Sometimes the universe chooses the path for you, ya know? And in this case, since the road less traveled is not necessarily the more difficult of the two (seeing as how there are growling, snarling monsters patrolling the other), how is a person to know which one would make "all the difference"? It's a conundrum, to be sure, and perhaps one which jefbot will learn the answer to very shortly.
Gobots! now that takes me back… heheh.
well, except for that awesome garbage lid throw a couple strips back, ZodiacMage! but, yeah – except for that, this trip hasn't been very lucky for him.
As Joss Whedon sort of said, there's a reason the road less traveled isn't one's first choice.
heh. SOOO true, Sarah W. Mr. Whedon's a smart man.
I can't tell you how much I'm enjoying this story. Thank you for all your hard work. This comic has become one of my favorites and everything you put into its creation is very much appreciated.
Just when you think you're safe and can let your guard down the universe kicks you in the rear and says "F#$% YOU!" with a smile on its face. Almost never fails.
although it makes the times when things DO go right that much sweeter, Fijiman! let's hope bot tastes some of that sweetness soon.
Shoot, 'bot…at least there's a path (which is more than I was expecting when I saw yesterday's comic). Also, good on you for having the cool-under-pressure-ness to say "two roads diverged in Hollywood." I guess when you're running from alien-enhanced creatures, you have to take your excuses for witty wordplay when you can get them.
I had no idea there was a play on words intended until I read your post. I typed in "two roads diverged" and I was pointed toward a site about the old Robert Frost poem "The Road Less Taken". Thanks for this educational experience.
I realized after I used my search engine and posted my last reply that I could have clicked above for the Robert Frost poem.
every so often my English degree rears its precious head, Rainey, so i fear the same happens to bot, especially when he comes across two diverging roads. and i love that you were intrigued enough to go on a search which led to Robert Frost! too cool.
no doubt bot will have to wander from that path sooner or later, JOE, but yes – for now, at least he has some direction (for good or ill)! and, yeah: you definitely have to take the opportunities for witty wordplay when you can. heheh. especially when in danger from mutated beasties!
Considering how full Jefbot's hands are, this would be a bad time for him to be forced to climb up the side of that mountain.
indeed, Rainey. let's hope that dark road doesn't cut out on him anytime soon.
Me likey the mysterious style in this strip. It brings out the beauty in a way.
I've come to realise, after living in it for 16 years, that the universe is not always fair.
Bot made a wise choice. Creepy forest over techmonsters, every time.
glad you found the strip, uh, "beauterious" today, DAS! and, yeah – the universe isn't always fair but it's not always unfair, either, which makes life "beauterious" in a way.
The nice thing is, from the top of the hill he can drop angsty teenagers on them. It's sort of a win/win that way.
ha! true that, Oz. if some "angsty teenagers" are up there, they're gonna get a big surprise when bot shows up with a large, unconscious, mutated hamster in his backpack and some big beasties in tow.
HEY! Not gonna be on frequently anymore. No more internet. The connection where I work is intermittent at best, so I'll pop in when I'm able. Don't know when I'll be getting a regular connection again, so please regard me kindly.
That said, good luck JB and Mu! So far, so good, though Murphy's starting to go after you.
poop! well, i hope you find another internet connection soon, KB – you're a valuable voice here and i'll miss it while you're gone. hope to see you back here soon.
and, yes: murphy's law is impossible to escape, it seems. damn murphy!
Please tell me they'll run into a mutated pug.
ha! that's verrrrrrry possible, GB!
As long nothing escaped from the zoo…. DUH DUH DUUUUUUH!!!
yikes! let's hope not, Raron – a mutant rhino would be bad news for bot!
One of my favorite poems, that.
it's a good 'un, Insectoid.
Poor Bot having to take the more difficult road – at least in terms of topology and quality of pavement. It looks MUCH less difficult in terms of the number of red, glowy, bad former-pets. Hopefully it will stay that way!
And hopefully Musashi will wake up soon and provide some guidance on whether Jefbot is heading the right way.
heheh. when you put it that way, the less glow-y road could be the much easier option, Shanna!
It suddenly occurred to me that in panel 2, Bot/Mu looks like Ratchet & Clank. Does Musashi give Bot a double jump ability?
a double jump ability would be extremely helpful right about now, Skreyola! also helpful? A CHICKEN GUN!
*gasp* A Chicken Gun or Sheep Gun?
Yo, I only hope Bot doesn't regret his decision in the road ahead. Then the irony of referencing the poem would be complete.
true that, Valorius. and i have a feeling there are going to be many opportunities for bot to regret referencing that poem during the rest of this storyline!
Lol. That was absolutely funny.
glad you like it, Jf#300!
I used to love "Choose Your Own Adventure" books. Now I read "Endless Quest" and "Choosamatic" books. Both of these are a considerably higher reading level than "Choose Your Own Adventure". I used to stink at decision making skills. After reading these, well… Decision making still isn't my strong point but at least my skills don't stink. If Morris Simon or Matthew Youngmark were the author of this strip, going headfirst into the mob of mutant creatures would be the correct choice. I remember these two authors just looove to put nasty twists in their stories in order to cause you to make the wrong choices.
huh. i've never read any "Endless Quest" or "Choosamatic" books, Rainey, as i never knew there was anything beyond CYOA books! i might have to check 'em out, if only for curiosity's sake.
I tune out to study for a few days, and Jefbot puts on the badass. Props, Jefbot. Props.
ha! glad you found his actions badass, Bender_Sastre. happy studying!
true dat, ROM! always tricky figuring out which way to go.
All caught up again! I had to laugh at Bot's "Damn universe" comment. And Bot looks way cool in the 2nd panel, great job Jeff!
glad you're all caught up, Natalie, and thanks for the kind words!
Now I know what to say when things get tough, "Damn universe". Gonna use it!


We at B.K. Electricals, established in the year 1996, are the leading manufacturer, exporter, importer, trader and supplier of industrial heaters and thermocouples. We are fast growing industrial units in the private sector in India and are engaged in manufacturing and supplying with precision components an ultra modern machinery and equipments. Our catalog products are different and are of excellent in comparison in the race. They are well appreciated in the global market and are a benchmark for others in the race. The quality and durability of our catalog products are inspiration for others and are felicitated by our customers and our valued clientele. Our products are of unbeatable prices and recommended in the global market.
On the professional front, the distribution activities are in the hands and under the supervision of our experienced and expert employees who are in panel division and therefore responsible for their panel activities of manufacturing and supplying. The aptitude and knowledge prospects of our professional are unmatchable and looked upon in the market for their best of the dealings. It is our prime responsibility to be well organized and coordinated towards distributing the catalog products to our valuable clientele. We believe in building friendly and informal relations with our esteemed clients so as to make the client-customer and industrial relations become stronger and strengthen our business deals. Our industry has grown on the strong customer and client base which is a foundation of lighting torch to the success of our company. Our company requirements are upgraded from time to time when needed and in accordance with the present scenario. The quality, sturdiness and durability of the products are well checked with precision by our quality control and logistics department and are then persuaded to our valued clientele and customers.
The continuous rise is with the possible inspirational mentor ship of our Mr. Sumeet Agrawal and Mr. N. K. Agrawal. He has strives and dedication towards the company is unmatched which has taken our organization to another level of success. His managerial skills to coordinate with the departments, giving instructions, supervision and motivation have made our professionals reach heights and have achieved their targets important for every position in the company. His hierarchical man of his motivation, feel free attitude to put forth the views and regular feedback o the performance of each department an each employee, despite the fact of position.
We run our business by adopting high business methodologies at our production base. With this approach and with the help of our expert engineers we are able to procure qualitative grade products with accurate dimensional measurements. In this span, we have gained a long list of satisfied buyers. Further, we believe to maintain transparent facilities at our unit, which means everyone at our base easily talk to the supreme management. Similarly, our trainers conduct regular training sessions for the update of market trends and latest technology.
Bound with the promise of quality, our unit is involved in manufacturing of high quality sphere of industrial heaters. To bring out innovative solutions we ensure procurement of reliable quality products within specified time limit. Further, we follow ethical business practices that make us to achieve higher standard in this competitive market. We design heater according to customer requirements with sound technical support and solutions with cost effective and energy efficiant heat solutions.


text:
Tag: Feng Shui
What are the five elements of fung shui? And how to use them?
What are the five elements of fung shui? The ancient Chinese believed that the five elements of fire, earth, metal, water, and wood represent the
Bagua Map Feng Shui and How to Use It?
Bagua map feng shui is a popular Taoist-based feng shui technique that uses the eight trigrams on the map to guide practitioners in decorating and
The Meaning of Yin and Yang in Feng Shui
The meaning of yin and yang is simply the relationship between opposites to create balance in any environment. This philosophy is a fundamental idea in
What is Feng Shui and How Can it Help You?
What is Feng Shui? Is it a branch of interior design, or it is something to do with moving furniture around and hanging mirrors? Don't
Mirrors in Feng Shui: Uncover the Secrets of Bedroom Mirrors.
Mirrors in Feng Shui for bedrooms and interior spaces have been used for centuries as a way to improve the flow of energy. While the
Facebook-f Globe Linkedin-in Pinterest Instagram
KHAMES © 2023 , All Rights Reserved.
Partner Projects: ALDECOR

redpajama_set_name: RedPajamaCommonCrawl


text:
Q: Multiline текст в checkedListBox с#(winforms) Есть варианты с очень длинным текстом, хотелось бы сделать перенос на другую строчку.Как это сделать?

