Invitation Only Research Event

Event participants

Professor Philippe Sands QC, Barrister, Matrix Chambers
Chris Whomersley, Deputy Legal Adviser, Foreign and Commonwealth Office (2002-14)
Professor Julia Xue, Academy Senior Fellow, International Law Programme, Chatham House
Chair: Elizabeth Wilmshurst, Distinguished Fellow, International Law Programme, Chatham House

21 July 2016

6 January 2017 , Volume 93, Number 1

weeLEGOman posted a photo:

Overwatch

Iker Merodio | Photography posted a photo:

Martinen dinosaurioa.

The investigation into the fatal shooting in Brunswick, Georgia, will also look at a neighbor of suspects Gregory and Travis McMichael who recorded video of the incident, authorities said.

GAZIANTEP, Turkey—After five years fighting to preserve Bashar al-Assad’s regime in Syria, Russia now appears inclined to dispose of its infamous client. Assad’s persistent brutality and corruption, and his inability to establish even the semblance of a functioning state, has grown to be a burden Moscow would prefer not to bear.And then there’s the problem of Iran. Assad, members of his family, and his Alawite clansmen enjoy close, perhaps unbreakable, bonds to the regime in Tehran and to Iranian-backed militias in Syria. All of which undermines Moscow’s primary mission there: to rehabilitate the Assad regime as a symbol of stability capable of attracting hundreds of billions of dollars of foreign investment for reconstruction, which Russian firms would then be poised to receive. As long as Assad’s relatives continue to function as a mafia and give free rein to Iranian troops using Syria as base of operations to threaten Israel and plan attacks against U.S. troops in Iraq, those countries likely to foot the bill for Syrian reconstruction—the nations of Europe and the Gulf—are unlikely to come up with the cash. Amid Escalating Syrian Carnage, Turkey Shoots Down Assad’s PlanesThis has not gone unnoticed by the United States.“Assad has done nothing to help the Russians sell this regime,”James Jeffrey, the U.S. special envoy for the Coalition to Defeat ISIS, told reporters in a State Department briefing on Thursday. “You find Assad has nothing but thugs around him, and they don't sell well either in the Arab world or in Europe. We have heard repeatedly from Russians we take as credible that they understand how bad Assad is.” The Syrian president’s “refusal to make any compromises” in order to secure diplomatic recognition and acceptance for his regime has jeopardized “hundreds of billions of dollars in reconstruction assistance” for Syria, according to Jeffrey. Yet the Trump administration is unlikely to exploit this growing rift. “Getting Russia out of Syria,” Jeffrey said, “has never been our goal. Russia has been there for 30 years. It has a long-term relationship with Syria. We don’t think it has been healthy for the region. We don’t think it really is even healthy for Russia. But that’s not our policy.”  MEDIA FRENZYJeffrey’s statements come just one week after Russian state media unleashed a slew of reports and editorials targeting Assad, portraying the beleaguered president as hopelessly corrupt and unfit to govern, and suggesting the time had come to replace him with a new leader.The first batch of articles was published by the Russia’s Federal News Agency (FNA), an outlet owned by Yevgeny Prigozhin, a Russian oligarch and chairman of several companies implicated in the 2016 U.S. elections scandal. Appearing over the course of a mere three hours on April 17, they would shake Syria to its core. The first of the three articles in question highlighted a corruption scheme carried out by the regime in summer 2019 in which the Syrian prime minister purportedly lied to citizens about oil and gas scarcities in order to justify the occurrence of long power outages while selling Syrian electricity to businessmen in Lebanon. The second piece cited an opinion poll claiming only 32 percent of Syrians would vote for Assad in the country’s upcoming 2021 presidential election. The third and final article, entitled, “Corruption is Worse than Terrorism,” chastized President Assad for personally failing to combat corruption, prevalent at all levels of the state.  That these were published by Prigozhin’s news agency was the kind of signal it would be hard for Assad to miss. Prigozhin, who first built his fortune as a caterer, is sometimes known as “Putin’s chef.” But of particular relevance to Syria is his role as chairman of the Wagner Group, whose mercenaries have fought alongside Assad regime forces since October 2015 and helped the latter take back control of key revenue generating infrastructure such as the al-Sha’ir gas field in Homs province.Deputy Assistant Secretary Christopher Robin told the same State Department briefing Thursday, “Wagner is often misleadingly referred to as a Russian private military company, but in fact it’s an instrument of the Russian government which the Kremlin uses as a low-cost and low-risk instrument to advance its goals.”The article on corruption would also point out, suggestively, that the Assads are not the only powerful family in Syria, “there are also the Makhloufs.”Rami Makhlouf, who is in fact Bashar al-Assad’s first cousin, is Syria’s wealthiest man, and also, it would seem, Russia’s man. Certainly he has strong ties to the Kremlin and for years has been one of the most vocal critics of Iran’s presence in Syria. In July 2018, the al-Watan newspaper, one Syria’s most prominent pro-regime mouthpieces and owned by Rami Makhlouf since 2006, published a then unprecedented public rebuke to Iran, accusing it of sponsoring Islamist fanaticism throughout the Middle East alongside Turkey and Qatar, the main backers of Syria’s opposition. (Rami Makhlouf’s father Muhammad and brother Hafiz meanwhile are alleged by some to be living in Russia.) The April 17 articles published by Prigozhin’s FNA preceded the release of a wave of other articles and items in the media over the next 12 days that would further drive home the point that Moscow was considering options other than Assad to rule Syria. TASS, Russia’s largest state-run news agency, wrote in one editorial that, “Russia suspects that Assad is not only unable to lead the country anymore, but also that the head of the Syrian regime is dragging Moscow towards the Afghani scenario.” This is like evoking the Vietnam War for an American audience, a reference to the Kremlin’s botched campaign through the 1980s that helped bankrupt the Soviet Union and finally break it apart.Amid this coverage, TASS would also take swipes at Iran, claiming that the Islamic Republic has “no interest in achieving stability in the region, because it considers it a battlefield with Washington”.On April 30, the Russian International Affairs Council (RIAC), a think tank established by Moscow’s Ministry of Foreign Affairs, released a scathing report saying Russia was in talks with other parties to the Syrian conflict to draw up plans for a political resolution that did not include Bashar al-Assad as president. The report highlighted purported Russian efforts to compel the Syrian regime to commit to ceasefires with both American-backed and Kurdish-led Syrian Democratic Forces (SDF), and the Turkish-backed Free Syrian Army (FSA) opposition, while beginning steps to form a new unity government that would include representatives from both. That day, Rami Makhlouf, whose assets were frozen five months earlier as part of a tax dispute, uploaded a video onto his personal Facebook page accusing the Assad regime of corruption. In a state known for carrying out the full-scale slaughter of those who test its authority, Makhlouf’s videos, coming on the heels of the unprecedented Russian attacks in the media, sent shockwaves throughout the country.  