Here are the latest developments in the coronavirus crisis.

Here is the latest news from around the world.

It is at times like these that we as a country rely on the relevant stakeholders to take care of our people and put into action the promises made in the preamble of our Constitution, particularly where it is stated that through our freely elected representatives, the quality of life of all citizens is to be improved.

The Economic Injury Disaster Loan program is now limiting both the amount and recipients it will consider for the emergency loans.

Disneyland (Strategy)

Walt Disney World

The Amateur Traveler talks to Zoë Dawes from TheQuirkyTraveler.com about England’s Lake District. This beautiful pastoral area was made famous by the poets and artists in the romantic period, most notably William Wordsworth. Not far from industrial Manchester, the lake district is still a refuge for hikers and and other tourists tucked away in northern England. Zoë describes some of her favorite hikes like the coffin trail along the shores of Lake Windermere and favorite villages like Ambleside.

Hear about hiking the Kings Trail (Kungsleden) in Swedish Lapland as the Amateur Traveler talks to Agata from nullnfull.com about this northern and rugged portion of Sweden. Agata says, “Lapland is all about nature and wilderness”.

The health expert who helped shaped Britain’s lockdown policies in response to the coronavirus has resigned after admitting he broke his own rules by having his married lover visit him at his home. Professor Neil Ferguson, 51, of Imperial College in London was a prominent member of the Scientific Advisory Group for Emergencies. The Telegraph…

The post Scientist Behind Social Distancing Breaks Own Rules To Cheat with Married Woman, Resigns appeared first on The Western Journal.

Eleven members of the Secret Service have tested positive for COVID-19, according a new report. Yahoo News reported Friday it has seen Department of Homeland Security documents which show that 11 individuals currently have the virus and that 23 members of the Secret Service have recovered from the disease. Another 60 employees of the agency…

The post Report: Secret Service Discovers Nearly a Dozen Coronavirus Cases in Its Own Ranks appeared first on The Western Journal.

At least 65,116 individuals have been "forcibly disappeared" by the Syrian government, according to a new report by Amnesty International.

The coronavirus pandemic has many of us nostalgic for a simpler time — maybe when we were teenagers and/or watching our favorite early oughts sitcoms. 

Actress Cobie Smulders seems to be feeling that itch, and sensed we are too. The former How I Met Your Mother actress decided to recreate her character Robin Scherbatsky's hit song from the series, "Let's Go to The Mall."

For those who need a refresher, "Let's Go to The Mall" was released during Robin's short career as a pop star (under the name Robin Sparkles) in her homeland of Canada. While it has all the flair of an '80s banger — jelly bracelets, worshipping the mall — it was released in the '90s because apparently that's when '80s culture reached Canada.  Read more...

Work(out) From Home is a weekly column where we review smart fitness machines and apps in the wake of the coronavirus outbreak. Thanks to technology, there are still plenty of ways to exercise if your gym is closed. Read more...

Data Governance sounds like a candidate for the most boring topic in technology: something dreamed up by middle-managers to add friction to data scientists’ lives. The funny thing about governance, though, is that it’s closely related to data discovery. And data discovery is neither dull nor additional friction; it’s an exciting process that enables great […]

Yesterday saw the online premier of a mini-episode of a new animated comic series based on the classic Gilbert Shelton underground comic, the Fabulous Furry Freak Brothers. As a hippie wannabe teen in the 70s, this (and Zap! Comics) was everything to me.

In 1969, life in San Francisco consists of free love, communal living, and political protest. Freewheelin’ Franklin Freek (Harrelson), Fat Freddy Freekowtski (Goodman), Phineas T. Phreakers (Davidson) and their mischievous, foul-mouthed cat, Kitty (Haddish) spend their days dodging many things —- the draft, the narcs, and steady employment -– all while searching for an altered state of bliss.

But after partaking of a genetically-mutated strain of marijuana, the Freaks wake up 50 years later to discover a much different society. Quickly feeling like fish out of water in a high-tech world of fourth-wave feminism, extreme gentrification and intense political correctness, the Freaks learn how to navigate life in 2020 -— where, surprisingly, their precious cannabis is now legal.

OK, sounds good. But is it? If the reaction to the first mini-episode is any indication, maybe the Freaks should have remained in their drug-induced coma. As one Facbooker commented: "Get yourself a collected set of the original comic and skip this drivel!"

Read the rest

Luis Spencer Freitas, digital marketing director at Pernod Ricard USA, explains what will drive greater innovation in display advertising next year.

It can scarcely be denied that the supreme goal of all theory is to make the irreducible basic elements as simple and as few as possible without having to surrender the adequate representation of a single datum of experience.

—Albert Einstein, “On the Method of Theoretical Physics” (1934)

Context: Last week, I pinched off one of my typically woolly emails in response to an acquaintance whom I admire. He’s a swell guy who makes things I love, and he'd written, in part, to express concern that my recent Swift impersonation had been directed explicitly at something he'd made. Which, of course, it hadn’t—but which, as I'll try to discuss here, strikes me as irrelevant.

To paraphrase Bogie, I played it for him, so now I suppose I might as well play it for you.

(n.b.: Excerpted, redacted, munged, and heavily expanded from my original email)

There are at least a couple things that mean a lot to me that I'm still just not very good at.

• Make nuanced points in whatever way they need to be made; even if that ends up seeming “un-nuanced”
• Never explain yourself.

I want to break both these self-imposed rules privately with you here. [Editor’s Note: Um.] Because, I hope to nuance the shit out of some fairly un-nuanced points. And, to do that, I'll also (reluctantly) need to explain myself. But, here goes.

First [regarding my goofing on “distraction-free writing environments”] I think there are some GIANT distinctions at play here that a lot of folks may not find nearly as obvious as I do:

1. Tool Mastery vs. Productivity Pr0n – Finding and learning the right tools for your work vs solely dicking around with the options for those tools is just so important, but also so different. And, admittedly, it’s almost impossible to contrast those differences in terms of hard & fast rules that could be true for all people in all situations. But, that doesn’t make the difference any less qualitatively special or real.
   