A: Создайте новый проект с формой. Обзовите форму MainForm. Киньте на форму DataGridView, обзовите его dataGridView. И скопируйте этот код:
public partial class MainForm : Form
{
    DataGridViewCheckBoxColumn _checkCol;
    DataGridViewTextBoxColumn _textCol;

    public MainForm()
    {
        InitializeComponent();

        _checkCol = new DataGridViewCheckBoxColumn();
        _checkCol.Name = "CheckCol";
        _checkCol.HeaderText = "CheckCol";
        _checkCol.Resizable = DataGridViewTriState.False;
        _checkCol.AutoSizeMode = DataGridViewAutoSizeColumnMode.DisplayedCells;

        var textColCellStyle = new DataGridViewCellStyle();
        textColCellStyle.WrapMode = DataGridViewTriState.True;

        _textCol = new DataGridViewTextBoxColumn();
        _textCol.AutoSizeMode = DataGridViewAutoSizeColumnMode.Fill;
        _textCol.Name = "TextCol";
        _textCol.HeaderText = "TextCol";
        _textCol.DefaultCellStyle = textColCellStyle;
        _textCol.ReadOnly = true;

        dataGridView.AllowUserToAddRows = false;
        dataGridView.AllowUserToDeleteRows = false;
        dataGridView.AllowUserToResizeColumns = false;
        dataGridView.AllowUserToResizeRows = false;
        dataGridView.AutoSizeRowsMode = DataGridViewAutoSizeRowsMode.AllCells;
        dataGridView.CellBorderStyle = DataGridViewCellBorderStyle.None;
        dataGridView.CellMouseDown += dataGridView_CellMouseDown;
        dataGridView.ColumnHeadersHeightSizeMode = DataGridViewColumnHeadersHeightSizeMode.AutoSize;
        dataGridView.ColumnHeadersVisible = false;
        dataGridView.BackgroundColor = Color.White;
        dataGridView.MultiSelect = false;
        dataGridView.RowHeadersVisible = false;
        dataGridView.SelectionMode = System.Windows.Forms.DataGridViewSelectionMode.FullRowSelect;

        dataGridView.Columns.AddRange(new DataGridViewColumn[] {
            _checkCol,
            _textCol
        });

        dataGridView.Rows.Add(true, "Какой то очень длиный текст который не влезает в одну строку");
        dataGridView.Rows.Add(false, "Какой то очень длиный текст который не влезает в одну строку");
        dataGridView.Rows.Add(true, "Какой то очень длиный текст который не влезает в одну строку");
        dataGridView.Rows.Add(false, "Какой то очень длиный текст который не влезает в одну строку");
        dataGridView.ClearSelection();
    }

    private void dataGridView_CellMouseDown(object sender, DataGridViewCellMouseEventArgs e)
    {
        var row = ((DataGridView)sender).Rows[e.RowIndex];
        if (row.Selected) {
            row.Cells[_checkCol.Index].Value = !(bool)row.Cells[_checkCol.Index].Value;
        }
    }
}

redpajama_set_name: RedPajamaStackExchange


text: Tyler Messerschmidt was last year's 100 and 200 freestyle state champ at the 4A-II state meet, but that doesn't quite tell the full story. In the 200, he broke the all-time state record with a time of 1:38.00, nearly eights seconds better than his next-fastest opponent. He also swam All-American qualifying times in both races and made the All-Arizona team in both events. He has also set sights on qualifying for the 2012 Olympics and according to an Arizona Republic article written last summer, he's already reached marks that would easily qualify him for the 2012 Olympic Trials (although the official marks for qualification haven't been released yet). He's also a lifeguard and plays the drums in a band with friends. Photo By Dave Cruz


text:
Q: Avoid scrollbar from print in yii In my Yii web application, I am using divToPrint for take print out of a div contents. In div having one class table-responsive for responsive table. Because of this class displayed scroll bar. When I am taking print out of this div, color of this scroll bar also present in that print. How to avoid that scroll bar from print out. For print the below function is used,
function printDiv(divName) {
    var divToPrint = document.getElementById(divName);
    var popupWin = window.open('', '', 'width=300,height=300');
    popupWin.document.open();
    popupWin.document.write('<html><body onload="window.print()">');
    popupWin.document.write('<link href="<?php echo Yii::app()->request->baseUrl ?>/css/assets/css/minified/ccebootstrap.min.css" rel="stylesheet" type="text/css">');
    popupWin.document.write('<link href="<?php echo Yii::app()->request->baseUrl ?>/css/assets/css/minified/core.min.css" rel="stylesheet" type="text/css">');
    popupWin.document.write('<link href="<?php echo Yii::app()->request->baseUrl ?>/css/reportcardstyle.css" rel="stylesheet" type="text/css">');
    popupWin.document.write(divToPrint.innerHTML + '</html>');
    popupWin.document.close();
}

A: It has nothing to do with Yii, as it's PHP framework.
Use css overflow: visible; on element that contains scrollbar.


text:
Comfortable accommodations, an inviting courtyard, centrally located, many nearby restaurants, friendly atmosphere and the alluring blue Pacific Ocean along the Southern California coast...the only thing missing is you. All accommodations (see our room photo gallery) have been recently renovated and are tastefully decorated. All guests of the hotel enjoy complimentary parking, free wi-fi, air-conditioning, cable tv, in-room refrigerator, iron and ironing board. Additionally, all rooms are non-smoking.
An hour before, in a maze of punch-colored lockers, I located number seven-thousand-something and began shoving all signs of winter weather into it. Puffy coat, recently unwound scarf, thick socks, boots, and sweater all went in, but not before I removed a bikini and beach gear from my overnight bag. An hour outside of Berlin, Tropical Islands (T.I.) fills an airfield hanger originally built in 1938 for the Nazi Luftwaffe. The Red Army overtook the site in 1945 and the Soviet Air Force stayed until 1992. A Malaysian company purchased the hanger at a steep discount in 2003, after some kind of German cargo-blimp enterprise went bankrupt. T.I. opened a year later.
© Snorkeling Report Int. is the owner of the intellectual property rights or holds the rights to use all the elements that can be accessed on the site, particularly the texts, pictures, graphics and logo. Any reproduction, representation, modification, publication or adaptation of all or part of the elements of the site, whatever the means or the procedure used, is forbidden, without prior written authorization from Snorkeling Report.
I am in love with this crochet bikini set. It is so incredible cute. For reference, I am 5'4", weigh 110lbs and normally wear a size 0 bottom and a 32B top. It fits perfectly. However, since it's crochet it could easily expand to accommodate larger sizes. I honestly bought this thinking I could wear it as a legit bikini to the beach, but I was extremely mistaken. It is 100% crochet. So, you can somewhat see through it all. I am going to instead wear it around the pool or boardwalk, but with a strapless bra and underwear on underneath. This is more of clothing than a swimwear bikini. The bottoms fit like booty-shorts and the top is just a typical bandeau. I think it would need an layer underneath of swimsuit material in order for it to be a true swimsuit. Either way, I still love the design/pattern, and the quality of the crochet is great. Can't wait to show this off this summer!
Dave and I honeymooned in the Dominican Republic so it has a very special place in our hearts. The Dominican is a part of the second largest island in the Caribbean so there are many places to capture beautiful tropical island pictures. But this one from Puerto Plata captures our view of what a tropical vacation should be about. Beautiful skies, over the water bungalows and a cruise ship off in the distance.
here's a spot that might be interesting to some people. I've spent a month two years ago in Cozumel, Mexico (right across from Cancun) and found it to be really cool. I worked on cruise ships and it is one of the major cruise ship stops so there are tourists but mostly during the day while the ships are in port. Other than that it's a cool and cheap place to stay. Using the app HomeAway (similar to AirBnB) I've found a place (it was a studio in a 7 apartment bulding, with a gate and a small pool in the back) that I paid $350 for a month for. The internet was very good and there were a few big supermarkets close by. Cozumel, for example is one of the top spots for scuba diving in the Caribbean and the side of the island that's opposite to downtown where the cruise ships dock is full of beaches that rarely anyone goes to but it's a great spot for windsurfing for example.
Give your suitcase a boho vibe with our range of crochet bikinis. Whether you'll be spending the day posing by the pool or bathing at the beach, we've got a crochet bikini for you. Keep it simple with muted tones or go for a bright and bold crochet bikini top with matching bottoms. Whether you're jetting off with the dolls or going to your annual family vacay, we've got a crochet bikini for you. Keep your poolside look authentically retro with crochet bathing suit, just add some hoops and heeled wedges to amp up your style. Whichever way you work it, a crochet bikini is a must-pack this season.
By making an analogy with words like bilingual and bilateral containing the Latin prefix "bi-" (meaning "two" in Latin), the word bikini was first back-derived as consisting of two parts, [bi + kini] by Rudi Gernreich, who introduced the monokini in 1964.[18][19] Later swimsuit designs like the tankini and trikini further cemented this derivation.[20] Over time the "–kini family" (as dubbed by author William Safire[21]), including the "–ini sisters" (as dubbed by designer Anne Cole[22]), expanded into a variety of swimwear including the monokini (also known as a numokini or unikini), seekini, tankini, camikini, hikini (also hipkini), minikini, face-kini, burkini, and microkini.[23] The Language Report, compiled by lexicographer Susie Dent and published by the Oxford University Press (OUP) in 2003, considers lexicographic inventions like bandeaukini and camkini, two variants of the tankini, important to observe.[24] Although "bikini" was originally a registered trademark of Réard, it has since become genericized.[25]
A bandeaukini (alternatively called a bandini)[133] is a bandeau top (no straps going over the shoulders) worn with any bikini bottom.[134][135] It is the oldest form of bikini, with earliest examples found in Sicilian Villa Romana del Casale (dubbed the "Bikini Girls) dating back to the 4th century AD.[29][30][136][137] Reintroduced, its appeal grew fast among young women,[138] with bandeau tops edging into the sales of the classic tankini.[139]


An interview DONE BY MATRICH!! The most awsome person to have stepped foot in the ns office.. ever! MATRICH was suposed to post this but something failed and now im posting it so give him the karma and not me.. I am a tool!


text:
Q: Vulkan SDK detects only 1 GPU at a time I have an Optimus notebook (meaning it has two graphics cards; one integrated and one discrete (dedicated)). Both of them support Vulkan and I can use each of them individually on my code. But there is a problem, I can't list or use them both on the same program. I must select on which GPU it runs before running my code (on Windows: right click on "myProgram.exe" -> Run with graphics processor, on Linux: ./"myProgram" for integrated GPU and primusrun ./"myProgram" for discrete) and it detects only the one I run my code on even when both GPUs are active at the same time.
My GPUs are Intel UHD Graphics 630 and NVIDIA GeForce GTX 1050 Ti. I am using LunarG Vulkan SDK 1.1.92, Intel driver version 25.20.100.6326, NVIDIA driver version 416.32 on Windows and 415.25 on Linux. As C compiler and operating system, MSVC 1912 on Windows 10 and gcc 8.2.1 on Arch Linux. Vulkan runtime and ICD loaders are installed and I get the same result on both OSs.
Here is my code:
uint32_t deviceCount;
vkEnumeratePhysicalDevices(instance, &deviceCount, NULL);
printf("Found: %d\n", deviceCount);

VkPhysicalDevice *devices = malloc(deviceCount * sizeof(VkPhysicalDevice));
vkEnumeratePhysicalDevices(instance, &deviceCount, devices);

for(uint32_t deviceIndex = 0; deviceIndex < deviceCount; deviceIndex++)
{
    VkPhysicalDeviceProperties deviceProperties;
    vkGetPhysicalDeviceProperties(devices[deviceIndex], &deviceProperties);
    printf("%s\n", deviceProperties.deviceName);
}

Here is the outputs I get:
If I run on integrated GPU:
Found: 1
Intel(R) UHD Graphics 630

If I run on dedicated GPU:
Found: 1
GeForce GTX 1050 Ti

What I am expecting is:
Found: 2
Intel(R) UHD Graphics 630
GeForce GTX 1050 Ti

And I should be able to pick either of them to use on my program without hassling with some driver level code or setting it on my OS before running. Vulkan is relatively low level API after all. Is there any way to force list all of the active GPUs on my system?