THE ROYAL FAMILYWhile the Makhlouf clan clearly has thrown its lot in with Russia, key members of Bashar al-Assad’s immediate family and others with ties to Qardaha in Syria’s largely Alawite Latakia province, are among the most prominent Iranian-backed militia leaders in Syria. It’s an alliance that traces back to his father Hafez al-Assad, who was born in Qardaha, and who forged ties with the Iranian revolution almost from its beginning more than 40 years ago. The Iranians responded by offering religious legitimacy to the Alawite sect, which is regarded as heretical by Sunnis and indeed by many Shi’a.These Qardaha militia leaders have regularly engaged in armed clashes against Russian backed units. They are among the most egregious violators and abusers of power, overseeing wide networks of corruption similar to those lamented in the Russian media. And foremost among them is Bashar’s younger brother, Maher al-Assad. Since April 2018, Maher al-Assad has commanded the Syrian Army’s 4th Armored Division, one of country’s oldest, best equipped and overwhelmingly Alawite brigades. After the 2011 outbreak of the Syrian revolution, when the loyalty of much of the army was in doubt, it became a refuge for numerous Alawite-Shi’a dominated pro-regime militias.Currently, the 4th Armored Division’s members control many smuggling operations throughout the country, in cities from Albu Kamel on Syria’s eastern border with Iraq to Latakia on the Syrian coast, where the port was leased to Iran on October 1 last year. It has since become one of the biggest export hubs for drugs headed to markets in Europe, the Middle East and North Africa. Examples abound: On July 5, 2019, Greek coast guard and drug enforcement officials announced the biggest drug bust in history, seizing 5.25 tons (33 million pills) of Captagon amphetamines worth $660m hidden in shipping containers loaded at the Latakia port in Syria. That followed a long string of such seizures made by Greek authorities. More recently, in late April, customs officials in both Saudi Arabia and Egypt also announced the seizure of similar quantities of drugs in containers traced back to Latakia. Local reports have accused a range of actors including Maher al-Assad’s 4th Division, Hizbollah, Rami Makhlouf, and others of profiting from the massive drug exports emanating from the port. In January 2019 the 4th Armored Division launched attacks on the Russian-backed Tiger Forces unit in an attempt to wrest control of smuggling routes between regime- and opposition-held territory in Idlib province. The clashes led to the death of 70 fighters. These and other skirmishes prompted Russia to back a major campaign to arrest 4th Division and other Iranian-backed units throughout the country beginning in April 2019, which succeeded in rounding up numerous mid-ranking Iranian-backed officers. Among those targeted in the campaign was Bashar Talal al-Assad, a cousin to the president (similar name, different people) who was wanted on drug and weapons trafficking charges. Unlike others who were detained in the roundup, Bashar Talal al-Assad and his ‘Areen Brigade managed to fight off Russian-backed forces that sought to arrest him in Qardaha. He then pledged to attack Russia’s Hmeimim military base, located 17 miles east of Latakia city, in the event the regime sought to arrest him again.For Russia, the threat of such attacks on its military infrastructure is a real concern. The Hmeimim base—from which Moscow has directed its entire military campaign in Syria—had already been subject to a series of attacks from January to October 2018 by other Iranian-backed militias in the area. The threat posed by both Iran’s acquisition of the Latakia port and its support for local Assad family proxies in Syria’s coastal region is exacerbated by the fact that Tehran has also begun making progress toward completing construction of its Shalamcha railroad, which, via stops in Basra, Baghdad, Albu Kamel and Damascus, will give Tehran direct access to the Syrian and Lebanese coasts. If Iran succeeds in integrating the Latakia port with the Shalamcha rail line, this will cut off Hmeimim from Russian forces in central and southern Syria and enable Tehran to quickly deliver weapons to proxy forces in Latakia that are already engaged in clashes against Russian-backed groups. WORLDWIDE CONSENSUSMoscow’s inability to control Iranian backed Syrian militiamen engaged in widespread crime, corruption, and assaults on Russian forces has infuriated the Kremlin. But Russia is not the only major player on the ground with scores to settle against Iran, and the Russian military leadership in Syria has ignored if not largely encouraged Israeli strikes on Iranian troops throughout the country.It may not be coincidental that the Israeli attacks have increased in pace and scope since April, following the flurry of Russian media articles attacking Assad and his regime. “We have moved from blocking Iran’s entrenchment in Syria to forcing it out of there, and we will not stop,” Israel’s new defense minister, Naftali Bennett, declared on April 28. Without Russia, Iran has found itself the odd man out in Syria, the single party still seeking to push for war at a time when most other international players have been struck with fatigue and simply seek to put Syria’s pieces back together. President Recep Tayyip Erdoğan of Turkey, the last patron of Syria’s battered FSA opposition, has himself made peace with Moscow, effectively agreeing last March to cede control of wide swaths of rebel held territory after a particularly bloody Russian led campaign against the last FSA holdout in Idlib province that ended in victory for regime forces. Ironically, Erdoğan’s long-held desire to overthrow Syria’s president may still come to fruition, albeit not as he expected, as Assad’s ouster may come at the hands of Russia itself, and not the revolution. Read more at The Daily Beast.Get our top stories in your inbox every day. Sign up now!Daily Beast Membership: Beast Inside goes deeper on the stories that matter to you. Learn more.