   Similarly…

2. Self-Help Vs. “Self”–“Help” – Solving the problem that caused the problem that caused the problem that caused the symptom we eventually noticed. Huge. Arguably, peerless.

   • Viz.: How many of us ignore the actual cause of our problem in favor of just reading dozens of blog posts about how to “turbocharge” its most superficial symptoms? Sick.

3. Focus & Play – Yes, focusing on important work is, as Ford used to say, Job 1. But, that focus benefits when we maintain the durable and unapologetic sense of play that affords true creativity and fosters an emergence of context and connection that’s usually killed by stress. BUT.

   • Again, what conceivable “rule” could ever serve to immutably declare that “THIS goofing-off is critical for hippocampal plasticity” vs. “THAT goofing-off is just dumb, distracting bullshit?”

   • Impossible. Because drawing those kinds of distinctions is one of our most important day-to-day responsibilities. Decisions are hard, and there’s no app or alarm gadget that can change that.

     • Although, they certainly can help mask the depth of the underlying problem that made them seem so—what’s the parlance?—“indispensable”.
     • Think: Elmo Band-Aids for that unsightly pancreatic tumor.

4. Reducing Distraction through Care (Rather than braces, armatures, and puppet strings). Removing interruptions and external distractions that harm your work or life? Great. Counting on your distraction-removal tool to supplement your non-existent motivation to do work that will never get done anyway? Pathetic.

   • Frankly, this is a big reason I'm so galled when anyone touts their tool/product/service as being the poor, misunderstood artist’s new miracle medicine—rather than just admitting they've made a slightly different spoon.
   • Because, let’s be honest: although most of us have plenty of perfectly serviceable spoons, everybody knows collecting cutlery is way more fun than using it to swallow yucky medicine.

5. Using a System Vs. Becoming a System. Having a system or process for getting work done vs. making the iteration of that system or process a replacement for the work. This is just…wow…big.
   
   But, maybe most importantly to me…

6. Embracing the Impossibles. Getting past these or any other intellectual koans by simply accepting life’s innumerable and unresolvable paradoxes, hypocrisies, and impossibilities as God-given gifts of creative constraint. Rather than, say, a mimeographed page of long division problems that must be solved for a whole number, n.

   • I just can’t ever get away from this one. For me, it’s what everything inevitably comes back to.
   • The very definition of our jobs is to solve the right problem at the right level for the right reason—based on a combination of the best info we have for now and a clear-eyed dedication to never pushing an unnecessary rock up an avoidable hill.

   • YET, we keep force-feeding the monster that tells us to fiddle and fart and blame the Big Cruel World whenever we face work that might threaten our fragile personal mythology.

     • “Sigh. I wish I could finally start writing My Novel….Ooooooh, if only I had a slightly nicer pen…and Zeus loved me more….”

All that stuff? That there’s a complex set of ideas to talk about for many complex reasons—not least of which being how many people either despise or (try to) deny the unavoidable impact of ol' number six.

But, here’s the thing: as much as saying so pisses anybody off, I think the topics we're NOT talking about whenever we disappear into Talmudic scholarship about “full-screen mode” or “minimalist desks” or whatever constitutes a “zen habit”—those shunned topics are precisely the things that I believe are most mind-blowingly critical to our real-world happiness as humans.

In fact, I believe that to such a degree that helping provide a voice for those unpopular topics that can be heard over the din is now (what passes for) my career. I really believe these deeper ideas are worth socializing on any number of levels and in many media. Even when it’s inconvenient and slightly disrespectful of someone’s business model.

So, that’s what I try to do. I talk about these things. Seldom by careful design. Often poorly. But, always because they each mean an awful lot to me.

[…]

But, no matter how I end up saying whatever the hell I say, I believe in saying it not simply to be liked or followed or revered as a “nice guy” who pushes out shit-tons of whatever to “help people.”

Because, believe me, friend, a great many of those apparently “nice guys” swarming around the web “helping people” these days are ass-fucking their audience for nickels and calling it a complimentary colonoscopy. And, while I absolutely think that in itself is empirically wrong, I also think it’s just as important to say that it’s wrong. Sometimes, True Things need to be said.

Which in this instance amounts to saying, a) selling people a prettier way to kinda almost but not really write is not, in the canonical sense, “nice”—but, far worse, b) leaving your starry-eyed customers with the nauseatingly misguided impression that their “distraction” originates from anyplace but their own busted-ass brain is really not “helping.” Not on any level. It is, literally, harmful.

“Helping” a junkie become more efficient at keeping his syringe loaded is hardly “nice.”

It’s the opposite of nice. And, it’s the opposite of helpful. These are my True Things.

And, to me, saying your True Things also means not watering down the message you care about in order to render it incapable of even conceivably hurting someone’s feelings—or of even conceivably losing you even one teeny-tiny slice of that precious “market share.”

Well, that’s the price, and I'm fine paying it—best money I've ever spent.

But, it also means trusting your audience by letting each of them decide to add water only as they choose to—by never corrupting the actual concentrate in a way that might make it less useful to the smartest or most eager 5% of people who'd like to try using it undiluted. Because, at that point, you're not only abandoning the coolest people you have the honor of serving—you risk becoming a charlatan.

And, that’s precisely what you become when you start to iteratively inbreed the kind of fucktard audience for whom daily buffets of weak swill and beige assurance are life’s most gratifying reward.

Sure. Those poor bastards may never end up using any of that watery information to do anything more ambitious than turbocharging their most regrettable symptoms. But, who’s the last person in the universe who’s going to grab them by the ears and tell them to get back to work? Exactly—that same “nice guy” whose livelihood now depends on keeping infantalized strangers addicted to his “help.”

Holy shit—no way could I ever live with that. It’s so wrong, it’s not even right. ESC, ESC, ESC!