A: I'm not sure what you mean by

If I run on integrated GPU

versus

If I run on dedicated GPU"...

If you're going into the nVidia control panel and specifying a target GPU for your application, then you're seeing pretty much what I would expect... only that GPU exposed to your app.
I believe what you should be doing is setting it to auto-detect, and then ensuring that you're exporting the "Optimus Aware" flag in your binary.  You should have something like this in your test code
extern "C" {
_declspec(dllexport) uint32_t NvOptimusEnablement = 0x00000001;
}

That should either cause Vulkan to see both GPUs, or force your application to automatically see only the discrete GPU, but without you having to do anything in the control panel, not sure which.


In the past I've found that political orientation items were little explained by the current MOTIV personality traits when I added them as experimental items. I unfortunately didn't pursue that research, writing off political orientation as maybe more like IQ and not being reflected well in personality dimensions. However, I recently reexamined the topic after being puzzled by people with nationalist/xenophobic orientations and reading more of the research on ethnocentrism which is the academic term for nationalist/xenophobic personality tendencies.
What has emerged in my research is a clear independent personality trait that explains the political orientation items I had previously been unable to chart well on MOTIV. More accessibly put, the political spectrum most understand as conservative vs liberal is mostly explained by in group vs the whole group orientation. Conservatives prioritize the in group, their tribe/country/nation/culture and liberals prioritize the whole group, the international, the world/globe. While conservatism is not perfectly analogous to racism, those that self report as conservative are far more likely to endorse racist personality items (such as enjoying racist jokes, preferring segregation to integration, endorsing a belief their race is superior). Sentiments on immigration/immigrants seems to be a central topic to the conservative vs liberal divide with self reported conservatives being far more likely to be distrustful/fearful/disdainful of immigrants/immigration and self reported liberals being more open and accepting of them.
So, I will eventually be adding a new trait to MOTIV labeled Ethnocentrism. I will refer to the low scoring end of this spectrum as Globalism. It is my speculation that Ethnocentrism evolved when humans lived in tribes more independent and cut off from other groups/tribes of humans. This caused more differences between different groups of humans and thus more difficulty getting along, cooperating, and likely more conflict (appropriately, those that score high on Ethnocentrism are more likely to endorse positive views on war and aggression). Globalism has evolved more recently as the human environment has become more connected, interdependent.
While Ethnocentrism is orthogonal to Vitality on an individual level, when I separated a sample of 8352 users into three equally sized groups (high Ethnocentric score, medium Ethnocentric score, low Ethnocentric score), I found that the medium Ethnocentric scoring group had the highest Vitality score (9.39), followed by low Ethnocentric scorers (8.95), with the high Ethnocentric scoring group having the lowest Vitality score (8.73). So this appears to be a trait where a middle preference may be healthiest (at least within the current population sample). Thus, this trait seems to mirror the Interpersonal trait, where the medium scoring group showed a higher Vitality score than the high (overly selfless) or lower (overly selfish) scoring group. If a balance of nationalism/globalism is ideal it might explain the intention-ed design of the American federal/state system, where the founding fathers understood that there needed to be a balance between more micro, local interests and more macro, national interests. Likewise, sometimes local interests need more consideration, and sometimes global interests need more consideration. Alternatively, we may simply be in a transitional mid point evolutionarily and as we continue to progress global interests/power will justifiably begin to take more importance than local interests/power.


text:
80s Song of the Day: If You Leave by Orchestral Manoeuvres in the Dark
April 21, 2017 April 20, 2017 / Janet / Leave a comment
If You Leave by Orchestral Manoeuvres in the Dark (OMD) was released on April 21, 1986, with La Femme Accident as the B side. It is a single from the Pretty in Pink soundtrack and is featured in the final scene of the movie. Continue reading →
80s Song of the Day: Don't You Want Me by The Human League
November 27, 2016 November 26, 2016 / Janet / Leave a comment
Don't You Want Me by The Human League was released November 27, 1981, with Seconds as the B side. It was the fourth single from the band's fourth studio album Dare.
In the United Kingdom, the song was the 1981 Christmas number one. The Human League's first single released in the United States, Don't You Want Me topped the Billboard Hot 100 for three weeks in July 1982. Continue reading →
80s Song of the Day: Obsession by Animotion
Obsession by Animotion was released November 23, 1984, with Turn Around as the B side. It was the debut single from the group's self-titled debut album.
The song originally was written and recorded by Holly Knight and Michael Des Barres in 1983. This version was featured in the 1983 film A Night in Heaven; however, Animotion's version was by far the big success of the two. It reached No. 5 in the United Kingdom and No. 6 in the United States. Continue reading →
80s Song of the Day: Take on Me by a-ha
October 19, 2016 October 18, 2016 / Janet / Leave a comment
In the "if at first you don't succeed" category, Take on Me by a-ha was originally released on October 19, 1984, with And You Tell Me as the B side. But this was not the version – or the accompanying music video – that would become one of the most iconic of the 80s.
The original version peaked at No. 3 in a-ha's native Norway but failed to achieve success elsewhere. Unhappy with the version, the band re-recorded the song with a new producer – Alan Tarney – and released it again in 1985, with Love Is Reason as the B side. Continue reading →
80s Song of the Day: Tonight She Comes by The Cars
Tonight She Comes by The Cars was released on October 14, 1985, with Just What I Needed as the B side. It was a single from the band's Greatest Hits album. Just What I Needed, which also appeared on the Greatest Hits album, was originally released on May 29, 1978 as the debut single from the band's self-titled debut album.
Tonight She Comes was the first of two songs to be released as a single from the Greatest Hits album. The second was a remixed version of I'm Not the One, previously recorded for the 1981 album Shake It Up.
The Cars' fourth Top 10 hit, Tonight She Comes reached No. 7 on the Billboard Hot 100 and No. 1 on the Billboard Mainstream Rock charts.
80s Song of the Day: Human by The Human League
August 11, 2016 August 4, 2016 / Janet / Leave a comment
So many nights I longed to hold you
So many times I looked and saw your face
Nothing could change the way I feel
No one else could ever take your place
Human by The Human League was released on August 11, 1986, with an extended instrumental version of the song as the B side. The song was the first single from the album Crash. Continue reading →
80s Song of the Day: What Have I Done to Deserve This? by Pet Shop Boys
August 10, 2016 August 4, 2016 / Janet / 1 Comment
I bought you drinks, I brought you flowers
I read you books and talked for hours
What Have I Done to Deserve This? by the Pet Shop Boys was released August 10, 1987, with A New Life as the B side. The song, featuring Dusty Springfield, was a single from the album Actually. Continue reading →
80s Song of the Day: Save a Prayer by Duran Duran
August 9, 2016 August 3, 2016 / Janet / Leave a comment
Don't save a prayer for me now
Save it til the morning after
Save a Prayer by Duran Duran was released on August 9, 1982, with Hold Back the Rain (Remix) as the B side. It was the third single from the album Rio. Continue reading →
80s Song of the Day: True Faith by New Order
July 20, 2016 July 16, 2016 / Janet / Leave a comment
I used to think that the day would never come
I'd see delight in the shade of the morning sun
My morning sun is the drug that brings me near
To the childhood I lost, displaced by fear
True Faith by New Order was released on July 20, 1987, with 1963 as the B side. The newly released singles appeared along with all of the band's previously released singles in their 12-inch versions on the compilation album Substance (also known as Substance 1987). Continue reading →
80s Song of the Day: Tainted Love by Soft Cell
July 7, 2016 July 3, 2016 / Janet / Leave a comment
This tainted love you've given
I give you all a boy could give you
Take my tears and that's not nearly all
Tainted Love by Soft Cell was released on July 7, 1981 in the United Kingdom. It featured Where Did Our Love Go as the AA side. (The U.S. single, released in 1982, featured Memorabilia as the B side.) The song, originally released in 1965 by Gloria Jones, was the second single from the album Non-Stop Erotic Cabaret. Continue reading →


This TurnLock part HHSTLHS-3Z is a heavy duty stud. We recommend the use of the following TurnLock retaining washers: HHSWPS or HHSWSS.
TurnLock can offer you stud HHSTLHS-3Z with Certificate of Quality, Certificate of Conformity and Certificate of Origin on request.
TurnLock is offering stud HHSTLHS-3Z as factory new.


14/ Do not use auto posting software. Violators will be banned.
15/ Do not flood. We tolerate 10-15 post per day. Please adhere to a reasonable time between posts of let's say 10-15 minutes. This concerns mostly those posting in single post per thread sections, but megathread owners please be aware of that aswell! Violators will be banned.


text:
Patricija je žensko osebno ime.

Izvor imena
Ime Patricija oziroma Patricia je latinskega izvora, in sicer ženska oblika imena Patricius. Ime Patricius pa izhaja iz latinske besede patricius »patricijski, plemenit«

Različice imena
ženske oblike imena: Patricia, Patriša, Patrišia
moške oblike imena: Patricij, Patrick, Patrik

Tujejezikovne oblike imena
 pri Angležih: Patrick (m), Patricia (ž), skrajšano Pat
 pri Francozih: Patrice (m,ž)
 pri Italijanih: Patrizia
 pri Nemcih: Patricia, Patrizia
 pri Poljakih: Patrycja
 pri Rusih: Patrikija

Pogostost imena
Po podatkih Statističnega urada Republike Slovenije je bilo na dan 31. decembra 2007 v Sloveniji število ženskih oseb z imenom Patricija: 2.073. Med vsemi ženskimi imeni pa je to ime  po pogostosti uporabe uvrščeno na 112. mesto.

Osebni praznik
V koledarju je Patricia 25. avgusta; na ta dan leta 670 je umrla devica Patricija Carigrajska.

Glej tudi
seznam osebnih imen na P
seznam najpogostejših imen v Sloveniji

Viri

Ženska osebna imena

redpajama_set_name: RedPajamaWikipedia


Dark Phoenix DVD and Blu-ray release date not announced.
As the second-most powerful mutant of the X-Men team, Dr. Jean Grey has always struggled with controlling her telepathic powers. As those powers begin to unfold, a dark side of Jean Grey emerges. The other members of the X-Men must find a way to contain the Dark Phoenix before the insidious alter ego of Jean Grey wreaks havoc on the human race. Each member of the team must decide if the life of Jean Grey is worth saving when the Dark Phoenix becomes impossible to control.
DVDs Release Dates is the best source to find out when does Dark Phoenix come out on DVD and Blu-ray. Dates do change so check back often as the release approaches.


text:
Q: Ontology design: Individual in Subclass not showing in Super Class? I just want to clarify as to why an Individual/Member in a subclass doesnt show up as a member in the superclass?
For example
Human
   Male
   Female

If Male has two members/individuals, namely "John" and "Shawn", shouldn't they show up in the superclass too?
From my understanding, "John" and "Shawn", are male and hence Human
Or should I have to explicitly state something to make that happen
Edit:
Okay, so i just tested something, if I use DL-QUERY to query the superclass, the Individual does in fact show up for the superclass …
This is weird. Is this is a bug?
I'm using Protégé 5.2.

