we give out read/write privs to guest which we don't want to do. > I think for this to be possible two separate connections would need to be maintained to the broker, one for guest and one for admin so as the simpleAuthenticationPlugin and authorizationPlugin can be used based on the user/password used to log on. Ideally the user/password entered during a basic authentication prompt could be mapped to the same user/password used to connect to the broker. Maybe this isn't possible if the web console only maintains one connection to the broker. Maybe the web console would need to be enhanced with a user/group security section to control what privs in the web console the logged on user has. An admin could then control whether a user has the right to delete a message, a queue, etc. and the web console has the smarts to display the delete link or not based on the privs of the logged on user. -- This message was sent by Atlassian JIRA (v6.1.5#6160)
|View raw message


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Foilacar Industries Corporation
- Alamat Kantor
- No. 10 Brixton Street Brgy. Kapitolyo, Pasig, Philippines
- Industri
- Real Estate / Mortgage
Foilacar Industries Corporation has branched out into surfaced design specializing in creating imagery for interior design for hotels, resorts, offices, clinics, business buildings.Foilacar Industries Corporation is the #1 premium installers of vehicle wraps in the market. Be it Motorcycles, car, SUV s to marine yachts to airplanes & helicopters; we use the best materials and our specially trained technicians to ensure upholding the quality standards we are best known for.
Taking this technological know-how and meticulous work ethic; Foilacar industries is venturing into designing and foiling internal and external home and work spaces. We have a team of Creative Designers, ready and capable of designing our customer's Spaces depending on their style-preferences or personalized taste.


text:
Activated Carbon
€ 49,90
Lieferbar innerhalb von 2-3 Tagen
Oktober 2015
BeschreibungAcacia modesta wood have been collected from mountain of (Ramora) Dir lower Khyber Pakhtukhwa Pakistan and then converted into activated carbon. The sample was activated chemically. The sample was characterized by BET surface area, scanning electron microscopy, FTIR and energy dispersive spectroscopy techniques. Adsorption of some dyes (basic green 5 and Methylen blue) are studied on treated and raw sample of Acacia modesta carbon powder by kinetics and batch adsorption technique at different temperatures. Intraparticle diffusion, Elovich and Bhangam models were used for analysis adsorption kinetics studies. From adsorption kinetic data the thermodynamic parameters were calculated from the adsorption data. The result shows that the adsorption process is a spontaneous process.
Portrait1. Dr.Sultan Alam is Associate Professor in Physical Chemistry, University of Malakand Pakistan. 2. Dr. Noor-ul-Amin is Associate Professor in In-Organic Chemistry, AWKU Mardan, Pakistan. 3. Mr.Aslam Khan is research student at Department of Chemistry, University of Malakand, Pakistan.
Untertitel: Activated Carbon as an Adsorbent for Dyes Removal from WasteWater. Paperback. Sprache: Englisch.
Verlag: LAP Lambert Academic Publishing
Erscheinungsdatum: Oktober 2015
Seitenanzahl: 92 Seiten
Format: kartoniert


text:
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How much does an Online Lab Test cost?
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text:
Due to our ongoing expansion in the U.S. motorcycle market, Silkolene Powersports Lubricants, a division of Fuchs Lubricants Co., is seeking a Sales Representative to work with our distributor's Sales reps and their Dealers.
The ideal person must have an extensive motorcycle industry background, with experience in dealer sales and be prepared to travel. If interested, please send a resume to email@example.com.


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I go both ways also. OWB carry is the G36 in a Tucker HF2. An untucked polo shirt and shorts. Good to go.
Printable View
I go both ways also. OWB carry is the G36 in a Tucker HF2. An untucked polo shirt and shorts. Good to go.
One thing to check for, especially since you're use to IWB carry. Put the OWB rig on and try reaching up high like for something on a top shelf. OWB carry requires a longer cover garment at times like that. I carry a cane left handed (weak side) so any reaching (i.e. for groceries) is done with my strong side hand.
Yes, OWB CAN be more comfortable but that's not always the case. It'll depend on your belt, holster and gun you're wearing. I carry my Glock 19 primarily OWB but my 1911s IWB because of their thin profile.
Give it a try and see what YOU think.
Thanks for the replies. I think I'll give the OWB a whirl since I received one with the gun purchase. If I don't like it, I can always change back to IWB since I'm comfortable with that option.
I really just wanted to get some feedback from experienced people using OWB regarding comfort and concealment.
I love my IWB holsters but seriously... how could not stuffing a holstered gun inside your pants possibly not be more comfortable? :rofl:
OH NO Slomo is biholsterual!!!!:rofl:
I carry both IWB and OWB. IWB conceals easier. OWB is more comfortable and quicker / easier to deploy my weapon from.
I carried IWB for a bit but it didn't take me long to realize that IWB wasn't for me. I went for a basic OWB Galco COP 3-slot holster and haven't looked back since.
I've been carrying OWB the last few days around the house to get a feel for it... So far it's very comfortable and conceals well with an untucked button down shirt. Not to mention, I can tighten my belt to where it's supposed to be!
I'd try it outside the house if I could find some 40 S&W to load it. Ammo is pretty scarce in Knoxville at the moment it appears. Unless I want to pay $20+ per box at the local range.
Open carry is legal here in Tennessee...good thing...I vary between concealed and open carry a Glock 27 in my Don Hume 721 OT holster. I used to carry IWB when I was younger and lighter, but now that I'm older and heavier, OWB is much more comfortable.