[…]

Okay. So anyhow, there’s a really long-winded, overly generous, and extremely pompous way of trying to say I don’t know how to do what I do except how I do it. But, I do genuinely feel awful when innocent people feel they have been publicly humiliated or berated simply because I'm some dick who hates people.

Which has to be my favorite irony of all.

When I was a kid, I thought my Mom was “mean” not to let me play in traffic on busy Galbraith Road. Today, I'm not simply grateful that she had the strength and resolve to be so “mean”—I actually can’t imagine how sad it would be to not have people in your life who care enough about your long-term welfare to tell you to stop fucking around in traffic. To where you eventually might start even seeking 12x-daily safety hacks from some of the very same drivers whose recklessness may eventually kill you. Wow.

[…]

Admitting when life is complicated or things aren’t shiny and happy all the time strikes me as a wonderfully sane and adult way to conduct one’s life. That there are so many folks offended by even the existence of this anarchic idea is not a problem I can solve.

No more than I can wish useless email away or pray hard enough that it never rains on anyone’s leaky roof. All out of scope.

And, then, I jizzed on at length about how much I admire the recipient’s work. Which I do.

Good work doesn’t need a cookie

I may admire your work, too. Especially if you care a lot about that work and don’t overly sweat peoples' opinions of it. Most definitely including my own.

For these purposes, it doesn’t really matter whether we're friends and, honestly, it doesn’t even matter whether I love, use, or agree with everything you do, say, or make in a given day.

It doesn’t matter because good work doesn’t need me to love it. Like tornadoes and cold sores, good work happens with total disregard to whether I'm “into it.”

But, conversely, let’s stipulate that the points-of-view undergirding our opinions—again, including mine—will and should survive either agreement or lack of agreement with equivalently effortless ease. Because, like really good work, a really good point-of-view doesn’t require another person’s benediction.

Guess we'll have to disagree to agree

Now, to be only vaguely clearer here, I'm not posting this circuitous ego dump in the service of altering your opinion of either me, my friend, his work, or practically anything else for that matter.

But, I would love it if we could all be more okay with the fact that real life means that we do each have a different, sometimes incongruous, and often totally incompatible point-of-view. Yes. Even you have a point-of-view that someone despises. Ready to change it now? Jesus, I sure hope not.

Then, to be only slightly more clear, I'm also not advocating for that fakey brand of web-based kum ba ya that gets trotted out alternately as “tolerance” or “inclusion” or some styrofoam miniature of “civility.”

I'm absolutely not against all of those things when authentically practiced, but I'm also really skeptical of the well-branded peacemakers who are forever appointing themselves the Internet’s “Now-Now-Let’s-All-Pretend-We're-Just-Saying-the-Same-Useless-Thing-Here” den mothers.

Because we're not all saying the same things. Not at all.

And, it infantalizes some important conversations when we tacitly demand that any instance of honest disagreement be immediately horseshat into a photo opp where some thought leader gets to hoist everyone’s hands in the air like he’s fucking Jimmy Carter.

Nope. Not saying that.

Who will you really rely on?

What I AM saying is that alllllll this seemingly unrelated stuff is absolutely related—that the pattern of not relying on other people for anything you really care about is arguably the great-grandaddy of every useful productivity, creativity, or self-help pattern.

Where’s this matter? Pretty much everywhere you have any sort of stake:

• Don’t rely on other people to remove your totally fake “distractions.”
• Don’t rely on other people to pat your beret, re-tie your cravat, and make you a nice cocoa whenever that mean man on the internet points out that your “distractions” are totally fake. (Which they are)
• Don’t rely on other people to tell you when or whether you have enough information.
• Don’t rely on other people to define your job.
• Don’t rely on other people to “design your lifestyle.”
• Don’t rely on other people to decide when your opinions are acceptable.
• Don’t rely on other people to tell you when you're allowed to be awesome.
• Don’t rely on other people to make you care.
• Don’t even rely on other people to tell you what you should or shouldn’t rely on.

Yes. I went there.

Because that’s the point. These hypocrisies, paradoxes, and ambiguities that people get so wound up about—that many of us are constantly (impotently) trying to resolve—cannot be resolved.

Because, yeah: all of these harrowingly unsolvable problems are immune to new notebooks and less-distracting applications and shinier systems and “nicer” self-“help” and pretty much anything else that is not, specifically, you walking straight into the angriest and least convenient shitstorm you can find and getting your ass kicked until the storm gets bored with kicking it.

Then, you find an even angrier storm. Then, another. And, so on.

“Get the fuck off of my obstacle, Private Pyle!”

Doing that annoying hard stuff is how you grow, get better, and learn what real help looks like. Even if that’s not the answer you wanted to hear. You get better by getting your ass out of your RSS reader and fucking making things until they suck less. Not by buying apps.

You don’t whine about distractions, or derail yourself over needing a nicer pencil sharpener, or aggravate your chronic creative diabetes by starting another desperate waddle to the self-help buffet. No. You work.

And, for what it’s worth, just like you can’t get to the moon by eating cheese, you'll also never leave boot camp with your original scrote intact by telling your drill sergeant to try using more honey than vinegar.

No. That sergeant’s job is to make you miserable. It’s his job to break down your callow conceits about what’s supposed to be easy and fair. It’s his job to emotionally pummel you into giving up and becoming a Marine.

You? You're not there to give the sergeant notes; you're there to sleep two hours a night, then not mind getting beaten for 20 hours until a decent Marine starts to fall out.

Who knows? He may even surprise you by introducing a surprisingly effective “distraction-free learning environment.”

“Tee ell dee ahr, Professor Brainiac.”

Like most humans, I like things I can understand. Like most readers, I love specificity. Like most thinkers, I love clarity. Like most students, I love relevance and practicality. And, like most busy people, believe it or not, I actually do really like it when someone gets straight to the point.

But, here’s the problem. If my 2-year-old daughter asks me about time travel, and I blithely announce, “E=mc²”, I will have said something that is entirely specific, clear, relevant, practical, and/or straight-to-the-point. For somebody.