A: This is a bug in the Protege 5.0b15. In Protege 4.3 both Shawn and John are showing as Members of class Human after running the reasoner. I filed a bug against Protege 5.0.


In an acidic and lysine-rich environment Escherichia coli induces expression of the cadBA operon which encodes CadA, the lysine decarboxylase, and CadB, the lysine/cadaverine antiporter. cadBA expression is dependent on CadC, a membrane-integrated transcriptional activator which belongs to the ToxR-like protein family. Activation of CadC requires two stimuli, lysine and low pH. Whereas lysine is detected by an interplay between CadC and the lysine-specific transporter LysP, pH alterations are sensed by CadC directly. Crystal structural analyses revealed a close proximity between two periplasmic cysteines, Cys208 and Cys272.
Substitution of Cys208 and/or Cys272 by alanine resulted in CadC derivatives that were active in response to only one stimulus, either lysine or pH 5.8. Differential in vivo thiol trapping revealed a disulfide bond between these two residues at pH 7.6, but not at pH 5.8. When Cys208 and Cys272 were replaced by aspartate and lysine, respectively, virtually wild-type behavior was restored indicating that the disulfide bond could be mimicked by a salt bridge.
A disulfide bond was found in the periplasmic domain of CadC that supports an inactive state of CadC at pH 7.6. At pH 5.8 disulfide bond formation is prevented which transforms CadC into a semi-active state. These results provide new insights into the function of a pH sensor.
Escherichia coli uses several strategies to maintain a neutral cytoplasmic pH in an acidic environment helping the bacterium to survive under this unfavorable condition. A number of genes is induced upon exposure of cells to low pH, among them the genes for the degradative amino acid decarboxylase systems adi, gad and cad. The Cad system consists of the cytoplasmic protein CadA and the transmembrane proteins CadB and CadC . CadA is a lysine decarboxylase that catalyzes decarboxylation of lysine to cadaverine whereby one proton is consumed resulting in a relief of the intracellular acid stress. The alkaline product cadaverine is concomitantly excreted by the lysine/cadaverine antiporter CadB [2, 3]. The genes cadA and cadB are organized in an operon [3, 4], which is under the control of the P Cad promoter. Expression of the cadBA operon is induced after external acidification, and simultaneous presence of extracellular lysine. CadC is the positive regulator of cadBA expression , and binds to two sites within the cadBA promoter .
cadC is located upstream of the cadBA operon and encodes a 58 kDa inner membrane protein. CadC, a member of the ToxR-like transcriptional activators , consists of a cytoplasmic N-terminal domain (amino acids 1-158), a single transmembrane domain (amino acids 159-187), and a periplasmic C-terminal domain (amino acids 188-512) [5, 8]. The cytoplasmic domain shows sequence similarity to the ROII-subgroup of DNA-binding domains of response regulators . However, contrary to prototypical response regulators  signal transduction in CadC functions without phosphorylation. Thus, CadC and all other ToxR-like proteins represent a one-component stimulus-response system.
Based on CadC derivatives with altered sensing properties due to single amino acid replacements within the periplasmic domain, it was suggested that this domain is the signal input domain . Recently, it became clear that CadC senses alterations of the external pH directly , but lysine is sensed only indirectly. The lysine-dependent regulation of CadC is exerted by an interplay with the lysine permease LysP, and it is proposed that in the absence of lysine, CadC is inactivated by an interaction with LysP .
Here, we investigated the role of the three cysteine residues in CadC. The best investigated member of the ToxR-like protein family, ToxR of Vibrio cholerae, contains two cysteines within the periplasmic domain. These cysteines were found to be involved in the formation of an intramolecular disulfide bond but also in the formation of intermolecular disulfide bonds between two ToxR molecules and between ToxR and a second transmembrane protein, ToxS [12, 13]. Although it was shown that ToxR binds to the DNA only in a dimeric form , ToxR oligomerization in vivo was independent of environmental changes , and thus evidence for the functional importance of the cysteines in ToxR is still lacking. Our studies indicated that a disulfide bond within the periplasmic domain of CadC is formed at pH 7.6, but these cysteines are in the reduced state at pH 5.8. These results give new insights into the switch between inactive and active states of a pH-responsive protein.
CadC of E. coli contains three cysteine residues, one in the transmembrane domain (C172), and two in the periplasmic domain (C208 and C272). Amino acid alignment of CadC from all available sequences indicated that C172 is found only in a few species, whereas the two periplasmic cysteines are well conserved in the order of Enterobacteriales (data not shown). In addition, the crystal structure of the periplasmic domain of CadC depicted a close proximity between C208 and C272  predicting an intramolecular disulfide bond. Thus, the role of the cysteines in CadC was studied in detail.
First, each cysteine in CadC was replaced with alanine, and the resulting derivatives CadC_C172A, CadC_C208A, CadC_C272A and CadC_C208A,C272A were used for complementation of the E. coli EP314 reporter strain (cadC::Tn10, cadA'::lacZ). β-Galactosidase activities were determined as a measurement for cadBA expression. CadC_C172A with a replacement of the cysteine in the transmembrane domain retained the activity pattern of wild-type CadC with induction of cadBA expression only at pH 5.8 in the presence of lysine (Figure 1). In contrast, replacement of cysteines at positions 208 and 272 in the periplasmic domain either alone or in combination resulted in CadC derivatives for which one stimulus was sufficient to activate cadBA expression (Figure 1). Specifically, cells expressing these derivatives induced cadBA expression at pH 5.8 regardless of the presence of lysine, and also at pH 7.6 when lysine was present. In general, β-galactosidase activities were significantly higher for these derivatives compared to the wild-type. Besides, a comparison of the activities in response to one or two stimuli revealed that the induction level significantly increased when cells expressing these derivatives were exposed to both stimuli (Figure 1). All CadC derivatives analyzed in reporter gene assays were produced and found to be membrane-integrated as the wild-type protein (Figure 1). In consequence, C208 and C272 are important for the regulation of CadC activity.
Influence of cysteine replacements in CadC on cadBA expression. Reporter gene assays were performed with E. coli EP314 (cadC::Tn10; cadA'::lacZ fusion) which was complemented with plasmid-encoded cadC or the indicated cadC derivatives. Cells were cultivated under microaerobic conditions in minimal medium at pH 5.8 or pH 7.6 in the presence or absence of 10 mM lysine at 37°C to mid-logarithmic growth phase, and harvested by centrifugation. The activity of the reporter enzyme β-galactosidase was determined  and served as a measurement for cadBA expression. Error bars indicate standard deviations of the mean for at least three independent experiments. To analyze production and membrane integration of the CadC derivatives, Western blot analysis of membrane fractions from E. coli BL21(DE3)pLysS transformed with plasmids encoding either wild-type or CadC derivatives was performed (inset). Each lane contains 25 μg of membrane protein (CadC derivatives are in the same order as in the graph). CadC was detected by a monoclonal mouse antibody against the His-Tag and an alkaline phosphatase coupled anti-mouse antibody.
In order to detect intermolecular disulfide bonds, membrane vesicles containing wild-type CadC or CadC derivatives with cysteine replacements were treated with copper phenanthroline, a Cys null crosslinker. Subsequent Western blot analysis revealed that in case of wild-type CadC and CadC with a single Cys at position 172, a fraction of the protein was transformed into an oligomeric form which might be related to the formation of an intermolecular disulfide bond at position 172 (data not shown). Since replacement of Cys172 was without effect on the CadC-mediated cadBA expression (Figure 1), it is concluded that an intermolecular disulfide bond is without functional importance for CadC.
To analyze whether a disulfide bond is formed in CadC, an in vivo differential thiol trapping approach with iodoacetamide and PEG-maleimide was used . For simplification, these studies were performed with CadC_C172A which contains only the two periplasmic cysteines. The method is based on the fact that both iodoacetamide and PEG-maleimide react only with free thiol groups. First, E. coli cells producing CadC_C172A were labeled with iodoacetamide during growth at pH 7.6 or pH 5.8. Subsequently, free iodoacetamide was removed, and all disulfide bonds were reduced by treatment with dithiothreitol (DTT). Free thiol groups were labeled with PEG-maleimide in a second step. In consequence, only cysteines that are present in an oxidized form and thus protected from iodoacetamide labeling in the first step, are labeled with PEG-maleimide resulting in a detectable increase of the molecular weight.
At pH 7.6 differential labeling of CadC_C172A clearly resulted in a labeling with PEG-maleimide (Figure 2). The band for unlabeled CadC decreased, and an additional higher molecular band appeared demonstrating labeling of C208 and C272 with PEG-maleimide (Figure 2a, lane 2). This additional band was only detectable when cells were treated with DTT (Figure 2a, lane 3 in comparison to lane 2). The PEG-ylated CadC_C172A runs as a smeared and broadened band which is probably due to the interaction between PEG and SDS . Addition of PEG-maleimide (regardless of the treatment with DTT) resulted in an additional labeling product that also appeared in cells producing the cysteine-free CadC. Therefore, this signal can be regarded as unspecific labeling product which might be related to a reactivity of maleimide with other residues (e.g., lysine or tyrosine) in CadC (Figure 2a, lanes 2, 3, and 7, 8). Labeling of CadC_C172A with PEG-maleimide implies that iodoacetamide was unable to react with the periplasmic cysteines. To prove that this result was due to the presence of a disulfide bond that protected the cysteines from labeling, CadC_C172A,C208A was subjected to the same labeling procedure (Figure 2a, lanes 4 and 5). This CadC derivative contains one cysteine that should be labeled with iodoacetamide in the first labeling step. As expected this derivative was hardly PEG-ylated under this condition (Figure 2a, lane 5). In contrast, this protein was completely PEG-ylated when iodoacetamide was omitted in the first step (Figure 2a, lane 4). The PEG-ylated products (Figure 2a, lanes 2 and 4) differed in size because of the different number of cysteines that were accessible for labeling. These data clearly demonstrate the presence of a disulfide bond between C208 and C272 in the inactive state of CadC at pH 7.6 (Figure 2b).
In vivo monitoring of the thiol/disulfide state of the periplasmic cysteines of CadC at pH 7.6 (a) and illustration of the results (b). (a) CadC_C172A, CadC_C172A,C208A or CadC_C172A,C208A,C272A (cysteine-free CadC) were overproduced in E. coli BL21(DE3)pLysS grown in phosphate buffered minimal medium at pH 7.6. To label free thiol groups irreversibly, 5 mM iodoacetamide was added directly to the living cells. After TCA precipitation and extensive washing, oxidized thiol groups were reduced by addition of 10 mM DTT in denaturing buffer. These reduced cysteines were then alkylated by addition of 10 mM PEG-maleimide. Samples were mixed with non-reducing SDS-sample buffer and 30 μg total cell protein were loaded onto 12.5% SDS-polyacrylamide gels. CadC was detected by Western blot analysis of the His-tagged proteins. Control experiments were done without DTT (lanes 3, 8) or PEG-mal (lanes 1, 6) or iam (lane 4). As a negative control the cysteine-free CadC derivative CadC_C172A,C208A,C272A was used. The iam control was performed with a CadC derivative that contains only one cysteine (CadC_C172A,C208A). iam = iodoacetamide, DTT = dithiothreitol, PEG = PEG-maleimide. (b) The results are schematically illustrated.
Since CadC becomes activated at low pH, the occurrence of the disulfide bond was also investigated under this condition (Figure 3). At pH 5.8 CadC_C172A was not labeled with PEG-maleimide (Figure 3a, lane 2). Addition of PEG-maleimide either in the absence or the presence of DTT produced only an unspecific band that was also observed for the cysteine-free CadC_C172A,C208A,C272A (Figure 3a, lanes 2, 3, and 7, 8). This result alludes to an efficient labeling of C208 and C272 with iodoacetamide in the first step, and implies that the periplasmic cysteines exist in a reduced form under acidic conditions. As a control, iodoacetamide was omitted and thereupon the typical PEG-maleimide labeling product appeared (Figure 3a, lane 4). Omittance of PEG-maleimide resulted in the disappearance of this band (Figure 3a, lane 5). These results reveal that PEG-ylation was possible under the used experimental conditions, but the cysteines in CadC_C172A were modified with iodoacetamide in the first step (Figure 3b). In conclusion, C208 and C272 are in a reduced form at low pH.