Depending on your build you might try AIWB. I find it a lot more comfortable than the traditional 3:30 - 4:00 position yet not as comfortable as OWB. It is a very fast presentation position for those who care about such things. :yup:


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AndyPhunkm... Photographer
Angiegirl223Model Female Las Vegas, Nevada, US
Mayhem # 3341123
About Me***:09/01/16 I'm finally all healed from my car accident months ago and have been working out, getting in shape. I'm busy but I'm ready to shoot again. Thanks for your patience with me. -Angela
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My name is Angela and I'm a young woman hoping to be featured in magazines, catalogs and posters. I'm on the hunt for photographers who are willing to trade Time For Photos. No nudes please.
Born in the beautiful state of California and currently residing in Las Vegas Nevada, I'm of Native American ancestry. I am willing to work with other models as well and hoping to gain as much experience out of this as I can. Even though I have not been doing this for very long, it's been fun and I enjoy the photographers I've collaborated with. Please note, in addition to several tattoos I have braces on my teeth and they will be coming off soon. :)
IMPORTANT NOTE #1: If you have a legitimate photoshoot offer, please utilize ModelMayhem messages! Photoshoot offers left as TAGs & Image Comments may not be noticed. Thank you.
IMPORTANT NOTE #2: I would prefer to meet in a public setting BEFORE the actual photoshoot so we can meet, go over ideas & details and make sure we're both comfortable working together. Thanks again.
Have a project, idea, concept or question? I want to hear it! Message me and I will get back to you asap.
Thank you!
Have you worked with Angiegirl223? Add Credits for Angiegirl223! >>
Albuquerque Images
"Start Modeling again Angela!!! We need you! :)"
See Sample Worked together 2-10 times; Most recent: May 2016
Credit Notes*´¨)
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(¸.·´ (¸.·´ Photographer(s)·*´¨) ¸.·*¨)
Albuquerque Images Ltd MM #3337648
Craig Solomon MM #2388399
Snapstreet MM #3204735
Geekslair Photography MM #224086
EtherealLight MM #2554864
David Bardes MM #1957957
In progress:
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Blended Faces Makeup Artist
Grin Without a Cat Artist/Painter
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text:
- //Home
- //Music
- //Lowland Hum
Lowland Hum
Native Air
US: 6 Aug 2013
UK: 6 Aug 2013
North Carolina folk-pop duo Lowland Hum are, in some ways, exactly what you might expect from a married couple playing intimate folk songs. Their vocal harmonies are airtight and sweet, their songs shadowy but bolstered by hope and by a shared vision. But where so many folk acts tend towards the spare and the slow—hoping to drag emotion out of us one pulled note at a time—Lowland Hum's sound is a vibrant one. The vocals on "War Is Over" rise up and out of the track, letting it bloom instead of whispering in the corner of some coffee shop.
The two work well singing together, on the fragile yet lush space of "Albatross" or the faint stomp of "My House Is Empty", but they also sound great on solo turns. Lauren Plank Goans' voice is sweet and approachable, so unassuming you might miss its subtle range, while Daniel Levi Goans can hit with a gruff, confessional whisper or a honeyed howl. The best moments here are personal and bracing—"but the strong desire keeps my feet moving," they sing at one point—which make some grand statements ("I watch my generation sleep") feel a bit too broad in comparison. Luckily, they focus mostly on small details writ large in these songs, making Native Air a confident and impressive debut.
Fleet Foxes "Fool's Errand" (Singles Going Steady) // Music
Vince Staples "Big Fish" (Singles Going Steady) // Music
*repeat repeat "Plugged In" (video) (premiere) // Music
Slowdive Sell Out Brooklyn and Release Bonus Song "30th June"
// Notes from the Road
"Although sound issues delayed their set on the second night, Slowdive put on an unforgettable show in Brooklyn, or rather two shows.READ the article


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Lamellipodium extension and membrane ruffling require different SNARE-mediated trafficking pathways
Intracellular membrane traffic is an essential component of the membrane remodeling that supports lamellipodium extension during cell adhesion. The membrane trafficking pathways that contribute to cell adhesion have not been fully elucidated, but recent studies have implicated SNARE proteins. Here, the functions of several SNAREs (SNAP23, VAMP3, VAMP4 and syntaxin13) are characterized during the processes of cell spreading and membrane ruffling.
We report the first description of a SNARE complex, containing SNAP23, syntaxin13 and cellubrevin/VAMP3, that is induced by cell adhesion to an extracellular matrix. Impairing the function of the SNAREs in the complex using inhibitory SNARE domains disrupted the recycling endosome, impeded delivery of integrins to the cell surface, and reduced haptotactic cell migration and spreading. Blocking SNAP23 also inhibited the formation of PMA-stimulated, F-actin-rich membrane ruffles; however, membrane ruffle formation was not significantly altered by inhibition of VAMP3 or syntaxin13. In contrast, membrane ruffling, and not cell spreading, was sensitive to inhibition of two SNAREs within the biosynthetic secretory pathway, GS15 and VAMP4. Consistent with this, formation of a complex containing VAMP4 and SNAP23 was enhanced by treatment of cells with PMA. The results reveal a requirement for the function of a SNAP23-syntaxin13-VAMP3 complex in the formation of lamellipodia during cell adhesion and of a VAMP4-SNAP23-containing complex during PMA-induced membrane ruffling.
Our findings suggest that different SNARE-mediated trafficking pathways support membrane remodeling during ECM-induced lamellipodium extension and PMA-induced ruffle formation, pointing to important mechanistic differences between these processes.
