A new children's book transforms a sad, scared and anxious little boy into a superhero. The book is called "Cape," in honor of the bright-red cape the little boy wears and finds comfort in following the death of his father. "Cape" is Kevin Johnson's debut picture book, and it's vividly illustrated by artist Kitt Thomas.

A North Texas man recently spent $4000 on a guitar he believed was signed by Taylor Swift at the Ellis County Wild Game Dinner in Waxahachie, Texas, and then immediately destroyed it with a hammer. Sounds like the guitar was an imposter. — Read the rest

The post Texas man wastes $4k smashing a Taylor Swift "themed" guitar appeared first on Boing Boing.

The following article, GOP Rep. Luna Says ‘Criminal Prosecutions Necessary’ For Anti-Trump Lawfare Schemers In Government (Video), was first published on Conservative Firing Line.

As President Donald Trump prepares to re-enter the White House after his landslide victory in Tuesday’s election, a U.S. congresswoman says “criminal prosecutions” are “necessary” for the government officials who have been promoting the massive lawfare campaign against the president-elect. On “Sunday Morning Futures” on the Fox News Channel, host Maria Bartiromo asked U.S. Rep. …

Continue reading GOP Rep. Luna Says ‘Criminal Prosecutions Necessary’ For Anti-Trump Lawfare Schemers In Government (Video) ...

A couple weeks ago, my pals at Twitter were kind enough to invite me in to visit with their (rapidly growing) team. The topic was meetings, so I used it as an opportunity to publicly premiere a talk I've been presenting to private clients over the past few months.

I hope you'll enjoy, Broken Meetings (and how you'll fix them).

Slides:

Supplementary links and commentary forthcoming, but I wanted to go ahead and post the talk as quickly as the video was available. Special thanks to Michelle, Jeremy, and the crackerjack Twitter crew for a swell afternoon.

I really like this talk and sincerely hope you will find it useful in helping to un-break your own meetings.

One of the greatest pass rushers in NFL history appears to be taking a pass himself when it comes to directly addressing a raging controversy. Pro Football Hall of Famer […]

The post NFL Analyst Michael Strahan Speaks Out After Becoming Embroiled in National Anthem Controversy appeared first on The Western Journal.

Nonphotochemical quenching (NPQ) is a mechanism of regulating light harvesting that protects the photosynthetic apparatus from photodamage by dissipating excess absorbed excitation energy as heat. In higher plants, the major light-harvesting antenna complex (LHCII) of photosystem (PS) II is directly involved in NPQ. The aggregation of LHCII is proposed to be involved in quenching. However, the lack of success in isolating native LHCII aggregates has limited the direct interrogation of this process. The isolation of LHCII in its native state from thylakoid membranes has been problematic because of the use of detergent, which tends to dissociate loosely bound proteins, and the abundance of pigment–protein complexes (e.g. PSI and PSII) embedded in the photosynthetic membrane, which hinders the preparation of aggregated LHCII. Here, we used a novel purification method employing detergent and amphipols to entrap LHCII in its natural states. To enrich the photosynthetic membrane with the major LHCII, we used Arabidopsis thaliana plants lacking the PSII minor antenna complexes (NoM), treated with lincomycin to inhibit the synthesis of PSI and PSII core proteins. Using sucrose density gradients, we succeeded in isolating the trimeric and aggregated forms of LHCII antenna. Violaxanthin- and zeaxanthin-enriched complexes were investigated in dark-adapted, NPQ, and dark recovery states. Zeaxanthin-enriched antenna complexes showed the greatest amount of aggregated LHCII. Notably, the amount of aggregated LHCII decreased upon relaxation of NPQ. Employing this novel preparative method, we obtained a direct evidence for the role of in vivo LHCII aggregation in NPQ.

Interview: Nicole Hemmer The World Today rescobales.drupal 27 September 2022

The political historian talks to Mike Higgins about how Donald Trump has transformed the US midterm elections and the Republican party’s lurch further right.

Partisans: The Conservative Revolutionaries Who Remade American Politics in the 1990s
Nicole Hemmer, Basic Books, $18.99

In Partisans, you argue that the American political far right had been preparing the conditions for Donald Trump for 25 years and ‘the puzzle pieces snapped into place’ when he was nominated in 2016. What did you mean by that?