But, not so much for my daughter. And, to be honest, not even to any useful degree for me.

She'd probably either laugh derisively at me (which she’s great at), or she'd pause and ask, “Whuh dat?” (which she’s even better at).

Thing is, her understanding that jumble of characters less than me—and my understanding it WAY less than Professor Al—has zero impact on the profundity, truth, beauty, or impact of the man’s theory.

Sure. You could quite accurately fault me for being a smartass and a poseur, and you could even berate my toddler for her unaccountably shallow understanding of Modern Physics. But, in any case, you can’t really blame either Albert or his theory.

You're turbocharging nothing

Specifically, Albert can’t begin to tell us what he really knows if we don’t understand math.

So, let’s say this theory you've been hearing about really interests you. And, let’s also pretend, just for the sake of the analogy, that you haven’t completed Calculus III (212) or Quantum Mechanics (403) or even something as elementary as, say, Advanced Astrophysics II (537). I know you have. Obviously. But, let’s pretend. Where do you start?

Well, you could read some tips about learning math. You could find a list of 500 indispensable resources for indispensable math resources. You could buy a new “distraction-free math environment.” Heck, there’s actually nothing to stop you from just declaring yourself a “math expert.” Congratulations, Professor.

Thing is: you still don’t know math.

Which means you still can’t really understand the theory—no more than a pathetic Liberal Arts refugee like me or a dullard Physics ignoramus like my kid can really grok relativity.

Difference is, you will have blown a lot of time hoping that actual expertise follows non-existent effort—while my daughter and I get to remain total novices without charge. Only, we don’t get all mad at the theory as a result; a staggering number of fake math experts do.

I mean, be honest—after all that recreational non-work and make-believe dedication almost trying to kinda learn math sorta—you might actually get frustrated at how brazenly Al defies your fondness for shortcuts by continuing to rely on so many terms and proofs and blah-blah-blah that you still just don’t understand. So annoying.

You may simply decide that Albert Einstein’s a huge dick for never saying things that can be completely understood solely by scanning a headline.

EPIC EINSTEIN FAIL, amirite?

You never really know what you didn’t know until you know it

But, Al just told the truth.

Problem is, Al’s truth not only requires fancy things in order to be truly understood—the more of those fancy things you take away from his truth, the less true it gets. And, by the time it’s been diluted to the point where you're comfortable that you understand it? You'd be understanding the wrong thing. Even I can understand that.

But, not one bit of any of this is Al’s fault. Al doesn’t get to control who uses, abuses, gets, or doesn’t get what he said or why it matters. Especially since he’s been dead for over fifty years.

All I know is, regardless of who has ears to hear it on a given day, it would be to Al’s credit never to mangle something important in order to get it into terms everybody’s ready to handle without actually trying.

And God bless him for never agreeing that your “distractions” to learning math are his problem.

So, yeah, if you only need to hand in a crappy 5-page paper, you could certainly Cliff’s Notes your way through Borges, Eliot, or Joyce in an afternoon, and feel like you haven’t missed a thing. Trouble is, if you did care even a little, it’s impossible to even say how much you're missing since you can’t be bothered to soldier through the source text. The text itself is the entire point.

Even the wonderfully cogent and readable layman’s explanations Einstein himself provided don’t really get to the nut, the application, and the implications of his real theory.

That all takes real math.

That “single datum of experience” matters

Sometimes, complex or difficult things stop being true when you try to make them too simple. Sometimes, you have to actually get laid to understand why people think sex is such a thing. Sometimes, you need to learn some Greek if you really want to understand The Gospel of John. And, yeah, sometimes, you're going to have to just work unbelievably hard at whatever you claim to care about before anyone can begin to help you get any better—or less “distracted”—at it.

The part I really know is what doesn’t work. Reading Penthouse Forum won’t help you CLEP out of Vaginal Intercourse 101. Watching a Rankin-Bass cartoon about the Easter Bunny will teach you very little about the intricacies of transubstantiation. And, if you can’t be troubled to care so much about your work that you reflexively force distractions away, dicking around with yet another writing application will merely aggravate the problem. Ironic, huh?

These quantum mechanics of personal productivity are rife with such frustrating “paradoxes.”

These are True Things.

Achieving expertise and doing creative work is all horribly complicated and difficult and paradoxical and frustrating and recursive and James Joyce-y—and any guide, blog, binary, guru, or “nice guy” that tries to suggest otherwise is probably giving you a complimentary colonoscopy. Do the math.

Want a new syllabus? Sure:

Run straight into your shitstorm, my friends. Reject the impulse to think about work, rather than finishing it. And, open your heart to the remote possibility that any mythology of personal failure that involves messiahs periodically arriving to make everything “easy” for you might not really be helping your work or your mental health or your long-standing addiction to using tools solely to ship new excuses.

Learn your real math, and any slide rule will suffice. Try, make, and do until you quit noticing the tools, and if you still think you need new tools, go try, make, and do more.

If you can pull off this deceptively simple and millennia-old pattern, you'll eventually find that—god by dying god—any partial truth that’s supported your treasured excuses for not working will be replaced by a no-faith-required knowledge that you're really, actually, finally getting better at something you care about.

Which is just sublimely un-distracting.

Dedication

This article is dedicated to my friend, Greg Knauss. No, he’s not the app guy–he’s just a good man who does good work, who accidentally/unintentionally helped me write this rant. He also happens to be a fella who could teach anyone a thing or two about writing with distractions. Thanks, Greg.

School bus drivers in the Upper Grand District School Board are transporting school work instead of students.

Grey Bruce provincial police say the OPP canine unit found human remains in Meaford, and one person is under arrest.

After being pulled over for what started as a traffic violation, two Windsor men were arrested and face multiple drug, property, and weapon related charges.