Cell adhesion to an extracellular matrix (ECM) is a fundamental feature of multicellular organisms and is a critical component of development, tissue function and homeostasis. Adhesive interactions between cells and ECM are complex, requiring the regulation of integrin localization and activity, reorganization of the actin cytoskeleton and localized membrane remodeling. After engagement of an ECM substrate, integrin-mediated biochemical signaling induces the actin-driven formation of Rac1-dependent lamellipodia and/or Cdc42-dependent filipodia
Integrin-mediated cell spreading is accompanied by dramatic alterations to the shape and content of the plasma membrane, particularly at sites where new cell-ECM contacts are formed. These alterations include the extension of the membrane (formation of a lamellipodium) and the enrichment of integrins and F-actin structures in or near the protrusion. Much evidence supports the notion that intracellular membrane trafficking is required for the remodeling of the plasma membrane as well as the redistribution of integrins that occurs during cell spreading. Studies have identified roles for the GTPases Arf6
Within the membrane trafficking pathways involved in cell adhesion, how the final stages of the pathways are regulated is not well understood. Most known membrane traffic pathways culminate in SNARE (
In the present study, we report the first identification of a SNARE complex, SNAP23-syntaxin13-VAMP3, that forms in response to cell adhesion to ECM. Using ectopic expression of inhibitory SNARE domains in CHO-K1 cells, we demonstrate that the functions of VAMP3, syntaxin13 and SNAP23 are required for the formation of lamellipodia during cell adhesion and migration. We also present the novel finding that the SNAP23-syntaxin13-VAMP3 complex functions in the trafficking of β1 integrin from a sorting endosome to a Rab11-containing recycling compartment during the delivery of adhesive receptors to the plasma membrane. As well, an interaction between VAMP4 and SNAP23 was identified that was enhanced specifically during PMA-stimulated membrane ruffle formation. Inhibition of SNAP23, VAMP4 and the Golgi SNARE, GS15, but not VAMP3 nor syntaxin13, impaired the formation of PMA-induced F-actin-rich ruffles. The results suggest that cell adhesion is dependent upon SNARE function for ECM-induced integrin trafficking and lamellipodium extension, while a distinct SNARE-dependent pathway leading from the trans-Golgi network is required for the formation of PMA-induced dorsal membrane ruffles. Our findings point to mechanistic differences between lamellipodium extension and membrane ruffle formation that include different SNARE-regulated pathways.
A SNAP23-syntaxin13-VAMP3 complex is induced in CHO cells attached to fibronectin
We have previously reported that the function of SNAREs, including SNAP23, are required for for endosomal trafficking and normal cell adhesion in CHO-K1 cells
Figure 1
SNAP23-syntaxin13-VAMP3 interaction in CHO cells is stimulated by adhesion to FN
SNAP23-syntaxin13-VAMP3 interaction in CHO cells is stimulated by adhesion to FN. CHO-K1 cells were held in suspension after pretreatment with 1 mM NEM and 2 mM DTT (
Additional file 1
Quantification of SNAREs. (A and B) CHO-K1 cells were held in suspension after pretreatment with 1 mM NEM and 2 mM DTT (
Click here for file
In several cell types, including glial cells
To address the possibility that interactions amongst SNAREs were occurring post-lysis, 'lysate-mixing experiments' were performed. CHO cells were transfected with FLAG-SNAP23, or GFP-syntaxin13, or cotransfected with both these tagged constructs. Cotransfected cells were held in suspension or plated on FN for 20 min, lysed and the GFP-syntaxin13 was immunoprecipitated with anti-GFP antibody. Cells transfected with FLAG-SNAP23 or GFP-syntaxin13 alone were held in suspension and lysed. After lysis, the FLAG-SNAP23-containing and GFP-syntaxin13-containing lysates were mixed for 1 h followed by immunoprecipitation of the GFP-syntaxin13 using GFP antibody for 15 h. GFP immunoprecipitates were probed for the presence of FLAG-tagged SNAP23. FLAG-SNAP23 only co-precipitated with GFP-syntaxin13 in cells cotransfected with GFP-syntaxin13 and FLAG-SNAP23, not in samples where FLAG-SNAP23 and GFP-syntaxin13 were mixed
Additional file 2
Complex containing Syntaxin13 and SNAP23 does not form after lysis of cells. Cells transfected with FLAG-SNAP23 or GFP-syntaxin13 alone were held in suspension and lysed. After lysis, the FLAG-SNAP23-containing and GFP-syntaxin13-containing lysates were mixed for 1 h (
Click here for file
VAMP3, syntaxin13 and SNAP23 are required for lamellipodium extension
To assess the function of VAMP3, syntaxin13 and SNAP23 in cell spreading, we examined the effect of inhibiting these SNAREs during cell adhesion. SNAREs were inhibited by expressing GFP-tagged, truncated constructs of VAMP3 (aa 1-77; VAMP3cyto), VAMP4 (aa 1-118; VAMP4cyto), syntaxin13 (aa 1-250; syntaxin13cyto) and GS15 (aa 1-85; GS15cyto). These constructs lack transmembrane domains, and are therefore cytosolic proteins that can bind cognate SNAREs through their SNARE domain, putatively blocking interactions with membrane-anchored endogenous SNAREs and exerting a dominant-negative effect. A GFP-tagged SNAP23 construct with the C-terminal nine amino acids deleted was used (SNAP23CΔ9) to inhibit SNAP23 function. The mutant SNARE constructs described here have been used previously to inhibit membrane trafficking in several experimental systems
Additional file 3
Dominant-negative SNAREs bind cognate SNAREs and impair endogenous binding. CHO-K1 cells were transiently transfected with truncated GFP-tagged constructs of syntaxin13 (syn13cyto-GFP), VAMP3 (VAMP3cyto-GFP), VAMP4 (VAMP4cyto-GFP) or SNAP23 (SNAP23CΔ9-GFP), lysed and cognate SNAREs were immunoprecipitated with antibodies coupled to sepharose beads. Immunoprecipitates were washed and analyzed by SDS-PAGE-western blotting for inhibitor and endogenous SNARE binding. (A) Syntaxin13 western blot of SNAP23 immunoprecipitaes from syn13cyto transfected cells. (B) VAMP3 western blot of SNAP23 and syntaxin13 immunoprecipitaes from VAMP3cyto transfected cells. (C) VAMP4 western blot of SNAP23 and syntaxin13 immunoprecipitaes from VAMP4cyto transfected cells. (D) SNAP23 western blot of VAMP3 and syntaxin13 immunoprecipitaes from SNAP23CΔ9 transfected cells.