There were changes happening in plain sight over a quarter of a century. Such as the rise of presidential candidates who had never held elected office but who had a following in conservative media and cable news more broadly. There was also a turn towards pessimism, to connections with extreme groups and to procedural extremism in the Republican Party, such as the impeachment of Bill Clinton over what most Americans agreed was a small indiscretion.

All that accelerates in the 2000s and 2010s to the point where if you look back from Trump’s election, he suddenly seems eligible in a way that I don’t think he was in 2015 during his ascendancy.

 How did the end of the Cold War open the door for the hard right in the early 1990s?

The Cold War is so important to making sense of conservative politics in the United States because it sat at the core of American conservatism for 50 years. Opposition to Soviet communism held together the conservative coalition. It brought together religious conservatives who talked about the atheistic Soviet Union with the libertarians who were fearful of a big state.

The Cold War forced conservatives to talk in a language of freedom and democracy because that was the way that the US was different from the Soviet Union. Ronald Reagan, in particular, was a master of this. It had real implications for domestic policy. Reagan embraced free trade and talked about immigration and diversity as things that were great for America.

And then the Cold War ends, and you have politicians like Pat Buchanan [a former White House Communications Director] who had been beating the Reagan line for years, coming out and saying: ‘You know what? Immigration is bad and immigrants are bad. They are harmful for the United States. Free Trade is bad. We need to have more protectionist economy and build Fortress America. And, by the way, democracy might not actually be the best form of government for our country. The Marine Corps and IBM are better run than the federal government, and those are autocratic governed institutions.’

It allowed a resurgence of a right-wing politics that the US hadn’t seen since the 1930s and 1940s. It would become a powerful political movement within conservatism that we ultimately see taking over the Republican Party by the 2010s.

 What was it that allowed far-right figures such as Rush Limbaugh to take advantage of the transformation of the media in the 1990s?

Part of what they saw in it was interactivity. We tend to associate that with the internet and with the rise of social media, but call-in national radio and cable TV gave people an opportunity to have their voices heard.

The most popular TV show on CNN was Larry King Live, a call-in show. That interactivity fed into this idea that a frustrated populace suddenly had a voice, and it was easy to play on that sense of frustration. Also, the right was innovative because they had been focused on alternative media for decades.

So, as you get new formats in the 1990s like political talk radio and cable news, conservatives were already in that space looking for ways to get their message across.

Also, journalistic objectivity in the 1990s became getting a voice from the right and a voice from the left, which opened the door for a new generation of conservative pundits such as TV hosts Laura Ingraham and Ann Coulter. You have these incentives drawing the right into this new media and all of these conservatives in the base who are looking for right-wing voices.

There are hearings into the January 6, 2021, Capitol invasion and an FBI investigation into the alleged presence of confidential papers at Trump’s residence Mar-a-Lago. Are we about to witness the insurgent far right within the Republican Party being brought to account legally?

There has been more accountability than I anticipated at the beginning of the Trump presidency. Some innovative legal techniques are being used successfully, such as sedition and conspiracy charges against some of the Capitol rioters.

Steve Bannon, one of Trump’s chief advisers in the 2016 campaign, has been charged with money laundering, conspiracy and fraud during the Trump presidency. Will that accountability lead to a fundamental transformation of right-wing politics in the US? I wouldn’t bet on that.

How is the radicalism of the Republican party shaping the upcoming mid-term elections?

The real radicalism in the Republican Party now is its willingness to use both procedural mechanisms but also mob violence to get the electoral outcomes that it wants – something that we saw with the insurrection on January 6, 2021.

That makes the mid-terms an existential election if Republicans are able to vote in candidates who are running on a platform of throwing out election results if Republicans don’t win. That is a real challenge to the future of American democracy.

As long as the political system is organized around one party that respects the rules of democracy and one party that fights against them, each election is going to be existential. It will make mid-term elections have the weight that the 2016 and 2020 presidential elections had.

Did the far right misunderstand the electorate when it comes to abortion, considering the strong reaction to the Dobbs ruling?

I think that is exactly right, and it’s fascinating. A major turn happens in American politics between the 1980s and the 1990s, especially on the right. Ronald Reagan built these broad coalitions. He was winning landslide elections, and building this ‘big tent’ party that would bring in people who didn’t identify as Republicans or conservatives.