Whenever Americans see videos of police brutality against black men and women, the first thing they do is assume they deserved their executionWhat skin color are the bad guys in America’s fantasies of vigilantism? When the proverbial “fellas” get together to drink beers and talk about their newest guns and who they’d take down, what race are the “criminals” in the theater of their minds?When Greg McMichael and his son, Travis, got the call from their neighbor that a “burglar” was running through their Brunswick, Georgia neighborhood that chilly February day, what color man do you think they imagined as they locked, loaded, and embarked on their “mission”?Ahmaud Arbery is dead today because when Americans dream of vigilante justice, black men are the villains of their imaginations.We as a nation are so comfortable with this baseline bigotry that our first assumption whenever we see videos of police brutality against or shootings of black men and women, the first thing we do is assume that the victims must have done something wrong to earn their own public execution.This assumption is both a function of white America having a completely different experience with police officers than black America as well as the hundreds of years of vilifying blackness in media and American culture.I will never forget the biggest and most uproarious applause during the theater debut of the lackluster 2007 vigilante film, Brave One, came when the protagonist Jodi Foster got her first vigilante kills of the movie – two threatening and scary black men. That theater filled with men the same age range as Greg and Travis McMichael erupted as if at that moment, all that they had ever imagined had been fulfilled on the big screen. Needless to say, I left that theater before the credits rolled.Across the country, our political leaders hold these same bigoted beliefs which inevitably lead to policies that directly assume criminality based on skin color.During his tenure as mayor of New York City, billionaire Michael Bloomberg made it explicitly clear why it was that he sent police officers into black and brown communities to “throw them” up against the wall. In his 2015 Aspen Institute speech he stated:“People say, ‘Oh my God, you are arresting kids for marijuana who are all minorities.’ Yes, that’s true. Why? Because we put all the cops in the minority neighborhoods. Yes, that’s true. Why’d we do it? Because that’s where all the crime is. And the way you should get the guns out of the kids’ hands is throw them against the wall and frisk them.”And it is for this reason that I do not distinguish between the violence committed by American citizens acting as vigilantes and the violence committed by so-called officers of the law when, in both cases, the working assumption and driving force behind that violence is the deeply bigoted and firmly American association between blackness and criminality.For Ahmaud, that association not only led to his brutal killing, but it also initially meant his killer not being arrested. It took more than two months for the father and son duo to be arrested. When explaining why they were not charged immediately the district attorney, George Barnhill, immediately stated that the victim, Ahmaud Arbery, was, in fact, the “criminal suspect”.“It appears that [Greg and Travis McMichael’s] intent was to stop and hold this criminal suspect until law enforcement arrived. Under Georgia Law [sic] this is perfectly legal.”Even after viewing the video and with no evidence beyond Ahmaud’s skin color, the top cop in the institution designed to bring equal justice under the law concluded that Ahmaud was a criminal suspect when he was simply a black man taking a jog.What are black Americans to do when justice is delayed or outright denied because of the assignment of innocence to vigilantes and police officers?What are black Americans to do when the assumption of guilt because of our skin color is as American as the guns they use to kill us?What are we to do when in our neighbors’ dreams and fantasies of cop-and-robber, the skin color of the bad guy matches our own?The very first thing we are going to do is defend ourselves as if our lives depend on it because when Americans fantasize about killing, those fantasies become our living nightmares. * Benjamin Dixon is the host of the Benjamin Dixon show.