Click here for file
Lamellipodium extension in CHO cells was quantified as the increase in ventral cell area after attachment to FN (see material and methods). Similar to previous results using tetanus toxin light chain (TeTx-LC) to inhibit VAMP3 function
Figure 2
Dominant-negative SNARE domains impair lamellipodium extension
Dominant-negative SNARE domains impair lamellipodium extension. (A) CHO-K1 cells were transiently transfected with GFP-tagged constructs of the indicated SNAREs: GS15, VAMP4, VAMP3, syntaxin13 or SNAP23. Constructs encoded either the full length protein (FL) or the dominant-negative form (cyto; CΔ9 in the case of SNAP23). HeLa cells were transfected with a SNAP23 shRNA construct (SNAP23 shRNA). The cells were plated on FN-coated (20 μg/ml) coverslips and the extent of cell spreading was calculated as the increase in ventral surface area that occurred between 30 and 90 mins. The results are presented as percent of non-transfected control sample. More than 50 randomly selected transfected cells were analyzed per experiment. Means +/- SEM of at least three independent experiments are shown. (B) CHO cells were transfected as above, serum starved for 3 hrs and placed into a transwell chamber containing polycarbonate membranes coated on the underside with FN. Cells were allowed to migrate for 4.5 h, the membranes were fixed and the percentage of transfected cells migrated was assessed. Data is presented as the percentage of transfected cells migrated normalized to full-length controls and represent at least three independent experiments. (C) A sample of the same HeLa cells used in (A) were lysed and analyzed by Western blot for knockdown of SNAP23 expression. Actin served as a loading control. A set of representative blots are shown.
To gain insight into the impact of SNARE inhibition on lamellipodium extension during directed cell migration, haptotactic transwell migration assays were performed. In these assays, cells in serum-free media were placed on top of a membrane with 8 μm pores and allowed to migrate through the pores toward FN coated on the underside of the membrane, as well as 10% serum in the lower chamber. Similar to the spreading assays, inhibition of GS15 and VAMP4 had little or no effect on migration, while syntaxin13cyto, VAMP3cyto and SNAP23CΔ9 inhibited migration by approximately 30%, 20% and 40%, respectively (Fig
The observation that blocking VAMP3, SNAP23 or syntaxin13 function led to an impairment of lamellipodium extension is consistent with two possible explanations. It is possible that cells with impaired SNARE function: 1) cannot extend lamellipodia efficiently; or 2) can extend lamellipodia, but the lamellipodia cannot form stable adhesive contacts with the substratum and therefore are retracted. To assess these two possibilities, lamellipodium protrusion and retraction were examined in real time from cells plated on fibronection for 90 minutes. Kymographs of cellular spreading were generated, and the rate and duration of protrusions and retractions were quantified. Representative kymographs are shown in Fig.
Figure 3
Inhibition of SNAREs reduces the rate of lamellipodium protrusion
Inhibition of SNAREs reduces the rate of lamellipodium protrusion. Transfected CHO-K1 cells were plated in serum-free media on fibronetin for 90 min and monitored in real-time. (A) Representative control (a) and SNAP23CΔ9 (b) cells are shown at initial acquisition (0 min) and after 90 min (90 min) in the top panels. Kymographs (a' and b') in lower panels represent progression of the cell edge along the lines indicated with the cell edge highlighted. (B) The protrusion and retraction rate of the cell edge was determined from kymographs using ImageJ software. Data is presented as the mean velocity of immediate edge movement from at least two cell edges from at least 3 cells (when possible the most protrusive regions of the cell were measured).
Inhibiting VAMP3, syntaxin13 or SNAP23 alters trafficking of β1-integrin
In previous studies, we have demonstrated, using TeTx-LC, that VAMP3 is required for trafficking of β1 integrin from the recycling endosome to the plasma membrane and that this inhibition in traffic is a possible explanation for reduced spreading
Figure 4
Dominant-negative SNARE domains inhibit β1 integrin exocytosis
Dominant-negative SNARE domains inhibit β1 integrin exocytosis. CHO-K1 cells were transiently transfected for 21 h with GFP-tagged constructs of truncated syntaxin13 (Syn13cyto)[D-F], VAMP3 (VAMP3cyto)[G-I], VAMP4 (VAMP4cyto) or SNAP23 (SNAP23CΔ9 - 15 h transfection) [J-L]. Untransfected control cells are shown in A-C. HeLa cells were transfected with a SNAP23 shRNA construct (SN23 shRNA) for 72 h. Cells were incubated with β1 integrin antibody in serum free media for 3 h to allowing internalization of the label. Then the cells were lifted with trypsin, removing remaining surface label, and were plated on 20 μg/mL FN for 10 min (A, D, G, J,), 20 min (B, E, H, K,) or 45 min (C, F, I, L). For exocytosis assays, cells were fixed and any surface exposed labeled-integrin was stained with AlexaFluor 594 secondary antibody (A-L, data for 20 min of adhesion shown in N). Insets show expression of GFP-tagged constructs (D-L). Images are 3 D reconstructions of a z-series. Scale bar represents 10 μm. (M and N) Integrin immunofluorescent staining at 20 min. was quantified in micrographs using ImageJ software. (M) To assess endocytosis of labeled integrin, cells were permeablized with 0.2% Trition X-100 in PBS before staining with AlexaFluor 594 secondary antibody (percent of non-transfected control). (N) Labeled integrin detected on the cell surface in non-permeabilized cells (percent of non-transfected control). Means ± SEM from three independent experiments (at least 20 cells per experiment) are shown in M and N.