In vivo monitoring of the thiol/disulfide state of the periplasmic cysteines of CadC at pH 5.8 (a) and illustration of the results (b). (a) CadC_C172A or CadC_C172A,C208A,C272A were overproduced in E. coli BL21(DE3)pLysS grown in phosphate buffered minimal medium pH 5.8. The labeling procedure was essentially the same as described in Figure 2, with the difference that the alkylation time was prolonged. Control experiments were done without DTT (lanes 3, 8), or PEG-mal (lanes 1, 5, 6) or iam (lane 4, 5). As a negative control the cysteine-free CadC derivative CadC_C172A,C208A,C272A was used. iam = iodoacetamide, DTT = dithiothreitol, PEG = PEG-maleimide. (b) The results are schematically illustrated.
The results obtained with the labeling experiments indicate a disulfide bond under non-inducing conditions, but this bond is not formed at pH 5.8. In the next experiments we asked the question whether the disulfide bond could be mimicked by a salt bridge, which is strongly pH-dependent . Therefore, C208 and C272 were replaced by lysine and aspartate in both combinations possible. Under non-inducing conditions (pH 7.6) these amino acids should be in their charged form, and thus be able to form a salt bridge that mimics a disulfide bond. At low pH formation of a salt bridge might be prevented due to the protonation of asparate. Indeed, the induction profile supported by CadC_C208D,C272K was comparable to wild-type CadC (Figure 4). These data imply that in CadC_C208D,C272K the charged amino acids are able to form a salt bridge that takes over the function of the disulfide bond. In contrast, cells producing CadC_C208K,C272D exhibited a deregulated induction pattern (Figure 4). This result suggested that in this construct salt bridge formation was prevented and therefore the replacements of the cysteines against charged amino acids had the same effect as the disruption of the disulfide bond by alanine replacements.
Generation of a functional cysteine-free CadC by replacement of the disulfide bond forming cysteines with charged amino acids. Reporter gene assays were performed with E. coli EP314 (cadC::Tn10; cadA'::lacZ fusion) which was complemented with plasmid-encoded cadC or the indicated cadC derivatives. Cells were cultivated under microaerobic conditions in minimal medium at pH 5.8 or pH 7.6 in the presence or absence of 10 mM lysine at 37°C to mid-logarithmic growth phase, and harvested by centrifugation. The activity of the reporter enzyme β-galactosidase was determined  and served as a measurement for cadBA expression. Error bars indicate standard deviations of the mean for at least three independent experiments. Western blot analysis (inset - see Figure 1 for details) of all CadC derivatives (same order as in the graph) indicates production and membrane insertion.
Importantly, the wild-type like regulation pattern of CadC_C208D,C272K offered the unique opportunity to generate a functional cysteine-free CadC variant required as prerequisite for site-specific labeling studies in future. As expected, the regulation pattern of cells producing the cysteine-free derivative CadC_C172A,C208D,C272K was almost comparable to cells producing the wild-type protein (Figure 4). These data indicate that a salt bridge can take over the function of the disulfide bond in CadC.
CadC activity is regulated by the two stimuli pH and lysine. CadC derivatives with a replacement of the periplasmic cysteines by alanine were inactive at pH 7.6 in the absence of lysine (Figure 1). Obviously, the inhibitory effect of LysP on the CadC derivatives was strong enough to prevent cadBA expression at pH 7.6. However, it remained unclear, why these CadC derivatives activated cadBA expression at low pH in the absence of lysine despite of the inhibitory effect of LysP on CadC. Thus the question arose, whether the disruption of the periplasmic disulfide bond alters the interaction between CadC and LysP. To answer this question, the interplay between CadC and LysP was disturbed, simply by overproduction of LysP [11, 19]. It is known, that LysP overproduction lowers wild-type cadBA expression significantly (57% reduction) (Figure 5). In contrast, CadC_C208A,C272A-mediated cadBA expression was slightly affected by LysP overproduction at pH 5.8 (17%), but severely affected at pH 7.6 (59%) (Figure 5). These results imply that the interaction between LysP and CadC_C208A,C272A is weaker at pH 5.8 than at pH 7.6, and in general weaker in comparison to wild-type CadC. Moreover, the weakened interaction between LysP and CadC_C208A,C272A might also account for the general higher ß-galactosidase activities measured for all derivatives with Cys replacements at positions 208 and/or 272 (Figures 1 and 5).
Influence of LysP overproduction on CadC-mediated cadBA expression. Reporter gene assays were performed with E. coli EP314 (cadC::Tn10; cadA'::lacZ fusion) which was co-transformed with plasmid-encoded cadC or cadC_C208A,C272A and with a second plasmid carrying the lysP gene (pBAD33-lysP). Cells were cultivated under microaerobic conditions in minimal medium at pH 5.8 or pH 7.6 in the presence of 10 mM lysine at 37°C to mid-logarithmic growth phase, and harvested by centrifugation. When indicated, overproduction of LysP was induced by addition of 0.2% (w/v) arabinose. The activity of the reporter enzyme β-galactosidase was determined  and served as a measurement for cadBA expression. Shaded numbers display the degree of inhibition of cadBA expression by LysP overproduction. It should be noted that wild-type CadC activates cadBA expression only at pH 5.8. Error bars indicate standard deviations of the mean for at least three independent experiments.
In another experiment CadC_C208A,C272A was introduced into the ΔlysP reporter strain E. coli EP-CD4 (cadC::Tn10, cadA'::lacZ, ΔlysP). In a lysP- background, wild-type CadC activates cadBA expression in a lysine-independent, but pH-dependent manner [11, 19]. As expected, in the lysP- background, CadC_C208A,C272A induced cadBA expression lysine- and pH-independently revealing that LysP is responsible for the inhibition of CadC_C208A,C272A in the absence of lysine at pH 7.6 (data not shown). As discussed below, these experiments revealed that CadC without a disulfide bond is transformed into a semi-active state with respect to both the pH and the lysine stimuli.
The results described above led to the hypothesis that at physiological pH CadC contains a disulfide bond which is reduced at low pH. Opening and formation of disulfide bonds requires either the corresponding environment (oxidizing or reducing) or enzymes that catalyze these processes. Therefore, we analyzed whether periplasmic proteins known to be involved in formation and opening of disulfide bonds during the protein folding process such as the Dsb proteins  have an influence on CadC activation. Six gene deletion mutants were constructed lacking the disulfide bond-modifying proteins DsbA, DsbB, DsbC, DsbD, DsbG and CcmG (also known as DsbE). CcmG does not belong to the Dsb system, but is a membrane-anchored protein with a periplasmic thiol:disulfide oxidoreductase domain involved in cytochrome c biogenesis . DsbA is a disulfide oxidase responsible for the formation of disulfide bonds and is recycled by the membrane protein DsbB . DsbC is an isomerase that opens wrongly formed disulfide bonds and introduces the correct ones and as such also exhibits a reductase activity. DsbG is a non-essential isomerase that is able to substitute for DsbC, and seems to protect single cysteines from oxidation that are needed in a reduced state to be catalytically active . Both, DsbC and DsbG, are recycled by DsbD. While DsbB and DsbD are membrane proteins, DsbA, DsbC and DsbG are soluble proteins located in the periplasm.
Mutants of E. coli MG1655 each lacking a single dsb gene were grown at pH 5.8 and 7.6 in the presence of external lysine, and lysine decarboxylase (CadA) activity was determined as a measurement for the expression level of cadBA and thus of the functionality of CadC (Figure 6). All strains tested exhibited a pH-dependent regulation that was comparable to the wild-type strain, though the fold-induction differed slightly in some mutants. Under inducing conditions (pH 5.8, lysine) CadA activity was more than twice as high in the mutant MG1655ΔdsbA, lacking the disulfide oxidase DsbA, as in the wild-type strain MG1655 [specific CadA activity of 2.96 μmol/(min*mg protein) instead of 1.27]. The same holds true for MG1655ΔdsbB, lacking the recycling protein for DsbA, and MG1655ΔdsbD, lacking the recycling protein for DsbC [specific CadA activity of 2.41 and 2.82 μmol/(min*mg protein), respectively].
Influence of periplasmic disulfide oxidoreductases on cadBA expression. Single gene deletions of dsbA, dsbB, dsbC, dsbD, dsbG and ccmG were constructed in E. coli MG1655, and cadBA expression was monitored in the corresponding mutants by determination of CadA activity. Cells were cultivated under microaerobic conditions in minimal medium at pH 5.8 or pH 7.6 in the presence of 10 mM lysine. The specific CadA activity was determined  and is given in μmol/(min*mg protein). Error bars indicate standard deviations of the mean for at least three independent experiments.
Cysteines are one of the most rarely used amino acids in proteins of all organisms studied so far . Conserved cysteines usually play crucial roles in the structure and function of proteins because of their ability to form intra- and intermolecular disulfide bonds and to coordinate transition metal ions . Disulfide bonds are often linked to protein stabilization, ligand binding, dimerization and activation. Examples of this are the eukaryotic cytokine receptor GHR  and the OxyR transcriptional factor of E. coli . The ArcB sensor kinase is another example for the participation of disulfide bond formation during signaling [27, 28]. Each ArcB monomer contains two cytosolic cysteines. Quinone-dependent inhibition of ArcB autophosphorylation involves the formation of one or two intermolecular disulfide bonds under aerobic conditions. The V. cholerae activator of virulence gene expression ToxR contains two periplasmic cysteines that are important for the formation of an intramolecular disulfide bond as well as for the formation of intermolecular disulfide bonds between two ToxR molecules and between ToxR and ToxS [12, 13]. ToxR binds to the DNA only in a dimeric form , and dimerization of full-length ToxR was shown in vivo . However, an influence of environmental conditions (pH, osmolarity) on ToxR oligomerization and hence disulfide bond formation in vivo was not detected .
CadC contains three cysteines. Here, we show that cysteines located at positions 208 and 272 are important for the function of CadC, whereas C172 is dispensable. Two stimuli are needed to activate wild-type CadC: low pH and lysine. Replacements of C208 and/or C272 resulted in CadC derivatives that partially induced cadBA expression at pH 7.6 indicating the transformation of CadC into a semi-active state. This discovery and structural data, according to which these residues are in close proximity to each other , prompted experiments to monitor the redox state of the thiol groups of the periplasmic cysteines in CadC in vivo. Differential thiol trapping performed with CadC_C172A producing cells that were grown in minimal medium at pH 7.6 revealed that a disulfide bond is present in CadC. In contrast, at pH 5.8 the periplasmic cysteines of CadC were found to be in the reduced state. These data indicate that CadC contains a functionally important disulfide bond. It is important to note, that CadC is hardly a redox sensor. The differences in the cadBA expression level found for anaerobic and aerobic growth conditions are dependent on H-NS . Therefore, it is proposed that the disulfide bond in CadC provides structural support for the switch of the sensor between the inactive and active state. This assumption is supported by the location of Cys208 within a flexible loop in the N-terminal subdomain .