Previous studies have shown that sphingosine kinase interacting protein (SKIP) inhibits sphingosine kinase (SK) function in fibroblasts. SK phosphorylates sphingosine producing the potent signaling molecule sphingosine-1-phosphate (S1P). SKIP gene (SPHKAP) expression is silenced by hypermethylation of its promoter in acute myeloid leukemia (AML). However, why SKIP activity is silenced in primary AML cells is unclear. Here, we investigated the consequences of SKIP down-regulation in AML primary cells and the effects of SKIP re-expression in leukemic cell lines. Using targeted ultra-HPLC-tandem MS (UPLC-MS/MS), we measured sphingolipids (including S1P and ceramides) in AML and control cells. Primary AML cells had significantly lower SK activity and intracellular S1P concentrations than control cells, and SKIP-transfected leukemia cell lines exhibited increased SK activity. These findings show that SKIP re-expression enhances SK activity in leukemia cells. Furthermore, other bioactive sphingolipids such as ceramide were also down-regulated in primary AML cells. Of note, SKIP re-expression in leukemia cells increased ceramide levels 2-fold, inactivated the key signaling protein extracellular signal-regulated kinase, and increased apoptosis following serum deprivation or chemotherapy. These results indicate that SKIP down-regulation in AML reduces SK activity and ceramide levels, an effect that ultimately inhibits apoptosis in leukemia cells. The findings of our study contrast with previous results indicating that SKIP inhibits SK function in fibroblasts and therefore challenge the notion that SKIP always inhibits SK activity.

The transcription factor forkhead box P3 (FOXP3) is a biomarker for regulatory T cells and can also be expressed in cancer cells, but its function in cancer appears to be divergent. The role of hepatocyte-expressed FOXP3 in hepatocellular carcinoma (HCC) is unknown. Here, we collected tumor samples and clinical information from 115 HCC patients and used five human cancer cell lines. We examined FOXP3 mRNA sequences for mutations, used a luciferase assay to assess promoter activities of FOXP3's target genes, and employed mouse tumor models to confirm in vitro results. We detected mutations in the FKH domain of FOXP3 mRNAs in 33% of the HCC tumor tissues, but in none of the adjacent nontumor tissues. None of the mutations occurred at high frequency, indicating that they occurred randomly. Notably, the mutations were not detected in the corresponding regions of FOXP3 genomic DNA, and many of them resulted in amino acid substitutions in the FKH region, altering FOXP3's subcellular localization. FOXP3 delocalization from the nucleus to the cytoplasm caused loss of transcriptional regulation of its target genes, inactivated its tumor-inhibitory capability, and changed cellular responses to histone deacetylase (HDAC) inhibitors. More complex FKH mutations appeared to be associated with worse prognosis in HCC patients. We conclude that mutations in the FKH domain of FOXP3 mRNA frequently occur in HCC and that these mutations are caused by errors in transcription and are not derived from genomic DNA mutations. Our results suggest that transcriptional mutagenesis of FOXP3 plays a role in HCC.

Prostate cancer (PCa) cells heavily rely on an active androgen receptor (AR) pathway for their survival. Enzalutamide (MDV3100) is a second-generation antiandrogenic drug that was approved by the Food and Drug Administration in 2012 to treat patients with castration-resistant prostate cancer (CRPC). However, emergence of resistance against this drug is inevitable, and it has been a major challenge to develop interventions that help manage enzalutamide-resistant CRPC. Erythropoietin-producing human hepatocellular (Eph) receptors are targeted by ephrin protein ligands and have a broad range of functions. Increasing evidence indicates that this signaling pathway plays an important role in tumorigenesis. Overexpression of EPH receptor B4 (EPHB4) has been observed in multiple types of cancer, being closely associated with proliferation, invasion, and metastasis of tumors. Here, using RNA-Seq analyses of clinical and preclinical samples, along with several biochemical and molecular methods, we report that enzalutamide-resistant PCa requires an active EPHB4 pathway that supports drug resistance of this tumor type. Using a small kinase inhibitor and RNAi-based gene silencing to disrupt EPHB4 activity, we found that these disruptions re-sensitize enzalutamide-resistant PCa to the drug both in vitro and in vivo. Mechanistically, we found that EPHB4 stimulates the AR by inducing proto-oncogene c-Myc (c-Myc) expression. Taken together, these results provide critical insight into the mechanism of enzalutamide resistance in PCa, potentially offering a therapeutic avenue for enhancing the efficacy of enzalutamide to better manage this common malignancy.

Heterotrimeric G proteins are the core upstream elements that transduce and amplify the cellular signals from G protein–coupled receptors (GPCRs) to intracellular effectors. GPCRs are the largest family of membrane proteins encoded in the human genome and are the targets of about one-third of prescription medicines. However, to date, no single therapeutic agent exerts its effects via perturbing heterotrimeric G protein function, despite a plethora of evidence linking G protein malfunction to human disease. Several recent studies have brought to light that the Gq family–specific inhibitor FR900359 (FR) is unexpectedly efficacious in silencing the signaling of Gq oncoproteins, mutant Gq variants that mostly exist in the active state. These data not only raise the hope that researchers working in drug discovery may be able to potentially strike Gq oncoproteins from the list of undruggable targets, but also raise questions as to how FR achieves its therapeutic effect. Here, we place emphasis on these recent studies and explain why they expand our pharmacological armamentarium for targeting Gq protein oncogenes as well as broaden our mechanistic understanding of Gq protein oncogene function. We also highlight how this novel insight impacts the significance and utility of using G(q) proteins as targets in drug discovery efforts.