Having shown that VAMP3, syntaxin13 and SNAP23 form a complex, and that inhibition of these SNAREs impairs integrin exocytosis, we sought to determine where within the cell β1 integrin was being trapped. CHO cells expressing dominant-negative SNARE domains were incubated with β1 integrin antibody for one hour followed by an acid wash to remove surface antibody. The subcellular localization of the internalized integrin and the endocytic compartment markers Rab11 and Rab4 was then assessed (Fig.
Figure 5
Dominant-negative SNARE domains disrupt β1 integrin trafficking into the recycling endosome
Dominant-negative SNARE domains disrupt β1 integrin trafficking into the recycling endosome. CHO-K1 cells were transiently transfected for 22 h with GFP [A-D] or GFP-tagged constructs of truncated VAMP3 (V3cyto) [E-H], syntaxin13 (Syn13cyto) [I-L], SNAP23 (SNAP23CΔ9 - 15 h transfection) [M-P, A'-D'], GS15 (GS15cyto) [Q-T], or VAMP4 (V4cyto) [U-X]. Cells were labeled with anti-β1 integrin antibody in serum free medium for 1 h to allow internalization of integrin-antibody complexes. Cells were then washed in 0.2 M glycine, pH 2.5, fixed, and stained for Rab11 [C, G, K, O, S, W and C'] and the intracellular integrin-antibody complexes [B, F, J, N, R, and V], or Rab4 [B']. Overlays of the Rab11 and β1 integrin staining are shown in D, H, L, P, T and X, and Rab4 and Rab11 overlays are shown in D'. Arrows indicate conventional colocalization of β1 integrin in a central Rab11-containing compartment. Images are 3 D reconstructions of a z-series. Scale bar represents 10 μm.
Additional file 4
In cells expressing dominant-negative SNARE domains integrin co-localizes with Rab4. CHO-K1 cells were transiently transfected for 22 h with GFP [A-D] or truncated GFP-tagged constructs of VAMP3 (VAMP3cyto) [E-H], syntaxin13 (syntaxin13cyto) [I-L], VAMP4 (VAMP4cyto) [Q-T] or 15 h with SNAP23 (SNAP23CΔ9)[M-P]. Cells were labeled with anti-β1 integrin antibody in serum free media for 1 h to allow internalization of the label. Cells were then washed in 0.2 M glycine, pH 2.5, fixed and stained for the labeled integrin [B, F, J, N, R] and Rab4 [C, G, K, O, S]. Overlays of the Rab4 and β1 integrin staining are shown in D, H, L, P and T. Images are 3 D reconstructions of a z-series. Scale bar represents 10 μm.
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Additional file 5
Dominant-negative SNARE domains inhibit trafficking of transferrin into a perinuclear compartment. CHO-K1 cells were transiently transfected for 22 h with truncated GFP-tagged constructs of VAMP3 (VAMP3-cyto) [A], syntaxin13 (syntaxin13-cyto) [C], VAMP4 (VAMP4-cyto) [G], or 15 h with SNAP23 (SNAP23CΔ9) [E]. To load the perinuclear recycling compartment with Tfn, cells were serum starved for 1 hr followed by incubation with 30 μg/mL rhodamine-Tfn in serum-free media for 15 min. Coverslips were briefly washed with 0.2 M glycine, pH 2.5, to remove surface bound Tfn, washed three times with PBS and fixed in 4% paraformaldehyde in PBS. Arrows in B, D and F point to perinuclear, rhodamine-Tfn-containing compartment in non-transfected control cells. Images are 3 D reconstructions of a z-series. Scale bar represents 10 μm
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To further assess the role of SNAP23 in trafficking of β1 integrin, we monitored trafficking of the integrin in HeLa cells expressing shRNA to reduce expression of SNAP23 (as described for Fig.
Figure 6
shRNA-mediated downregulation of SNAP23 disrupts β1 integrin trafficking into the recycling endosome
shRNA-mediated downregulation of SNAP23 disrupts β1 integrin trafficking into the recycling endosome. Hela cells were transiently transfected for 72 h with GFP + PLOK-1 vectror [A-D] or GFP + vector containing SNAP23-targeting shRNA [E-H]. Cells were labeled with anti-β1 integrin antibody in serum free media for 1 h to allow internalization of integrin-antibody complexes. Cells were then washed in 0.2 M glycine, pH 2.5, fixed, and stained for the intracellular integrin-antibody complexes [B, F] and Rab11 [C, G]. Overlays of the Rab11 and β1 integrin staining are shown in D and H. Arrows indicate conventional colocalization of β1 integrin in a central Rab11-containing compartment. Images are 3 D reconstructions of a z-series. Scale bar represents 10 μm.
SNAP23 and VAMP4 function are required for PMA-induced membrane ruffling
The observation that blocking VAMP3, SNAP23 or syntaxin13 function inhibits integrin trafficking suggested that SNARE-mediated trafficking supports lamellipodium extension by providing ECM receptors at the leading edge to aid in attachment and stabilization of lamellipodia. Previous studies, however, have shown that when membrane trafficking is inhibited integrin-mediated cell attachment is not altered
In CHO cells, endogenous SNAP23 was detected in PMA-induced membrane ruffles (Fig.