Pakistanis crowded markets on Saturday after a nationwide coronavirus lockdown was eased, despite the country declaring its second highest daily infection toll. Prime Minister Imran Khan has allowed businesses to reopen in phases from the weekend, citing the economic havoc the virus restrictions have wreaked on the improvised nation. In the garrison city of Rawalpindi, thousands of shoppers were preparing for Eid, which follows the Muslim fasting month of Ramadan, with many flouting social distancing rules and advice to wear masks.

President Trump said Friday he had watched the video of Ahmaud Arbery being shot and found it “very disturbing” and “heartbreaking,” but said he was confident that the Georgia legal system would come down on the side of justice.

Travel blogger and photographer Andy Luten drove 4,200 miles across six states to see the grounded jets, detailing the shocking state of aviation.

Attorney General Bill Barr explained that the FBI did not conduct “a bonafide counterintelligence investigation” in the case that led former Trump national security adviser Michael Flynn to plead guilty to federal investigators in 2017.Barr, speaking in an exclusive interview with CBS News after the Justice Department dropped its case against Flynn on Thursday, said that his review of the case found Bureau investigators laid “a perjury trap” for Flynn in a January 2017 White House interview.“They didn’t warn him, the way that would usually be required by the Department, they bypassed the Justice Department, they bypassed the protocols at the White House, and so forth,” Barr stated. “These were things that persuaded me that there was not a legitimate counterintelligence investigation.”Former FBI director James Comey admitted in a December 2018 interview that he “sent” the agents to interview Flynn, adding that it was “something I probably wouldn’t have done or maybe gotten away with in a more organized administration.”In its Thursday court filing, the Justice Department explained that it was “not persuaded” that Flynn’s interview, which led to his guilty plea for lying to FBI agents Peter Strzok and Joe Pientka, had proper predication and was materially relevant.Comey tweeted his disappointment, following the decision, saying "the DOJ has lost its way."> The DOJ has lost its way. But, career people: please stay because America needs you. The country is hungry for honest, competent leadership.> > -- James Comey (@Comey) May 7, 2020Barr pointed to recently-released information that showed the FBI moved to close its surveillance of Flynn after finding “no derogatory information” about the retired general’s contacts with Russians, only for Strzok to keep the case open, leading to the eventual interview.“They were closing the investigation, in December [2016], they started that process and on January 4, they were closing it. When they heard about the phone call, which the FBI had the transcripts to — there was no question as to what was discussed, the FBI knew exactly what was discussed — and General Flynn, being the former director of the DIA, said to them, ‘you listen to everything, you know what was said,’” Barr explained.“So there’s no mystery about the call, but they initially tried some theories of how could open another investigation, which didn’t fly, and then they found out that they had not technically closed the earlier investigation, and they kept it open for the expressed purpose of trying to catch — lay a perjury trap — for General Flynn,” he added. A different filing released last week showed handwritten notes from an FBI official that questioned if the goal of Flynn’s White House interview was “to get him to lie, so we can prosecute him or get him fired.”Barr also did not comment on whether those that sought to entrap Flynn would face criminal charges, pointing to U.S. Attorney John Durham’s probe into the origins of the Trump-Russia investigation and saying his team was “in the middle” of “looking at the whole pattern of conduct.”“I’m going to wait until all the evidence is [in], and I get their recommendations as to what they found and how serious it is. But, if we were to find wrongdoing, in the sense of any criminal act, obviously we would follow through on that,” Barr said. “But again, just because something may even stink to high heaven, and appear to everyone to be bad, we still have to apply the right standard and be convinced that there is a violation of a criminal statute and that we can prove it beyond a reasonable doubt. The same standard applies to everybody.”

In a tweet Saturday morning, Tesla's chief executive said it would file a lawsuit against county officials over not being able to run its factory.