The question arose, how the disulfide bond might be formed and opened in vivo. Enzymes responsible for these processes might be the periplasmic disulfide oxidoreductases of the Dsb system. CadA activity as indication for cadBA expression was monitored in single dsb and ccmG deletion mutants. However, none of these deletions altered the CadC-mediated induction profile. In all deletion mutants induction of cadBA expression was prevented at pH 7.6, and CadA activity was significantly increased at low pH. These data imply that none of these proteins was essential for the formation or opening of the disulfide bond in CadC. It is worth mentioning, that we found an elevated CadA activity in the dsbA (encoding a disulfide oxidase), dsbB (encoding a protein that regenerates DsbA) and dsbD (encoding a recycling enzyme for an isomerase/reductase) deletion mutants. DsbA/DsbB are responsible for the introduction of disulfide bonds in newly synthesized proteins, thus their lack might support a higher probability of CadC molecules without a disulfide bond and thus the increased CadA activity. The role of DsbD in CadC activation remains unknown. Nevertheless, either these enzymes are functionally redundant, or the spontaneous oxidation by oxygen or low molecular compounds might be responsible for the formation of a disulfide bond in CadC. cadC belongs to the genes/operons with the shortest half-lives of the mRNA . Based on this result and our finding of a transient activation of CadC , we speculate that there is a rapid turnover of CadC and that the disulfide bond is preferentially introduced during de novo synthesis of CadC. The periplasm is accessible for oxygen and therefore allows the spontaneous oxidation of two neighboring cysteines in proteins [32, 33]. Expression of the cadBA operon is induced at low pH, and the induction level is higher in the absence of oxygen . Under these conditions the oxidation of cysteines to cystine is minimized due to the lack of oxygen as well as the surplus of protons which prevents the formation of thiolate anions, the prerequisite for disulfide bond formation . Thus, this shift in the external conditions already dramatically reduces the probability to form a disulfide bond in CadC. Based on these results it is suggested that under non-inducing conditions (pH 7.6) a disulfide bond in the periplasmic domain holds the sensor in an inactive state. Under inducing conditions (pH 5.8) formation of the disulfide bond is either prevented or the disulfide bond is opened by a still unknown mechanism. Concomitantly, CadC undergoes conformational changes due to the protonation of negatively charged amino acids located in a patch at the CadC dimer interface .
This proposal is in accordance with the finding that the disulfide bond could be mimicked by a salt bridge. When C208 was replaced with an aspartate and C272 with a lysine, a CadC derivative was generated that supported cadBA expression comparable to the wild-type protein. Functional substitution of a disulfide bond by a salt bridge in CadC requires formation of the salt bridge at pH 7.6, which is conceivable (aspartate deprotonated, lysine protonated), and an opening of the salt bridge, which might depend on the protonation of aspartate at low pH [36, 37]. In contrast, a CadC derivative in which the cysteines were replaced by the same charged amino acids but at opposite positions (CadC_C208K,C272D) caused deregulation of cadBA expression. It is suggested that a salt bridge was not formed in this derivative due to an unfavorable orientation of the amino acid side chains to each other.
The results obtained in this study illuminate the activation mechanism, specifically the sequential events to transform CadC into an active form (Figure 7). Derivative CadC_C208A,C272A induced cadBA at pH 7.6, however, its activity further increased at pH 5.8. Thus, the lack of the disulfide bond seems to be only one part of the pH-dependent structural transitions in CadC. Whether reduction of the cysteines is a prerequisite for or a consequence of additional conformational changes cannot be decided yet. Nevertheless, CadC without a disulfide bond is held in a semi-active state. This derivative also induces cadBA expression at low pH regardless of the lysine concentration. This result suggests that the interaction between LysP and a CadC derivative without a disulfide bond is weaker in comparison to the wild-type. In agreement, CadC lacking the periplasmic cysteines is hardly subject to LysP-mediated inhibition in cells that overproduce LysP. Our experimental data also revealed that the interaction between LysP and CadC is stronger at pH 7.6.
Model of the lysine- and pH-dependent activation of wild-type CadC and CadC_C208A,C272A. The different transcription activities are indicated by the arrows below CadC. Under non-inducing conditions (no lysine, pH 7.6) CadC-mediated cadBA expression is inhibited by two mechanisms, the interaction with LysP and a disulfide bond in the periplasmic domain. CadC with a disulfide bond remains inactive even when the interaction with LysP is released in the presence of lysine (lysine, pH 7.6). A shift to low pH causes conformational changes and prevents formation of a disulfide bond (lysine, pH 5.8). In the absence of lysine, CadC activity is blocked by the interplay with LysP (no lysine, pH 5.8). In CadC_C208A,C272A formation of a disulfide bond is prevented by amino acid replacements. As consequence, this derivative displays semi-active states at pH 7.6 in the presence of lysine (lysine, pH 7.6), and at pH 5.8 in the absence of lysine (no lysine, pH 5.8). See text for further details.
Based on these results, a two step activation mechanism for CadC is proposed (Figure 7). Under non-inducing conditions (no lysine, pH 7.6) CadC-mediated cadBA expression is inhibited by two mechanisms. At pH 7.6 a disulfide bond is formed, and CadC is in an inactive form. Moreover, CadC is inhibited through the interplay with the lysine permease LysP in the absence of lysine . CadC with a disulfide bond remains inactive even when the interaction with LysP is released in the presence of lysine (lysine, pH 7.6). Exposure of CadC to low pH is accompanied by conformational changes and reduction of the cysteines resulting in an active CadC (lysine, pH 5.8). Alternatively, at low pH in the absence of lysine, CadC is still locked in an inactive conformation due to the interplay with LysP (no lysine, pH 5.8). The presence of lysine suspends the interaction with LysP, and CadC is transformed into the active state (lysine, pH 5.8).
In CadC_C208A,C272A formation of a disulfide bond is prevented by amino acid replacements (Figure 7). As consequence, this derivative displays semi-active states at pH 7.6 in the presence of lysine (lysine, pH 7.6) or at low pH in the absence of lysine (no lysine, pH 5.8). Additional pH-dependent conformational changes or the presence of lysine are required to fully activate this CadC derivative (lysine, pH 5.8).
Previously, it was proposed that the two stimuli, lysine and low pH, affect CadC activation independently from each other . Here, we gained new insights into the molecular mechanism how CadC processes these stimuli, particularly that a disulfide bond is involved in the function of CadC.
Strains and plasmids are listed in Tables 1 and 2. E. coli JM109 served as carrier for the plasmids described. E. coli BL21(DE3)pLysS was used for expression of cadC and cadC derivatives from the T7 promoter. E. coli EP314 and EP-CD4 were complemented with plasmids (pET16b) encoding cadC and its derivatives, and used for cadBA transcriptional analysis. E. coli EP314 and EP-CD4 carry a cadA'::lacZ fusion and an inactivated cadC. Additionally, EP-CD4 is also lysP-. Overproduction of LysP was performed in E. coli EP314 transformed with plasmid pBAD33-lysP by inducing the arabinose promoter with 0.2% (w/v) arabinose. E. coli MG1655 was used for construction of gene deletion strains. E. coli strains were grown in Luria-Bertani (LB) medium  for strain maintenance and protein overproduction. To probe signal transduction in vivo, cells of E. coli EP314 transformed with the indicated plasmids were grown in minimal medium ; the phosphate buffer of the medium was adjusted to either pH 5.8 or pH 7.6. Lysine was added at a concentration of 10 mM. For selection of plasmid-containing cells appropriate antibiotics were added at concentrations of 100 μg/ml (ampicillin sodium salt), 50 μg/ml (kanamycin sulfate) and 34 μg/ml (chloramphenicol).
Site-directed mutants are designated as follows: The one letter code is used, followed by a number indicating the position of the amino acid in wild-type CadC.
The sequence is followed by a second letter denoting the amino acid replacement at this position.
All cadC derivatives were constructed by polymerase chain reaction (PCR) with mismatch primers either by single step or by two step PCR . To facilitate construction, a cadC gene with two additional unique restriction sites was employed . All site-specific mutations were directed by synthetic oligonucleotide primers containing the required nucleotide exchanges. PCR fragments were cloned into the expression vector pET16b with the restriction enzymes NdeI and BamHI so that all constructs carried the sequence encoding an N-terminal His-Tag of 10 histidine residues. E. coli EP-CD4, E. coli MG1655ΔdsbA, E. coli MG1655ΔdsbB, E. coli MG1655ΔdsbC, E. coli MG1655ΔdsbD, MG1655ΔdsbG and MG1655ΔccmG were constructed by deleting the genes lysP, dsbA, dsbB, dsbC, dsbD, dsbG and ccmG, respectively, via the Quick & Easy E. coli Gene Deletion Kit (Gene Bridges) according to the manufacturer's instructions.
The thiol/disulfide state of the periplasmic cysteines of CadC was monitored in vivo by differential thiol trapping according to . The procedure was modified as follows: E. coli BL21(DE3)pLysS carrying one of the plasmids pET-CadC-C172A, pET-CadC-C172A,C208A or pET-CadC-C172A,C208A,C272A was grown in phosphate buffered minimal medium with a pH of 7.6 or 5.8 to an OD600 of 0.5. Subsequently, overproduction of the CadC derivatives was induced by addition of 0.5 mM IPTG. After an additional hour of growth at 37°C, the OD600 was adjusted to 1, and 5 mM iodoacetamide (dissolved in 0.1 M Tris) was added to 1 ml cell suspension. At pH 7.6, incubation was performed for 15 min (37°C), at pH 5.8 the incubation time was prolonged to 150 min to compensate the lower alkylation rate of iodoacetamide at low pH. This first alkylation procedure irreversibly modified all free thiol groups directly in the living cells. Subsequently, cells were harvested into 100 μl ice-cold 100% (w/v) trichloric acid (TCA) and stored on ice for at least 30 min. The TCA treated cells were centrifuged (16.000 g, 4°C, 15 min), and the resulting pellet was washed with 200 μl of ice-cold 10% (w/v) TCA followed by a wash with 100 μl of ice-cold 5% (w/v) TCA. The supernatant was removed completely, and the pellet was resuspended in 100 μl of denaturing buffer [6 M urea, 200 mM Tris-HCl (pH 8.5), 10 mM EDTA, 0.5% (w/v) SDS] supplemented with 10 mM DTT to reduce disulfide bonds. After one hour of incubation in the dark (37°C, gentle agitation at 1300 rpm), 10 μl ice-cold 100% (w/v) TCA was added, and the sample was stored on ice for at least 30 min. After centrifugation, the resulting pellet was again washed with 200 μl of ice-cold 10% (w/v) TCA followed by a wash with 100 μl of ice-cold 5% (w/v) TCA. Finally, the pellet was resuspended in 50 μl of denaturing buffer containing 10 mM PEG-maleimide (Iris Biotech GmbH, Marktredwitz/Germany) to alkylate all newly reduced cysteines. The reaction (37°C, gentle agitation at 1300 rpm, in the dark) was stopped after one hour by addition of 5 μl ice-cold 100% (w/v) TCA. After precipitation on ice (30 min) and centrifugation, the pellet was washed first with 100 μl of 10% and then with 50 μl of 5% ice-cold (w/v) TCA. After removing the TCA, the pellet was washed twice with 500 μl acetone and resuspended in 50 μl denaturing buffer. Samples were mixed with non-reducing SDS-sample buffer and loaded onto 12.5% SDS-polyacrylamide gels . CadC was detected by Western blot analysis .
For the detection of intermolecular disulfide bonds, wild-type CadC and all available CadC derivatives with Cys replacements (CadC_C172A; CadC_C208A; CadC_C272A; CadC_C172A,C208A; CadC_C172A,C272A; CadC_C208A,C272A; CadC_C172A,C208A,C272A) were overproduced in E. coli BL21(DE3)pLysS, and membrane vesicles were prepared . CadC-containing membrane vesicles [1 mg protein/ml in TG-buffer, 50 mM Tris/HCl, pH 7.5; 10% (v/v) glycerol] were treated with 0.2 mM copper phenanthroline at 25°C for 30 min. The reaction was stopped by addition of 10 mM EDTA. Samples were mixed with non-reducing SDS-sample buffer and loaded onto 7.5% (w/v) SDS-polyacrylamide gels . CadC was detected by Western blot analysis .
Signal transduction activity of different CadC derivatives in vivo was probed with a β-galactosidase based reporter gene assay as previously described . Using a pET-based vector in combination with the reporter strain E. coli EP314 that does not possess a T7 polymerase resulted in a low expression that was sufficient to allow complementation but did not lead to overproduction of CadC which would result in stimulus-independent cadBA expression. β-galactosidase activity was determined from at least three independent cultures, and is given in Miller units (MU) calculated as described . The activity of the lysine decarboxylase CadA as a measurement for cadBA expression was determined according to  with the following changes: for the assay cells corresponding to an optical density of 1 (600 nm) were resuspended in 20 mM potassium phosphate buffer (pH 5.6) and lysed by the addition of chloroform. One unit is defined as 1 μmol decarboxylated lysine produced per minute and specific activities were calculated for 1 mg of protein [μmol/(min*mg)]. Insertion of the CadC derivatives into the cytoplasmic membrane was analyzed after overproduction of CadC, isolation of membrane vesicles and subsequent Western blot analysis as previously described [11, 45].
This work was supported by the Deutsche Forschungsgemeinschaft (JU270/5-3 and Exc114/1). We thank Teresa Friedrich for the construction of E. coli MG1655ΔdsbA, MG1655ΔdsbB, MG1655ΔdsbC and MG1655ΔdsbD and Korinna Burdack for technical assistance.
LT, CK and KJ designed research experiments; AD performed experiments; LT performed experiments and analyzed data. LT and KJ wrote the manuscript. All authors have read and approved the final manuscript.