Figure 7
Analysis of SNARE function in PMA-induced membrane ruffle formation
Analysis of SNARE function in PMA-induced membrane ruffle formation. CHO-K1 cells were treated with 500 nM PMA in DMEM (PMA) or DMEM alone (control) for 10 mins, fixed, permeabilized and stained with anti-SNAP23 antibody, Alexa-488-conjugated secondary antibody (A, D) and rhodamine-phalloidin (B, E). A-C, control cell; D-F, PMA-treated cell. Some cross-reactive binding of the SNAP23 antibody in the nucleus is observed (A, D). C and F are overlays of panels A and B and D and E, respectively. Scale bar represents 10 μm. (G) CHO-K1 cells were transfected with GFP vector alone, GFP-tagged dominant-negative constructs of the indicated SNAREs, or GFP together with the light chain of tetanus toxin. HeLa cells were transfected for 72 h with GFP + PLOK-1 or GFP + vector containing SNAP23-targeting shRNA. Cells were treated with 500 nM PMA in DMEM (PMA) or DMEM alone (control) for 10 mins, fixed, permeabilized and stained with rhodamine-phalloidin. Samples were then imaged by confocal microscopy and actin-rich ruffles on the dorsal region of the cells were quantified as described previously. Ruffle indices (expressed as mean +/- SEM) are shown from three independent experiments (more than 25 randomly selected transfected cells were analyzed per condition in each experiment). (H and I) CHO-K1 cells were treated as above, lysed, and VAMP4 (H) or VAMP3 (I) were immunoprecipitated. Immunoprecipitates were washed and analyzed by SDS-PAGE-western blotting for SNAP23,
Inhibition of VAMP3 or syntaxin13 did not significantly alter PMA-induced membrane ruffle formation (Fig.
The observation that VAMP4cyto, SNAP23CΔ9 inhibited membrane ruffling suggested that these SNAREs may function in the same ruffling-dependent trafficking pathway. Therefore, we examined whether PMA could induce the formation of a VAMP4-SNAP23 complex in serum starved CHO cells. Stimulation with PMA increased the amount of SNAP23 that immunoprecipitated with VAMP4 (Fig.
In the present study, we have shown that VAMP3, syntaxin13 and SNAP23 form a complex during CHO cell adhesion to FN. While previous studies have documented the existence of a VAMP3-SNAP23-containing SNARE complex
The cellular importance of the SNAP23-syntaxin13-VAMP3 complex is revealed by data showing that cell spreading and β1 integrin exocytosis were impaired when the function of these SNAREs was disrupted. Our findings are consistent with much evidence indicating that cellular adhesion requires regulated membrane traffic, including the recycling of integrins and the redistribution of integrins to sites of lamellipodium protrusion
Our findings do not exclude the possibility of other SNAREs being involved in the formation of lamellipodia, but they do suggest that SNAP23 function may be central in this process. It is possible that SNAP23 functions at the plasma membrane and at internal endosomes to facilitate the delivery of secreted and/or recycled components during lamellipodium extension. Other SNAREs (e.g. v-SNAREs in endosomal compartments) may function in compartment-specific pathways, but all these pathways may, in turn, depend on SNAP23 function. If so, then SNAP23CΔ9 would be expected to be a more potent inhibitor of cell motility than other soluble SNARE domains, as was observed. In support of this model, others have reported that a VAMP2-syntaxin13-SNAP25 complex mediates traffic from a sorting endosome to a recycling endosome in neurons
The effects of SNARE inhibition on cell spreading and migration suggest that SNARE function is required to support the localized extension of the plasma membrane to form lamellipodia. It is interesting to note that, compared to the effects on lamellipodium extension (Fig.
The fact that inhibition of VAMP4 or GS15 impaired membrane ruffle formation is consistent with a previous study that suggested PMA-induced ruffle formation is dependent on Arf1 activity
The results described here are the first to directly demonstrate that formation of a SNAP23-syntaxin13-VAMP3 complex is modulated during cell-ECM adhesion. The observations contribute to a growing body of evidence describing an important role for SNARE-mediated membrane traffic in cell adhesion. Importantly, the present findings advance our understanding of the role of SNARE-mediated membrane traffic in cell adhesion; it supports transport from a sorting endosome to a recycling compartment during the delivery of adhesive receptors to the plasma membrane. Furthermore, blocking the function of SNAP23 impaired the formation of membrane ruffles in response to PMA, a process that is expected to be independent of adhesive receptor localization. Thus, SNARE-mediated traffic may be needed to deliver membrane components that allow the plasma membrane to be remodeled during the elaboration of membrane ruffles, as well as lamellipodia. The fact that different SNAREs are required for the formation of ruffles and lamellipodia suggests that these processes have partially separate molecular mechanisms. We have now identified a SNARE complex that is induced in cells in response to ECM attachment and that is essential for normal cell adhesion. Future work to elucidate how SNARE complex formation is regulated during cell adhesion will further advance the understanding of cell-ECM interactions.
Reagents and cDNA constructs
All chemicals were purchased from Sigma Chemical Co. (St. Louis, MO) or Fisher Scientific (Nepean, ON) unless otherwise indicated. Antibodies against Rab4 and Rab11 were obtained from Cedarlane Laboratories Ltd. (Mississauga, ON) and BD Biosciences (Mississauga, ON), respectively. All secondary antibodies, rhodamine-labeled transferrin and rhodamine-labeled phalloidin were purchased from Invitrogen (Mississauga, ON).