The C-type lectin receptors (CLRs) form a family of pattern recognition receptors that recognize numerous pathogens, such as bacteria and fungi, and trigger innate immune responses. The extracellular carbohydrate-recognition domain (CRD) of CLRs forms a globular structure that can coordinate a Ca2+ ion, allowing receptor interactions with sugar-containing ligands. Although well-conserved, the CRD fold can also display differences that directly affect the specificity of the receptors for their ligands. Here, we report crystal structures at 1.8–2.3 Å resolutions of the CRD of murine dendritic cell-immunoactivating receptor (DCAR, or Clec4b1), the CLR that binds phosphoglycolipids such as acylated phosphatidyl-myo-inositol mannosides (AcPIMs) of mycobacteria. Using mutagenesis analysis, we identified critical residues, Ala136 and Gln198, on the surface surrounding the ligand-binding site of DCAR, as well as an atypical Ca2+-binding motif (Glu-Pro-Ser/EPS168–170). By chemically synthesizing a water-soluble ligand analog, inositol-monophosphate dimannose (IPM2), we confirmed the direct interaction of DCAR with the polar moiety of AcPIMs by biolayer interferometry and co-crystallization approaches. We also observed a hydrophobic groove extending from the ligand-binding site that is in a suitable position to interact with the lipid portion of whole AcPIMs. These results suggest that the hydroxyl group-binding ability and hydrophobic groove of DCAR mediate its specific binding to pathogen-derived phosphoglycolipids such as mycobacterial AcPIMs.

Although a robust inflammatory response is needed to combat infection, this response must ultimately be terminated to prevent chronic inflammation. One mechanism that terminates inflammatory signaling is the production of alternative mRNA splice forms in the Toll-like receptor (TLR) signaling pathway. Whereas most genes in the TLR pathway encode positive mediators of inflammatory signaling, several, including that encoding the MyD88 signaling adaptor, also produce alternative spliced mRNA isoforms that encode dominant-negative inhibitors of the response. Production of these negatively acting alternatively spliced isoforms is induced by stimulation with the TLR4 agonist lipopolysaccharide (LPS); thus, this alternative pre-mRNA splicing represents a negative feedback loop that terminates TLR signaling and prevents chronic inflammation. In the current study, we investigated the mechanisms regulating the LPS-induced alternative pre-mRNA splicing of the MyD88 transcript in murine macrophages. We found that 1) the induction of the alternatively spliced MyD88 form is due to alternative pre-mRNA splicing and not caused by another RNA regulatory mechanism, 2) MyD88 splicing is regulated by both the MyD88- and TRIF-dependent arms of the TLR signaling pathway, 3) MyD88 splicing is regulated by the NF-κB transcription factor, and 4) NF-κB likely regulates MyD88 alternative pre-mRNA splicing per se rather than regulating splicing indirectly by altering MyD88 transcription. We conclude that alternative splicing of MyD88 may provide a sensitive mechanism that ensures robust termination of inflammation for tissue repair and restoration of normal tissue homeostasis once an infection is controlled.

Protease-activated receptor 2 (PAR-2) is activated by secreted proteases from immune cells or fungi. PAR-2 is normally expressed basolaterally in differentiated nasal ciliated cells. We hypothesized that epithelial remodeling during diseases characterized by cilial loss and squamous metaplasia may alter PAR-2 polarization. Here, using a fluorescent arrestin assay, we confirmed that the common fungal airway pathogen Aspergillus fumigatus activates heterologously-expressed PAR-2. Endogenous PAR-2 activation in submerged airway RPMI 2650 or NCI–H520 squamous cells increased intracellular calcium levels and granulocyte macrophage–colony-stimulating factor, tumor necrosis factor α, and interleukin (IL)-6 secretion. RPMI 2650 cells cultured at an air–liquid interface (ALI) responded to apically or basolaterally applied PAR-2 agonists. However, well-differentiated primary nasal epithelial ALIs responded only to basolateral PAR-2 stimulation, indicated by calcium elevation, increased cilia beat frequency, and increased fluid and cytokine secretion. We exposed primary cells to disease-related modifiers that alter epithelial morphology, including IL-13, cigarette smoke condensate, and retinoic acid deficiency, at concentrations and times that altered epithelial morphology without causing breakdown of the epithelial barrier to model early disease states. These altered primary cultures responded to both apical and basolateral PAR-2 stimulation. Imaging nasal polyps and control middle turbinate explants, we found that nasal polyps, but not turbinates, exhibit apical calcium responses to PAR-2 stimulation. However, isolated ciliated cells from both polyps and turbinates maintained basolateral PAR-2 polarization, suggesting that the calcium responses originated from nonciliated cells. Altered PAR-2 polarization in disease-remodeled epithelia may enhance apical responses and increase sensitivity to inhaled proteases.

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.

Prevention of aberrant cutaneous wound repair and appropriate regeneration of an intact and functional integument require the coordinated timing of fibroblast and keratinocyte migration. Here, we identified a mechanism whereby opposing cell-specific motogenic functions of a multifunctional intracellular and extracellular protein, the receptor for hyaluronan-mediated motility (RHAMM), coordinates fibroblast and keratinocyte migration speed and ensures appropriate timing of excisional wound closure. We found that, unlike in WT mice, in Rhamm-null mice, keratinocyte migration initiates prematurely in the excisional wounds, resulting in wounds that have re-surfaced before the formation of normal granulation tissue, leading to a defective epidermal architecture. We also noted aberrant keratinocyte and fibroblast migration in the Rhamm-null mice, indicating that RHAMM suppresses keratinocyte motility but increases fibroblast motility. This cell context–dependent effect resulted from cell-specific regulation of extracellular signal-regulated kinase 1/2 (ERK1/2) activation and expression of a RHAMM target gene encoding matrix metalloprotease 9 (MMP-9). In fibroblasts, RHAMM promoted ERK1/2 activation and MMP-9 expression, whereas in keratinocytes, RHAMM suppressed these activities. In keratinocytes, loss of RHAMM function or expression promoted epidermal growth factor receptor–regulated MMP-9 expression via ERK1/2, which resulted in cleavage of the ectodomain of the RHAMM partner protein CD44 and thereby increased keratinocyte motility. These results identify RHAMM as a key factor that integrates the timing of wound repair by controlling cell migration.