text:
The best car for 2021 is also the cheapest: Dacia's new £8,000 Sandero is named 'Car of the Year' - see which other models are recommended to drivers
Drivers who fall behind on car finance payments can have their vehicles repossessed by lenders from February as FCA's Covid-19 ban ends
Just remove a zero! Take a look around the cheapest house for sale in Britain which costs just £30,000 - or a TENTH of a typical property
Santander joins mortgage lenders turning off the tap for self-employed with new 40% deposit requirement
Free data for home schooling, help for vulnerable customers and WiFi vouchers: What is your telecoms firm doing to help in lockdown?
BUSINESS LIVE: Economy contracts 2.6% in November; Crash in hospitality sales; Meggitt exercises caution
Halfords benefits from lockdown cycle boom with bike sales up 34%
Pubs, restaurants and bars have 'dreadful' Christmas as sales tumble over 70% after already being hammered by the November lockdown
Tycoons cash in £250m as shares in ecommerce business Hut Group surge following last year's London float
British Gas owner Centrica manages to curb customer exits for the first time in years with 6.9m clients staying put
Lockdowns to wipe £1bn off Primark sales... but Boohoo booms as internet shoppers stock up on casualwear
Budget airline Norwegian axes its long-haul network, leading to 1,100 job losses at Gatwick Airport
Brexit casts a cloud over Tesco's festive feat as delivery woes hit Ireland
ALEX BRUMMER: Paris plays the food gambit as Canadian convenience store chain makes £14bn bid for Carrefour
Beyond Bitcoin: Number two crypto Ethereum is climbing faster and gave rise to the 'DeFi' industry that's netted some investors up to 11,000% in a year
Bitcoin bubble: Can you bank on the cryptocurrency? Five crucial questions answered before you dive in
Christine Lagarde demands action on bitcoin 'funny business' days after the UK's City watchdog issues warning around the cryptocurrency
Savers will be offered four simple ways to invest and draw on pension pots: How will this work, could it be better and what will it cost?
GRACE ON THE CASE: Our travel provider is taking the Mickey! Why haven't we received a refund for our dream £8,000 Disney World trip?
Are we about to see a burst of big ticket inflation? SIMON LAMBERT on why treating ourselves in 2021 is likely to be expensive
House prices set to fall 'significantly' in the coming months as property market braces for the end of stamp duty holiday and furlough
After yet another humbling U-turn... Why does Bank of England boss Mark Carney keep getting it SO wrong?
His six-year tenure at the Bank of England has divided opinion
Admited he was too gloomy about the immediate impact of the Brexit vote
By Hugo Duncan for the Daily Mail
On the day of his appointment four years ago, Mark Carney was hailed by George Osborne as 'the outstanding central banker of his generation'.
He was the rock star banker who said he was coming to 'where the challenge is greatest' by swapping the Bank of Canada for the Bank of England.
But his tenure at the Old Lady of Threadneedle Street, which will now run for six years to June 2019, has divided opinion – particularly since the EU referendum campaign.
Stage struck: Mark Carney was the rock star banker who said he was coming to 'where the challenge is greatest'
And yesterday the governor was forced to admit he was far too gloomy about the outlook for the UK following the Brexit vote.
To his critics, it was just the latest embarrassment since the 51-year-old started the job.
ALEX BRUMMER: Love bombing of Mark Carney by PM and... As the Bank of England rethinks post-Brexit GDP prospects,... Mark Carney is a man under fire, but his departure would... ALEX BRUMMER: Mark Carney lives up to nickname as...
The whopping pay deal
As the first foreign governor in the Bank's three-century history, he arrived in London in July 2013 with his wife Diana and four daughters to great fanfare. He took the Tube to work, arriving before 7am. But his pay – a thumping £874,000 a year, including a £250,000 housing allowance – had already raised eyebrows.
His flagship policy
The Canadian's first big decision as governor, in August 2013, was to launch his flagship policy of 'forward guidance' as he sought to kick-start an economic recovery.
The idea behind this was to reassure households and businesses over the future so they could plan accordingly.
He began by saying interest rates, which had been at 0.5 per cent since March 2009, would not rise until unemployment had fallen from the level then of 7.8 per cent to 7 per cent – something he did not expect to happen until late 2016. Unemployment in fact fell below 7 per cent in early 2014 (and has now reached 4.9 per cent), but interest rates remained unchanged.
Previous role: Mark Carney was Governor of the Bank of Canada before he took over the top job in the UK
The unreliable boyfriend
In a speech at Mansion House in the heart of the City in June 2014, the governor conceded that 'growth has been much stronger and unemployment has fallen much faster' than expected.
This, he said, had triggered 'great speculation about the exact timing of the first rate hike'.
It was widely thought that rates would rise in the first half of 2015, but in a fresh attempt to offer some sort of guidance, Carney said: 'It could happen sooner than markets currently expect.'
The comments sent shockwaves through the financial markets and drew criticism from MPs, with one famously accusing the Bank, and by extension the governor, of behaving like an 'unreliable boyfriend' for blowing hot and cold over when rates would rise.
Rather than giving certainty, it was the first sign that Carney's flagship policy could actually create confusion.
New rate rise warning
Despite his comments at Mansion House the previous summer, and solid economic growth, by July 2015 rates were still frozen at 0.5 per cent, but Carney again suggested this would soon change.
In another headline-grabbing speech, this time at Lincoln Cathedral, he said the decision over when to raise rates 'will likely come into sharper relief around the turn of this year'.
...and then a U-turn
In January 2016, with a new year under way, Carney conceded that there would be no interest rate hike any time soon.
'The year has turned, and, in my view, the decision proved straightforward: now is not yet the time to raise interest rates,' he said.
With speculation mounting that there would be no change in interest rates until 2017, the Canadian said the decision would 'depend on economic prospects, not the calendar'.
Carney also revealed that he may serve a full eight-year term as governor, having initially agreed to do the job for just five.
He told MPs that he would make a decision by the end of the year, but was urged by Andrew Tyrie, chairman of the Treasury Select Committee, to decide as soon as possible. 'The sooner you feel able to give us this clarity, the better,' he said.
Accountable: Mark Carney giving evidence to the Economic Affairs Committee at the House of Lords
Another flip-flop
Suddenly faced with questions in February 2016 over whether interest rates could in fact be cut, Carney was asked if the next move was still likely to be up.
'Absolutely,' he said. 'More likely than not the next move in rates is up.'
But just weeks later Carney appeared to have changed his mind, and said: 'We could cut interest rates towards zero.'
Joins Project Fear
With the EU referendum campaign in full swing in May, Carney warned that the vote was 'the most immediate and significant risk' for the UK economy.
Incredibly for a sitting central bank governor, he claimed that a vote to leave could lead to a 'technical recession'.
His comments sparked fury among Brexit supporters as Osborne – the architect of Project Fear – said the intervention was 'a very big moment in the debate about the economic costs of leaving the EU'.
His Brexit gloom
On June 24, the day after the referendum, Carney addressed the nation just hours after David Cameron announced he was standing down as prime minister.
With the financial markets in turmoil, Carney warned of 'volatility' but insisted: 'We are well prepared for this.'
He offered to pump an extra £250bn into the financial system to stabilise the markets and said 'we will not hesitate to take any additional measures required' to keep the economy on track.
Working together: George Osborne with Mark Carney, both of whom were accused of being too gloomy about the impact of Brexit on the UK economy
Then, in August, the Bank cut interest rates to a new low of 0.25 per cent and warned that a further cut to 0.1 per cent was likely before the end of the year.
It also restarted the money printing presses, pledging to pump another £170bn of emergency cash into the economy.
Although Carney ditched his warnings of recession, the Bank said that the UK 'is likely to see little growth in the second half of this year' and pencilled in expansion of just 0.1 per cent in the third quarter and none at all in the fourth.
And crucially, the Bank slashed its growth forecast for next year from 2.3 per cent to 0.8 per cent.
Changing forecasts: The August inflation report (left) predicted much slower GDP growth than the November report (right) published this week
Growth surprises
By October, official figures showed the economy actually grew by 0.5 per cent in the third quarter – five times faster than predicted by the Bank.
After days of frenzied speculation over his future, Carney said that he will stay at the Bank until the end of June 2019, extending his term by one year, rather than until 2021 as many had thought.
Changing expectations: The Bank of England now expects inflation to rise more sharply than it had previous thought (Source: Bank of England)
Now more humiliation
Carney was yesterday forced to admit that he was far too gloomy about the impact of the Brexit vote on the economy.
The Bank raised its growth forecasts for 2016 from 2 per cent to 2.2 per cent and for 2017 from 0.8 per cent to 1.4 per cent. That was the biggest upgrade to a growth forecast the Bank has ever made.
The Bank also resisted another cut in interest rates, saying the latest 'guidance' it offered in August has now 'expired'.
Carney even warned rates may have to rise to combat inflation.
To many observers, it was just the latest U-turn from the 'unreliable boyfriend'.
Why does Bank of England boss Mark Carney keep getting it SO wrong?