The pcDNA3.1-SNAP23fl, pcDNA3.1-SNAP23CΔ9, pcDNA3.1-tetanus toxin light chain (TeTx-LC) and peGFP-VAMP3 were kind gifts from Dr. W.S. Trimble (Hospital for Sick Children, Toronto). The pEGFP-SNAP23CΔ9 and pEGFP-SNAP23fl constructs were generated by inserting the
Cell culture and transfection
Chinese hamster ovary (CHO)-K1 or HeLa cells were grown in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% (v/v) fetal bovine serum (FBS; HyClone, Missisauga, ON) at 37°C and 5% (v/v) CO2. Cells were transfected using FuGENE6 transfection reagent following the manufacture's suggested protocol (Roche Applied Science, Laval, QC) or calcium phosphate-DNA precipitation. DNA complexes were incubated on the cells for 18 h with GFP-SNAP23 constructs, 48-72 h with the shRNA SNAP23 construct (Open Biosystems, Huntsville, AL) or 24 h when transfecting with other plasmids. Co-transfection studies were performed at a 10:1 molar ratio of DNA-to-marker DNA, except for FLAG-SNAP23/GFP-syntaxin13 co-transfection where the molar ratio was 1:1. Cells were pretreated for 3 h with 5 μg/mL Brefeldin A (BfA) and BfA was maintained in media during experiments where appropriate.
Immunofluorescence spreading and transwell migration assays
To monitor spreading, cells were plated onto fibronectin (FN) coated (20 μg/ml) glass coverslips for 1.5 h, washed and fixed with 4% (w/v) paraformaldehyde (w/v)/PBS. Samples were then permeabilized and blocked before staining with primary and secondary antibody, followed by washing and mounting. Samples were imaged using a 40X or 63X (NA 1.4) lens on a Leica DM-IRE2 inverted microscope with a Leica TCS SP2 system (Leica, Heidelberg, Germany). Images were captured and 3 D reconstructions were performed using Leica Confocal Software package.
Spreading was quantified by capturing digitized images using a 40X lens. The images were analyzed by outlining cells stained with rhodamine-phalloidin and measuring ventral cell area in ImageJ software. Increases in ventral cell area after 30 min attachment were considered as lamellipodia extension
For live cell imaging, CHO cells were transfected, lifted in serum- and phenol red-free DMEM/F12 (Gibco, Missisauga, ON), supplemented with 15 mM HEPES, pH 7.4, plated on FN coated coverslips inserted in a Series 40 chamber in a QE-1 heated platform (Harvard Apparatus Canada, Saint-Laurent, QC), sealed on top with a coverslip and mounted on the microscope stage. Temperature was maintained at 37°C with a TC-324B temperature controller (Harvard Apparatus Canada, Saint-Laurent, QC) for 90 min. Time series were imported into ImageJ software for the generation of kymographs and analysis.
Integrin and transferrin trafficking assays
To monitor integrin exocytosis, cells were incubated in β1 integrin antibody (AB1950 'fibronectin receptor', Chemicon, Temecula, CA) diluted 1:100 in DMEM for 3 h (internalization), washed with ice-cold PBS and then lifted with trypsin-EDTA (removing surface-bound antibody). Cells were plated on FN-coated coverslips in serum-free DMEM, incubated at 37°C for 10, 20 or 45 min, fixed, and then stained with labeled secondary antibody, with and without permeablization. To monitor integrin localization, transfected CHO cells were incubated with AIIB2 antibody (developed by C.H. Damsky, obtained from the Developmental Studies Hybridoma Bank, Iowa City, IA) diluted 1:75 in DMEM for 1-2 h (internalization). Cells were then quickly washed in cold 0.2 M glycine, pH 2.5, to remove surface label and washed several times with PBS before fixing at 4°C with 4% paraformaldehyde/PBS. For transferrin trafficking assays, transfected CHO cells were grown on coverslips, serum-starved for 1 h and then incubated with 30 μg/mL rhodamine-labeled holo-transferrin (Invitrogen, Mississauga, ON). Cells were quickly washed with cold 0.2 M glycine, pH 2.5, PBS and fixed at 4°C with 4% paraformaldehyde/PBS. Samples were counterstained where appropriate and imaged as outlined above. Integrin endoyctosis/exocytosis was quantified by analyzing integrin fluorescence staining intensities from micrographs in ImageJ software.
Immunoprecipitation and Western blotting
CHO cells were pretreated for 10 min with 1 mM
Sequential immunoprecipitates were performed as above except that after adsorption of lysates to immobilized syntaxin13 antibodies for 4 h, the beads were washed three times with lysis buffer and eluted with 100 μL 0.2 M glycine, pH 2.5. The pH of eluate was raised to 7.5 with 1 M Tris, pH 8.5 and a sample was set aside for the detection of VAMP3 and syntaxin13 before the rest was diluted to one milliliter with lysis buffer. Then the diluted elute was adsorbed to immobilized VAMP3 antibodies for 4 h. The beads were washed twice with lysis buffer before elution with hot 2X Lamelli buffer and analysis by Western blot. Membranes were blotted for SNAP23.