Inter-α-inhibitor is a proteoglycan essential for mammalian reproduction and also plays a less well-characterized role in inflammation. It comprises two homologous “heavy chains” (HC1 and HC2) covalently attached to chondroitin sulfate on the bikunin core protein. Before ovulation, HCs are transferred onto the polysaccharide hyaluronan (HA) to form covalent HC·HA complexes, thereby stabilizing an extracellular matrix around the oocyte required for fertilization. Additionally, such complexes form during inflammatory processes and mediate leukocyte adhesion in the synovial fluids of arthritis patients and protect against sepsis. Here using X-ray crystallography, we show that human HC1 has a structure similar to integrin β-chains, with a von Willebrand factor A domain containing a functional metal ion-dependent adhesion site (MIDAS) and an associated hybrid domain. A comparison of the WT protein and a variant with an impaired MIDAS (but otherwise structurally identical) by small-angle X-ray scattering and analytical ultracentrifugation revealed that HC1 self-associates in a cation-dependent manner, providing a mechanism for HC·HA cross-linking and matrix stabilization. Surprisingly, unlike integrins, HC1 interacted with RGD-containing ligands, such as fibronectin, vitronectin, and the latency-associated peptides of transforming growth factor β, in a MIDAS/cation-independent manner. However, HC1 utilizes its MIDAS motif to bind to and inhibit the cleavage of complement C3, and small-angle X-ray scattering–based modeling indicates that this occurs through the inhibition of the alternative pathway C3 convertase. These findings provide detailed structural and functional insights into HC1 as a regulator of innate immunity and further elucidate the role of HC·HA complexes in inflammation and ovulation.

β-Glucocerebrosidase (GBA) hydrolyzes glucosylceramide (GlcCer) to generate ceramide. Previously, we demonstrated that lysosomal GBA1 and nonlysosomal GBA2 possess not only GlcCer hydrolase activity, but also transglucosylation activity to transfer the glucose residue from GlcCer to cholesterol to form β-cholesterylglucoside (β-GlcChol) in vitro. β-GlcChol is a member of sterylglycosides present in diverse species. How GBA1 and GBA2 mediate β-GlcChol metabolism in the brain is unknown. Here, we purified and characterized sterylglycosides from rodent and fish brains. Although glucose is thought to be the sole carbohydrate component of sterylglycosides in vertebrates, structural analysis of rat brain sterylglycosides revealed the presence of galactosylated cholesterol (β-GalChol), in addition to β-GlcChol. Analyses of brain tissues from GBA2-deficient mice and GBA1- and/or GBA2-deficient Japanese rice fish (Oryzias latipes) revealed that GBA1 and GBA2 are responsible for β-GlcChol degradation and formation, respectively, and that both GBA1 and GBA2 are responsible for β-GalChol formation. Liquid chromatography–tandem MS revealed that β-GlcChol and β-GalChol are present throughout development from embryo to adult in the mouse brain. We found that β-GalChol expression depends on galactosylceramide (GalCer), and developmental onset of β-GalChol biosynthesis appeared to be during myelination. We also found that β-GlcChol and β-GalChol are secreted from neurons and glial cells in association with exosomes. In vitro enzyme assays confirmed that GBA1 and GBA2 have transgalactosylation activity to transfer the galactose residue from GalCer to cholesterol to form β-GalChol. This is the first report of the existence of β-GalChol in vertebrates and how β-GlcChol and β-GalChol are formed in the brain.

Actinobacillus pleuropneumoniae (App) is the etiological agent of acute porcine pneumonia and responsible for severe economic losses worldwide. The capsule polymer of App serotype 1 (App1) consists of [4)-GlcNAc-β(1,6)-Gal-α-1-(PO4-] repeating units that are O-acetylated at O-6 of the GlcNAc. It is a major virulence factor and was used in previous studies in the successful generation of an experimental glycoconjugate vaccine. However, the application of glycoconjugate vaccines in the animal health sector is limited, presumably because of the high costs associated with harvesting the polymer from pathogen culture. Consequently, here we exploited the capsule polymerase Cps1B of App1 as an in vitro synthesis tool and an alternative for capsule polymer provision. Cps1B consists of two catalytic domains, as well as a domain rich in tetratricopeptide repeats (TPRs). We compared the elongation mechanism of Cps1B with that of a ΔTPR truncation (Cps1B-ΔTPR). Interestingly, the product profiles displayed by Cps1B suggested processive elongation of the nascent polymer, whereas Cps1B-ΔTPR appeared to work in a more distributive manner. The dispersity of the synthesized products could be reduced by generating single-action transferases and immobilizing them on individual columns, separating the two catalytic activities. Furthermore, we identified the O-acetyltransferase Cps1D of App1 and used it to modify the polymers produced by Cps1B. Two-dimensional NMR analyses of the products revealed O-acetylation levels identical to those of polymer harvested from App1 culture supernatants. In conclusion, we have established a protocol for the pathogen-free in vitro synthesis of tailored, nature-identical App1 capsule polymers.