Director and Photographer Rodney Rascona is back and forward shooting in Cambodia with RED DIRT ROAD, INC.
The name "Red Dirt Road" comes from the road that connects Phnom Penh with the village of Tramung Chrum, where the Red Dirt Road organization first started. This is the road that young women from the village travel when they go to work in the garment factories. It is a long, dusty, dirt road that looks red under the scorching Cambodia sun. It is the hope of the organization that one day, young women will not have to travel the 'red dirt road' to endure working in the garment factories. That they will be self-sufficient women able to help their villages become sustainable communities.
Rodney's latest film explores sisterhood spanning across the globe, where women in First World countries play an active, responsible role for women in the Third World in order to enjoy choices, enhanced education and applied business skills in achieving their goals for a better life.
To know more about the film click in the name above or HERE.
The footage is now with editors with the expertise at WORK POST LTD and below is a selection of images from the trip.


text:
Lost treasures of egypt narrator
How to stop apple music playing in background
What is bongo bingo youtube
Live dealer online casino australia
Free vip betting tips app
Head tennis 2 player
Pokerstars pc requirements
What does ceo mean on tiktok
How to make a good volcano eruption
Icsi login uk
Grand casino hinckley poker room
Which qb has the most game winning drives
Live casino jackpot 247
Slot russian rock band
Gambling and information theory
These 10 Restaurants Serve The Best Grits In Georgia.
Georgia might be known for their peaches nationwide, but the local cuisine is much more expansive than just fruit. Specializing in cooking up some of the best comfort food you'll ever have the pleasure of eating, the Southern cooking here is so delicious you'll constantly be looking forward to your next meal. A few of the more popular dishes include grits, Brunswick stew, fried chicken.
To Participate in the Georgia Registry of Immunizations (GRITS) INSTRUCTIONS: 1. Complete the Enrollment information below for your organization. 2. If applicable, Complete an Add Site form for EACH ADDITIONAL Site location associated with your organization. 3. Read the Confidentiality Policy and keep for your record and review with any staff you assign to access GRITS. 4. Read and sign the.
From grits to UGA football to the Atlanta Opera, legendary humorist and columnist Lewis Grizzard wrote about it all. On Nov. 7, 2019, Grizzard, one of Atlanta's most beloved columnists, will be.
GRITS FAQs: This provides some of the most frequently asked questions and answers that Healthcare Professionals have inquired upon for GRITS GRITS Imm Record Request Form: Request for State of Georgia Official Immunization Record. Opt-Out of Registry Form: This form is required to allow an individual to request that a person's Immunization history be removed from the registry and no further.
Georgia also recognizes a state fruit, a state vegetable, and a state crop). All State Foods. Grits are a unique southern food first prepared by native Americans centuries ago. Southern grits are bits of ground corn or hominy prepared as a simple breakfast dish or incorporated into gourmet recipes. Corn is a prominent Georgia crop grown.
WARWICK, GA (WALB) - A popular south Georgia festival has been canceled this year. The National Grits Festival in Warwick will not be held because its long time sponsor backing out of the annual.
Grits definition, coarsely ground hominy, boiled and sometimes then fried, eaten as a breakfast dish or as a side dish with meats. See more.
The act of shitting in diarrhea form on a womans chest and smearing it all over her boobs and then titty fucking her. Fraternal twin to the Cleveland Steamer.
The GRITS in 60 seconds video provides a brief overview of the features and benefits of the Green Revolving Investment Tracking System, and a longer walkthrough reviews the Dashboard, Project Details page, Library, and Reporting feature in greater detail. Go to Videos. Webinars. The Sustainable Endowments Institute offers live webinars to demonstrate how GRITS is used. If you'd like to.
GRITS stands for the Georgia Registry of Immunization Transactions and Services. It was created under a 1996 state law requiring mandatory reporting of vaccinations given to children and teens.
The official website of the Georgia Pharmacy Association. This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License. Site design and construction by Andrew Kantor.
Georgia Registry of Immunization Transactions and Services. Use the GRITS system to access information and generate documentation only as necessary to properly conduct the administration and management of immunizations. Safeguard user ID and passwords granted through the GRITS system in accordance with generally accepted standards of system security management. Deny use of user ID or.
Retention: 10 years after the client reaches age of majority (18 in Georgia) or 28 years from date of last service. Children's High-Risk Screening and Case Management Records (Babies Can't Wait, Children First) Description: All documents relating to health services provided to Babies Can't Wait and Children First clients (0427-019) Retention: 5 years from date of last service; for Children.
Find Grits in Ga on Hotfrog. Get reviews and contact details for each business including phone number, postcode, opening hours and photos.
GRITS: Girls Riding in the Streets: GRITS: Georgia Rails Into Trails Society: GRITS: Godly Revelations in Today S: GRITS: Girls Riding in the South: GRITS: Gang Related Information Tracking System: GRITS: Girls Reading in the South: GRITS: Greater Raleigh Intrepid Tandem Society: GRITS: Gender Researchers in Tourism Studies: GRITS: Girls Racing.
The official prepared food of Georgia is grits, so it comes as no surprise that the best grits in Atlanta are some of the creamiest, tastiest, and most well-prepared that you'll get anywhere in the country. This unique Southern food is a must-try if you're travelling through the Southern United States. More specifically, it's a dish you absolutely have to sample if you're visiting.
Grits Cafe will open for outdoor dining on a closed Johnston Street for the next two weeks after the city approved it Monday. The city is allowing the popular restaurant, closed for dine-in since March, to block Johnston Street on the square from Lee Street to Jackson Street from 10 a.m.-10-p.m. Tuesdays-Saturdays from June 16-27 so Grits can use the street for dining.
These grits are produced in the mountains of North Georgia, where the process of making the grits provides them with a unique taste. A stone-grinding process that is rarely found in modern grain.
About me. I am a Senior Autopilot Planning and Controls Engineer at Tesla. I graduated from Georgia Tech in 2018 May with a Ph.D. degree in ECE supervised by Dr. Magnus Egerstedt.My Ph.D. research interests lie in the intersection of control theory, robotics, and machine learning.More specifically, my Ph.D. research focuses on the provably safe collaboration of multiple robots (quadrotors and.
The Georgia Registry of Immunization Transactions and Services, better known as GRITS, is the Immunization Information System developed to comply with Georgia Law (OCGA 32-12-3.1). Immunization registries are confidential, computerized information systems that contain information.
Ultimate Guide to Visiting River Street in Savannah, GA; TIP: Parking is plentiful in downtown Savannah, but my top place to park when visiting River Street is in Emmet Park off o.
If anthills are located where they cause problems, feed the ants grits. That's right — grits, the breakfast food. Sprinkle some dry grits around the anthill. The ants will eat the grits, drink.
When ready to mill, we belt up the millstones, fan suction system, and grits separator, and step up the speed of the water wheel, which drives all the machinery through a series of gears, flats belts, line shafts, and pulleys. The millstones we use are a set of 42-inch horizontal granite buhrs manufactured by the R.D. Cole Company of Newnan, Georgia, around the turn of the century. The granite.