The Mycobacterium tuberculosis virulence factor EsxA and its chaperone EsxB are secreted as a heterodimer (EsxA:B) and are crucial for mycobacterial escape from phagosomes and cytosolic translocation. Current findings support the idea that for EsxA to interact with host membranes, EsxA must dissociate from EsxB at low pH. However, the molecular mechanism by which the EsxA:B heterodimer separates is not clear. In the present study, using liposome-leakage and cytotoxicity assays, LC-MS/MS–based proteomics, and CCF-4 FRET analysis, we obtained evidence that the Nα-acetylation of the Thr-2 residue on EsxA, a post-translational modification that is present in mycobacteria but absent in Escherichia coli, is required for the EsxA:B separation. Substitutions at Thr-2 that precluded Nα-acetylation inhibited the heterodimer separation and hence prevented EsxA from interacting with the host membrane, resulting in attenuated mycobacterial cytosolic translocation and virulence. Molecular dynamics simulations revealed that at low pH, the Nα-acetylated Thr-2 makes direct and frequent “bind-and-release” contacts with EsxB, which generates a force that pulls EsxB away from EsxA. In summary, our findings provide evidence that the Nα-acetylation at Thr-2 of EsxA facilitates dissociation of the EsxA:B heterodimer required for EsxA membrane permeabilization and mycobacterial cytosolic translocation and virulence.

Type 2 diabetes mellitus (T2DM) is characterized by impaired glucose-stimulated insulin secretion and increased peripheral insulin resistance. Unremitting endoplasmic reticulum (ER) stress can lead to beta-cell apoptosis and has been linked to type 2 diabetes. Although many studies have attempted to link ER stress and T2DM, the specific effects of ER stress on beta-cell function remain incompletely understood. To determine the interrelationship between ER stress and beta-cell function, here we treated insulin-secreting INS-1(832/13) cells or isolated mouse islets with the ER stress–inducer tunicamycin (TM). TM induced ER stress as expected, as evidenced by activation of the unfolded protein response. Beta cells treated with TM also exhibited concomitant alterations in their electrical activity and cytosolic free Ca2+ oscillations. As ER stress is known to reduce ER Ca2+ levels, we tested the hypothesis that the observed increase in Ca2+ oscillations occurred because of reduced ER Ca2+ levels and, in turn, increased store-operated Ca2+ entry. TM-induced cytosolic Ca2+ and membrane electrical oscillations were acutely inhibited by YM58483, which blocks store-operated Ca2+ channels. Significantly, TM-treated cells secreted increased insulin under conditions normally associated with only minimal release, e.g. 5 mm glucose, and YM58483 blocked this secretion. Taken together, these results support a critical role for ER Ca2+ depletion–activated Ca2+ current in mediating Ca2+-induced insulin secretion in response to ER stress.

Bacterial type VII secretion systems secrete a wide range of extracellular proteins that play important roles in bacterial viability and in interactions of pathogenic mycobacteria with their hosts. Mycobacterial type VII secretion systems consist of five subtypes, ESX-1–5, and have four substrate classes, namely, Esx, PE, PPE, and Esp proteins. At least some of these substrates are secreted as heterodimers. Each ESX system mediates the secretion of a specific set of Esx, PE, and PPE proteins, raising the question of how these substrates are recognized in a system-specific fashion. For the PE/PPE heterodimers, it has been shown that they interact with their cognate EspG chaperone and that this chaperone determines the designated secretion pathway. However, both structural and pulldown analyses have suggested that EspG cannot interact with the Esx proteins. Therefore, the determining factor for system specificity of the Esx proteins remains unknown. Here, we investigated the secretion specificity of the ESX-1 substrate pair EsxB_1/EsxA_1 in Mycobacterium marinum. Although this substrate pair was hardly secreted when homologously expressed, it was secreted when co-expressed together with the PE35/PPE68_1 pair, indicating that this pair could stimulate secretion of the EsxB_1/EsxA_1 pair. Surprisingly, co-expression of EsxB_1/EsxA_1 with a modified PE35/PPE68_1 version that carried the EspG5 chaperone-binding domain, previously shown to redirect this substrate pair to the ESX-5 system, also resulted in redirection and co-secretion of the Esx pair via ESX-5. Our results suggest a secretion model in which PE35/PPE68_1 determines the system-specific secretion of EsxB_1/EsxA_1.

The rapid emergence and dissemination of methicillin-resistant Staphylococcus aureus (MRSA) strains poses a major threat to public health. MRSA possesses an arsenal of secreted host-damaging virulence factors that mediate pathogenicity and blunt immune defenses. Panton–Valentine leukocidin (PVL) and α-toxin are exotoxins that create lytic pores in the host cell membrane. They are recognized as being important for the development of invasive MRSA infections and are thus potential targets for antivirulence therapies. Here, we report the high-resolution X-ray crystal structures of both PVL and α-toxin in their soluble, monomeric, and oligomeric membrane-inserted pore states in complex with n-tetradecylphosphocholine (C14PC). The structures revealed two evolutionarily conserved phosphatidylcholine-binding mechanisms and their roles in modulating host cell attachment, oligomer assembly, and membrane perforation. Moreover, we demonstrate that the soluble C14PC compound protects primary human immune cells in vitro against cytolysis by PVL and α-toxin and hence may serve as the basis for the development of an antivirulence agent for managing MRSA infections.

Assembled α-synuclein in nerve cells and glial cells is the defining pathological feature of neurodegenerative diseases called synucleinopathies. Seeds of α-synuclein can induce the assembly of monomeric protein. Here, we used sucrose gradient centrifugation and transiently transfected HEK 293T cells to identify the species of α-synuclein from the brains of homozygous, symptomatic mice transgenic for human mutant A53T α-synuclein (line M83) that seed aggregation. The most potent fractions contained Sarkosyl-insoluble assemblies enriched in filaments. We also analyzed six cases of idiopathic Parkinson's disease (PD), one case of familial PD, and six cases of multiple system atrophy (MSA) for their ability to induce α-synuclein aggregation. The MSA samples were more potent than those of idiopathic PD in seeding aggregation. We found that following sucrose gradient centrifugation, the most seed-competent fractions from PD and MSA brains are those that contain Sarkosyl-insoluble α-synuclein. The fractions differed between PD and MSA, consistent with the presence of distinct conformers of assembled α-synuclein in these different samples. We conclude that α-synuclein filaments are the main driving force for amplification and propagation of pathology in synucleinopathies.