Lens proteins become increasingly cross-linked through nondisulfide linkages during aging and cataract formation. One mechanism that has been implicated in this cross-linking is glycation through formation of advanced glycation end products (AGEs). Here, we found an age-associated increase in stiffness in human lenses that was directly correlated with levels of protein–cross-linking AGEs. α-Crystallin in the lens binds to other proteins and prevents their denaturation and aggregation through its chaperone-like activity. Using a FRET-based assay, we examined the stability of the αA-crystallin–γD-crystallin complex for up to 12 days and observed that this complex is stable in PBS and upon incubation with human lens–epithelial cell lysate or lens homogenate. Addition of 2 mm ATP to the lysate or homogenate did not decrease the stability of the complex. We also generated complexes of human αA-crystallin or αB-crystallin with alcohol dehydrogenase or citrate synthase by applying thermal stress. Upon glycation under physiological conditions, the chaperone–client complexes underwent greater extents of cross-linking than did uncomplexed protein mixtures. LC-MS/MS analyses revealed that the levels of cross-linking AGEs were significantly higher in the glycated chaperone–client complexes than in glycated but uncomplexed protein mixtures. Mouse lenses subjected to thermal stress followed by glycation lost resilience more extensively than lenses subjected to thermal stress or glycation alone, and this loss was accompanied by higher protein cross-linking and higher cross-linking AGE levels. These results uncover a protein cross-linking mechanism in the lens and suggest that AGE-mediated cross-linking of α-crystallin–client complexes could contribute to lens aging and presbyopia.

The dextransucrase DSR-OK from the Gram-positive bacterium Oenococcus kitaharae DSM17330 produces a dextran of the highest molar mass reported to date (∼109 g/mol). In this study, we selected a recombinant form, DSR-OKΔ1, to identify molecular determinants involved in the sugar polymerization mechanism and that confer its ability to produce a very-high-molar-mass polymer. In domain V of DSR-OK, we identified seven putative sugar-binding pockets characteristic of glycoside hydrolase 70 (GH70) glucansucrases that are known to be involved in glucan binding. We investigated their role in polymer synthesis through several approaches, including monitoring of dextran synthesis, affinity assays, sugar binding pocket deletions, site-directed mutagenesis, and construction of chimeric enzymes. Substitution of only two stacking aromatic residues in two consecutive sugar-binding pockets (variant DSR-OKΔ1-Y1162A-F1228A) induced quasi-complete loss of very-high-molar-mass dextran synthesis, resulting in production of only 10–13 kg/mol polymers. Moreover, the double mutation completely switched the semiprocessive mode of DSR-OKΔ1 toward a distributive one, highlighting the strong influence of these pockets on enzyme processivity. Finally, the position of each pocket relative to the active site also appeared to be important for polymer elongation. We propose that sugar-binding pockets spatially closer to the catalytic domain play a major role in the control of processivity. A deep structural characterization, if possible with large-molar-mass sugar ligands, would allow confirming this hypothesis.

Bacterial biofilms are cellular communities that produce an adherent matrix. Exopolysaccharides are key structural components of this matrix and are required for the assembly and architecture of biofilms produced by a wide variety of microorganisms. The human bacterial pathogens Escherichia coli and Salmonella enterica produce a biofilm matrix composed primarily of the exopolysaccharide phosphoethanolamine (pEtN) cellulose. Once thought to be composed of only underivatized cellulose, the pEtN modification present in these matrices has been implicated in the overall architecture and integrity of the biofilm. However, an understanding of the mechanism underlying pEtN derivatization of the cellulose exopolysaccharide remains elusive. The bacterial cellulose synthase subunit G (BcsG) is a predicted inner membrane–localized metalloenzyme that has been proposed to catalyze the transfer of the pEtN group from membrane phospholipids to cellulose. Here we present evidence that the C-terminal domain of BcsG from E. coli (EcBcsGΔN) functions as a phosphoethanolamine transferase in vitro with substrate preference for cellulosic materials. Structural characterization of EcBcsGΔN revealed that it belongs to the alkaline phosphatase superfamily, contains a Zn2+ ion at its active center, and is structurally similar to characterized enzymes that confer colistin resistance in Gram-negative bacteria. Informed by our structural studies, we present a functional complementation experiment in E. coli AR3110, indicating that the activity of the BcsG C-terminal domain is essential for integrity of the pellicular biofilm. Furthermore, our results established a similar but distinct active-site architecture and catalytic mechanism shared between BcsG and the colistin resistance enzymes.