Quantitative ruffling assay
Cells were plated on FN-coated (10 μg/ml) glass coverslips for 3 h in serum-free DMEM. The cells were then treated with 500 nM PMA in DMEM, or DMEM alone as control, for 10 min. Cells were fixed with 4% paraformaldehyde/PBS and stained with rhodamine-phalloidin (Invitrogen, Mississauga, ON). An automated method to detect and quantify F-actin-containing ruffles in confocal micrographs was used and this approach is described in detail elsewhere
CHO: Chinese hamster ovary; DMEM: Dulbecco's modified Eagle's medium; ECM: extracellular matrix; FBS: fetal bovine serum; FN: fibronectin; GFP: green fluorescent protein; NSF:
Authors' contributions
MS conceived and carried out the immunoprecipitation, lammellipodia extension and immunofluorescence trafficking assays, and drafted the initial manuscript. QY conceived and performed the ruffling assays. MJK carried out the migration assays and contributed to the ruffling assays. DWM assisted in the immunoprecipitation assays. KCW contributed experimentally to the shRNA knock-down studies. AB generated several of the SNARE constructs and contributed to the spreading assays. MGC conceived/designed the study, analyzed/processed data, assembled figures and edited the manuscript. All authors read and approved the final manuscript.
The authors would like to thank David Foster and Kevin Luc for the generation of the GS15, VAMP4 and FLAG-SNAP23 constructs. This work was supported by the Natural Sciences and Engineering Research Council of Canada (NSERC) and the Ontario Ministry of Research and Innovation. M.S. holds an Ontario Graduate Scholarship.
Rho, rac, and cdc42 GTPases regulate the assembly of multimolecular focal complexes associated with actin stress fibers, lamellipodia, and filopodia
Focal adhesions: a nexus for intracellular signaling and cytoskeletal dynamics
Stimulation-dependent recycling of integrin beta1 regulated by ARF6 and Rab11
Requirement for Arf6 in cell adhesion, migration, and cancer cell invasion
PDGF-regulated rab4-dependent recycling of alphavbeta3 integrin from early endosomes is necessary for cell adhesion and spreading
PKD1/PKCmicro promotes alphavbeta3 integrin recycling and delivery to nascent focal adhesions
Neuropilin-1/GIPC1 signaling regulates alpha5beta1 integrin traffic and function in endothelial cells
Rho and Rab small G proteins coordinately reorganize stress fibers and focal adhesions in MDCK cells
A Rab8-specific GDP/GTP exchange factor is involved in actin remodeling and polarized membrane transport
Dynamin II interacts with syndecan-4, a regulator of focal adhesion and stress-fiber formation
The road taken: past and future foundations of membrane traffic
The mechanisms of vesicle budding and fusion
SNARE-mediated membrane fusion
Inhibition of SNARE-mediated membrane traffic impairs cell migration
Tetanus neurotoxin-mediated cleavage of cellubrevin impairs epithelial cell migration and integrin-dependent cell adhesion
SNARE-mediated trafficking of alph-5beta-1 integrin is required for spreading in CHO cells
SNARE-mediated membrane traffic modulates RhoA-regulated focal adhesion formation
A second SNARE role for exocytic SNAP25 in endosome fusion
Cultured glial cells express the SNAP-25 analogue SNAP-23
Subcellular distribution of 3 functional platelet SNARE proteins: human cellubrevin, SNAP-23, and syntaxin 2
Intracellular localisation of SNARE proteins in rat parotid acinar cells: SNARE complexes on the apical plasma membrane
Vesicle-associated membrane protein 3 (VAMP-3) and VAMP-8 are present in human platelets and are required for granule secretion
Early/recycling endosomes-to-TGN transport involves two SNARE complexes and a Rab6 isoform
Syntaxins 13 and 7 function at distinct steps during phagocytosis
Activation of store-operated calcium channels: assessment of the role of snare-mediated vesicular transport
Syntaxin 13 is a developmentally regulated SNARE involved in neurite outgrowth and endosomal trafficking
Cholecystokinin-regulated exocytosis in rat pancreatic acinar cells is inhibited by a C-terminus truncated mutant of SNAP-23
GS15 forms a SNARE complex with syntaxin 5, GS28, and Ykt6 and is implicated in traffic in the early cisternae of the Golgi apparatus
VAMP3, syntaxin 13 and SNAP23 are involved in secretion of matrix metalloproteinases, degradation of the extracellular matrix and cell invasion
Tetanus toxin-mediated cleavage of cellubrevin impairs exocytosis of transferrin receptor-containing vesicles in CHO cells
Automated classification and quantification of F-actin-containing ruffles in confocal micrographs
Inhibition by brefeldin A of protein secretion from the apical cell surface of Madin-Darby canine kidney cells
Dominant inhibitory mutants of ARF1 block endoplasmic reticulum to Golgi transport and trigger disassembly of the Golgi apparatus
Interaction of Munc-18-2 with syntaxin 3 controls the association of apical SNAREs in epithelial cells
Integrin trafficking and the control of cell migration
SNAP25, but not syntaxin 1A, recycles via an ARF6-regulated pathway in neuroendocrine cells
Two distinct actin networks drive the protrusion of migrating cells
Actin microfilament dynamics in locomoting cells
Omega-3 and omega-6 fatty acids stimulate cell membrane expansion by acting on syntaxin 3
Studies of the roles of ADP-ribosylation factors and phospholipase D in phorbol ester-induced membrane ruffling
ARF1-mediated actin polymerization produces movement of artificial vesicles
Actin and Arf1-dependent recruitment of a cortactin-dynamin complex to the Golgi regulates post-Golgi transport
Participation of the syntaxin 5/Ykt6/GS28/GS15 SNARE complex in transport from the early/recycling endosome to the trans-Golgi network
GGA function is required for maturation of neuroendocrine secretory granules
Difference in distribution of membrane proteins between low- and high-density secretory granules in parotid acinar cells
Biogenesis of regulated exocytotic carriers in neuroendocrine cells
Homotypic fusion of immature secretory granules during maturation requires syntaxin 6
Mapping of R-SNARE function at distinct intracellular GLUT4 trafficking steps in adipocytes


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