Cone photoreceptors in the retina enable vision over a wide range of light intensities. However, the processes enabling cone vision in bright light (i.e. photopic vision) are not adequately understood. Chromophore regeneration of cone photopigments may require the retinal pigment epithelium (RPE) and/or retinal Müller glia. In the RPE, isomerization of all-trans-retinyl esters to 11-cis-retinol is mediated by the retinoid isomerohydrolase Rpe65. A putative alternative retinoid isomerase, dihydroceramide desaturase-1 (DES1), is expressed in RPE and Müller cells. The retinol-isomerase activities of Rpe65 and Des1 are inhibited by emixustat and fenretinide, respectively. Here, we tested the effects of these visual cycle inhibitors on immediate, early, and late phases of cone photopic vision. In zebrafish larvae raised under cyclic light conditions, fenretinide impaired late cone photopic vision, while the emixustat-treated zebrafish unexpectedly had normal vision. In contrast, emixustat-treated larvae raised under extensive dark-adaptation displayed significantly attenuated immediate photopic vision concomitant with significantly reduced 11-cis-retinaldehyde (11cRAL). Following 30 min of light, early photopic vision was recovered, despite 11cRAL levels remaining significantly reduced. Defects in immediate cone photopic vision were rescued in emixustat- or fenretinide-treated larvae following exogenous 9-cis-retinaldehyde supplementation. Genetic knockout of Des1 (degs1) or retinaldehyde-binding protein 1b (rlbp1b) did not eliminate photopic vision in zebrafish. Our findings define molecular and temporal requirements of the nonphotopic or photopic visual cycles for mediating vision in bright light.

The plasmas of diabetic or uremic patients and of those receiving peritoneal dialysis treatment have increased levels of the glucose-derived dicarbonyl metabolites like methylglyoxal (MGO), glyoxal (GO), and 3-deoxyglucosone (3-DG). The elevated dicarbonyl levels can contribute to the development of painful neuropathies. Here, we used stimulated immunoreactive Calcitonin Gene–Related Peptide (iCGRP) release as a measure of nociceptor activation, and we found that each dicarbonyl metabolite induces a concentration-, TRPA1-, and Ca2+-dependent iCGRP release. MGO, GO, and 3-DG were about equally potent in the millimolar range. We hypothesized that another dicarbonyl, 3,4-dideoxyglucosone-3-ene (3,4-DGE), which is present in peritoneal dialysis (PD) solutions after heat sterilization, activates nociceptors. We also showed that at body temperatures 3,4-DGE is formed from 3-DG and that concentrations of 3,4-DGE in the micromolar range effectively induced iCGRP release from isolated murine skin. In a novel preparation of the isolated parietal peritoneum PD fluid or 3,4-DGE alone, at concentrations found in PD solutions, stimulated iCGRP release. We also tested whether inflammatory tissue conditions synergize with dicarbonyls to induce iCGRP release from isolated skin. Application of MGO together with bradykinin or prostaglandin E2 resulted in an overadditive effect on iCGRP release, whereas MGO applied at a pH of 5.2 resulted in reduced release, probably due to an MGO-mediated inhibition of transient receptor potential (TRP) V1 receptors. These results indicate that several reactive dicarbonyls activate nociceptors and potentiate inflammatory mediators. Our findings underline the roles of dicarbonyls and TRPA1 receptors in causing pain during diabetes or renal disease.

Manganese (Mn) is an essential micronutrient required for the normal development of many organs, including the brain. Although its roles as a cofactor in several enzymes and in maintaining optimal physiology are well-known, the overall biological functions of Mn are rather poorly understood. Alterations in body Mn status are associated with altered neuronal physiology and cognition in humans, and either overexposure or (more rarely) insufficiency can cause neurological dysfunction. The resultant balancing act can be viewed as a hormetic U-shaped relationship for biological Mn status and optimal brain health, with changes in the brain leading to physiological effects throughout the body and vice versa. This review discusses Mn homeostasis, biomarkers, molecular mechanisms of cellular transport, and neuropathological changes associated with disruptions of Mn homeostasis, especially in its excess, and identifies gaps in our understanding of the molecular and biochemical mechanisms underlying Mn homeostasis and neurotoxicity.

Treatment of patients with triple-negative breast cancer (TNBC) is limited by a lack of effective molecular therapies targeting this disease. Recent studies have identified metabolic alterations in cancer cells that can be targeted to improve responses to standard-of-care chemotherapy regimens. Using MDA-MB-468 and SUM-159PT TNBC cells, along with LC-MS/MS and HPLC metabolomics profiling, we found here that exposure of TNBC cells to the cytotoxic chemotherapy drugs cisplatin and doxorubicin alter arginine and polyamine metabolites. This alteration was because of a reduction in the levels and activity of a rate-limiting polyamine biosynthetic enzyme, ornithine decarboxylase (ODC). Using gene silencing and inhibitor treatments, we determined that the reduction in ODC was mediated by its negative regulator antizyme, targeting ODC to the proteasome for degradation. Treatment with the ODC inhibitor difluoromethylornithine (DFMO) sensitized TNBC cells to chemotherapy, but this was not observed in receptor-positive breast cancer cells. Moreover, TNBC cell lines had greater sensitivity to single-agent DFMO, and ODC levels were elevated in TNBC patient samples. The alterations in polyamine metabolism in response to chemotherapy, as well as DFMO-induced preferential sensitization of TNBC cells to chemotherapy, reported here suggest that ODC may be a targetable metabolic vulnerability in TNBC.