Extracellular matrix-evoked angiostasis and autophagy within the tumor microenvironment represent two critical, but unconnected, functions of the small leucine-rich proteoglycan, decorin. Acting as a partial agonist of vascular endothelial growth factor 2 (VEGFR2), soluble decorin signals via the energy sensing protein, AMP-activated protein kinase (AMPK), in the autophagic degradation of intracellular vascular endothelial growth factor A (VEGFA). Here, we discovered that soluble decorin evokes intracellular catabolism of endothelial VEGFA that is mechanistically independent of mTOR, but requires an autophagic regulator, paternally expressed gene 3 (PEG3). We found that administration of autophagic inhibitors such as chloroquine or bafilomycin A1, or depletion of autophagy-related 5 (ATG5), results in accumulation of intracellular VEGFA, indicating that VEGFA is a basal autophagic substrate. Mechanistically, decorin increased the VEGFA clearance rate by augmenting autophagic flux, a process that required RAB24 member RAS oncogene family (RAB24), a small GTPase that facilitates the disposal of autophagic compartments. We validated these findings by demonstrating the physiological relevance of this process in vivo. Mice starved for 48 h exhibited a sharp decrease in overall cardiac and aortic VEGFA that could be blocked by systemic chloroquine treatment. Thus, our findings reveal a unified mechanism for the metabolic control of endothelial VEGFA for autophagic clearance in response to decorin and canonical pro-autophagic stimuli. We posit that the VEGFR2/AMPK/PEG3 axis integrates the anti-angiogenic and pro-autophagic bioactivities of decorin as the molecular basis for tumorigenic suppression. These results support future therapeutic use of decorin as a next-generation protein therapy to combat cancer.

Endorepellin, the C-terminal fragment of the heparan sulfate proteoglycan perlecan, influences various signaling pathways in endothelial cells by binding to VEGFR2. In this study, we discovered that soluble endorepellin activates the canonical stress signaling pathway consisting of PERK, eIF2α, ATF4, and GADD45α. Specifically, endorepellin evoked transient activation of VEGFR2, which, in turn, phosphorylated PERK at Thr980. Subsequently, PERK phosphorylated eIF2α at Ser51, upregulating its downstream effector proteins ATF4 and GADD45α. RNAi-mediated knockdown of PERK or eIF2α abrogated the endorepellin-mediated up-regulation of GADD45α, the ultimate effector protein of this stress signaling cascade. To functionally validate these findings, we utilized an ex vivo model of angiogenesis. Exposure of the aortic rings embedded in 3D fibrillar collagen to recombinant endorepellin for 2–4 h activated PERK and induced GADD45α vis à vis vehicle-treated counterparts. Similar effects were obtained with the established cellular stress inducer tunicamycin. Notably, chronic exposure of aortic rings to endorepellin for 7–9 days markedly suppressed vessel sprouting, an angiostatic effect that was rescued by blocking PERK kinase activity. Our findings unravel a mechanism by which an extracellular matrix protein evokes stress signaling in endothelial cells, which leads to angiostasis.

Immigration is a prominent part of the United States’ DNA, despite concerns about immigrants’ ability to integrate. An examination of recent immigrant inflows shows newcomers to the United States are integrating well, based on language proficiency, socioeconomic attainment, political participation, residential locale, and social interaction indicators.

This final report summarizes and reflects upon the key findings of the Improving EU and U.S. Immigration Systems: Learning from Experience comparative research project undertaken by MPI and the European University Institute through a grant from the European Commission.

Migration and development have emerged as a pressing policy priority on the global agenda. This report identifies critical lessons from the past decade of policy experimentation and offers recommendations for migration and development policy.

BRIDGETOWN, Barbados (CMC): Former top West Indies cricket administrator, 76-year-old Deryck Murray, has warned against a forced restart of cricket amid the current challenges posed by the deadly coronavirus pandemic, and says any premature actions...

ROSEAU, Dominica (CMC): Well-travelled Barbadian umpire Gregory Brathwaite has been tipped to become the next Test umpire from the region. WEST INDIES Cricket Umpires Association secretary, Vivian Johnson, said the 50-year-old Brathwaite was...

Declaring it has been “pained” by the Jodian Fearon situation, the embattled Andrews Memorial Hospital (AMH) is speaking out, saying it has never been approached for or rejected efforts at registration or certification since it began operating 76...

For drummer Conroy Gordon, the hardest part of being locked away day after day in the narrow confines of a stranded cruise ship is his inability to tell his two daughters when Daddy will be home. For the past month, he has been battling this...

To say Judith-Miranda Townsend has a special love for children would not even begin to capture the essence of the Westmoreland supermom. Affectionately called ‘Mama Sweetie’ by foster children and members of the Holly Hill community in Darliston,...