Research Event

Event participants

Extracytoplasmic sugar decoration of glycopolymer components of the bacterial cell wall contributes to their structural diversity. Typically, the molecular mechanism that underpins such a decoration process involves a three-component glycosylation system (TGS) represented by an undecaprenyl-phosphate (Und-P) sugar-activating glycosyltransferase (Und-P GT), a flippase, and a polytopic glycosyltransferase (PolM GT) dedicated to attaching sugar residues to a specific glycopolymer. Here, using bioinformatic analyses, CRISPR-assisted recombineering, structural analysis of cell wall–associated polysaccharides (CWPS) through MALDI-TOF MS and methylation analysis, we report on three such systems in the bacterium Lactococcus lactis. On the basis of sequence similarities, we first identified three gene pairs, csdAB, csdCD, and csdEF, each encoding an Und-P GT and a PolM GT, as potential TGS component candidates. Our experimental results show that csdAB and csdCD are involved in Glc side-chain addition on the CWPS components rhamnan and polysaccharide pellicle (PSP), respectively, whereas csdEF plays a role in galactosylation of lipoteichoic acid (LTA). We also identified a potential flippase encoded in the L. lactis genome (llnz_02975, cflA) and confirmed that it participates in the glycosylation of the three cell wall glycopolymers rhamnan, PSP, and LTA, thus indicating that its function is shared by the three TGSs. Finally, we observed that glucosylation of both rhamnan and PSP can increase resistance to bacteriophage predation and that LTA galactosylation alters L. lactis resistance to bacteriocin.

Bacterial type VII secretion systems secrete a wide range of extracellular proteins that play important roles in bacterial viability and in interactions of pathogenic mycobacteria with their hosts. Mycobacterial type VII secretion systems consist of five subtypes, ESX-1–5, and have four substrate classes, namely, Esx, PE, PPE, and Esp proteins. At least some of these substrates are secreted as heterodimers. Each ESX system mediates the secretion of a specific set of Esx, PE, and PPE proteins, raising the question of how these substrates are recognized in a system-specific fashion. For the PE/PPE heterodimers, it has been shown that they interact with their cognate EspG chaperone and that this chaperone determines the designated secretion pathway. However, both structural and pulldown analyses have suggested that EspG cannot interact with the Esx proteins. Therefore, the determining factor for system specificity of the Esx proteins remains unknown. Here, we investigated the secretion specificity of the ESX-1 substrate pair EsxB_1/EsxA_1 in Mycobacterium marinum. Although this substrate pair was hardly secreted when homologously expressed, it was secreted when co-expressed together with the PE35/PPE68_1 pair, indicating that this pair could stimulate secretion of the EsxB_1/EsxA_1 pair. Surprisingly, co-expression of EsxB_1/EsxA_1 with a modified PE35/PPE68_1 version that carried the EspG5 chaperone-binding domain, previously shown to redirect this substrate pair to the ESX-5 system, also resulted in redirection and co-secretion of the Esx pair via ESX-5. Our results suggest a secretion model in which PE35/PPE68_1 determines the system-specific secretion of EsxB_1/EsxA_1.

A new book argues that Moscow’s approach to regional and global affairs reflects the tension between two very different worlds—the perceptual and the actual.

Chatham House is pleased to announce that Tim Benton has joined the institute as a Distinguished Visiting Fellow in the Energy, Environment and Resources Department.

Invitation Only Research Event

15 April 2020

Members Event Webinar

Event participants

Professor Johan Giesecke, MD, PhD, Professor Emeritus of Infectious Disease Epidemiology, Karolinska Institute Medical University, Stockholm; State Epidemiologist, Sweden (1995-05)
Professor David Heymann CBE, Distinguished Fellow, Global Health Programme, Chatham House; Executive Director, Communicable Diseases Cluster, World Health Organization (1998-03)
Chair: Emma Ross, Senior Consulting Fellow, Global Health Programme, Chatham House

Alfred C. O. Vertegaal
Dec 1, 2006; 5:2298-2310
Research

Guoan Chen
Apr 1, 2002; 1:304-313
Research

Heterotrimeric G proteins are the core upstream elements that transduce and amplify the cellular signals from G protein–coupled receptors (GPCRs) to intracellular effectors. GPCRs are the largest family of membrane proteins encoded in the human genome and are the targets of about one-third of prescription medicines. However, to date, no single therapeutic agent exerts its effects via perturbing heterotrimeric G protein function, despite a plethora of evidence linking G protein malfunction to human disease. Several recent studies have brought to light that the Gq family–specific inhibitor FR900359 (FR) is unexpectedly efficacious in silencing the signaling of Gq oncoproteins, mutant Gq variants that mostly exist in the active state. These data not only raise the hope that researchers working in drug discovery may be able to potentially strike Gq oncoproteins from the list of undruggable targets, but also raise questions as to how FR achieves its therapeutic effect. Here, we place emphasis on these recent studies and explain why they expand our pharmacological armamentarium for targeting Gq protein oncogenes as well as broaden our mechanistic understanding of Gq protein oncogene function. We also highlight how this novel insight impacts the significance and utility of using G(q) proteins as targets in drug discovery efforts.

7 May 2020 , Volume 96, Number 3

Invitation Only Research Event

Event participants

Paul D’anieri, Professor of Public Policy and Political Science, University of California, Riverside
Vlad Mykhnenko, Associate Professor of Sustainable Urban Development, St Peter’s College, University of Oxford
Chair: Orysia Lutsevych, Research Fellow and Manager, Ukraine Forum, Chatham House

Invitation Only Research Event

Event participants

Members Event Webinar

Event participants

Professor Johan Giesecke, MD, PhD, Professor Emeritus of Infectious Disease Epidemiology, Karolinska Institute Medical University, Stockholm; State Epidemiologist, Sweden (1995-05)
Professor David Heymann CBE, Distinguished Fellow, Global Health Programme, Chatham House; Executive Director, Communicable Diseases Cluster, World Health Organization (1998-03)
Chair: Emma Ross, Senior Consulting Fellow, Global Health Programme, Chatham House

Research Event

Event participants

Members Event

Event participants

Oded Haklai, Associate Professor, Department of Political Studies, Queen's University, Ontario
Neophytos Loizides, Reader, University of Kent; Leverhulme Trust Research Fellow
Madurika Rasaratnam, Lecturer, International Conflict Analysis, University of Kent
Chair: Evangelos Liaras, IE University, Madrid; Academy Associate, Chatham House