By the time you get to the 1990s, you have this idea of the Rino – politicians who are ‘Republican in name only’. You had to be purer and purer when it came to your right-wing politics. And the Republican Party grew a lot more unpopular. It has lost elections because of the extremism of its candidates, but it has still been able to win democratic elections without majorities, something we saw in 2000 and 2016.

The outcome of it all is exactly what you are seeing right now, that in many ways the conservative movement has got something it wanted for half a century, which is the overturning of Roe v Wade – something that energized its space for a very long time. Now that the dog has caught the fire truck, it is looking around and saying: ‘Oh, people do not like this.’

August’s referendum in Kansas that upheld the right to abortion mobilized lots of voters who normally wouldn’t have come out for an off-year election.

As you get more stories of 10-year-old rape victims who have to go to a different state to get an abortion, or stories of women who have high-risk pregnancies not able to get medical care because of these new extremist laws, that really does mobilize a population who understand what is at stake.

It is not just their access to abortion, but to reproductive healthcare more broadly. Not everywhere, but in some places that is going to change the dynamics of these mid-term races.

Ulrich Rothbauer
Feb 1, 2008; 7:282-289
Research

David A. Mitchell
Jun 1, 2006; 47:1118-1127
Thematic Reviews

Chi-Liang Eric Yen
Nov 1, 2008; 49:2283-2301
Thematic Reviews

Mohamad Navab
Jun 1, 2004; 45:993-1007
Thematic Reviews

John M. Dietschy
Aug 1, 2004; 45:1375-1397
Thematic Reviews

Weerapan Khovidhunkit
Jul 1, 2004; 45:1169-1196
Thematic Reviews

Dawn L. Brasaemle
Dec 1, 2007; 48:2547-2559
Thematic Reviews

Expert

In animals, the response to chronic hypoxia is mediated by prolyl hydroxylases (PHDs) that regulate the levels of hypoxia-inducible transcription factor α (HIFα). PHD homologues exist in other types of eukaryotes and prokaryotes where they act on non HIF substrates. To gain insight into the factors underlying different PHD substrates and properties, we carried out biochemical and biophysical studies on PHD homologues from the cellular slime mold, Dictyostelium discoideum, and the protozoan parasite, Toxoplasma gondii, both lacking HIF. The respective prolyl-hydroxylases (DdPhyA and TgPhyA) catalyze prolyl-hydroxylation of S-phase kinase-associated protein 1 (Skp1), a reaction enabling adaptation to different dioxygen availability. Assays with full-length Skp1 substrates reveal substantial differences in the kinetic properties of DdPhyA and TgPhyA, both with respect to each other and compared with human PHD2; consistent with cellular studies, TgPhyA is more active at low dioxygen concentrations than DdPhyA. TgSkp1 is a DdPhyA substrate and DdSkp1 is a TgPhyA substrate. No cross-reactivity was detected between DdPhyA/TgPhyA substrates and human PHD2. The human Skp1 E147P variant is a DdPhyA and TgPhyA substrate, suggesting some retention of ancestral interactions. Crystallographic analysis of DdPhyA enables comparisons with homologues from humans, Trichoplax adhaerens, and prokaryotes, informing on differences in mobile elements involved in substrate binding and catalysis. In DdPhyA, two mobile loops that enclose substrates in the PHDs are conserved, but the C-terminal helix of the PHDs is strikingly absent. The combined results support the proposal that PHD homologues have evolved kinetic and structural features suited to their specific sensing roles.

A. G. Sergeev, A. Kh. Khachatryan and Kh. A. Khachatryan
Trans. Moscow Math. Soc. 83 (), 55-65.
Abstract, references and article information

‘Hong Kong is now in the hands of its people – they cannot rely on others to stick up for them now.’ Expert comment sysadmin 29 June 2017

Kerry Brown on ‘one country, two systems’, the UK’s diminishing influence and the territory’s future, 20 years after the handover.

1 July marks the 20th anniversary of the transfer of Hong Kong’s sovereignty from Britain to China. Kerry Brown speaks with Jason Naselli about what the future holds for the territory.

How sustainable is the ‘one country, two systems’ framework? Will the arrangement last the full 50 years (until 2047) as originally envisioned?

It is questionable whether the arrangement that exists today was the one envisioned in 1997 when the handover happened. It was always a very abstract, flexible system, granting Hong Kong a high degree of autonomy, meaning it could maintain its capitalist system. Of course, in the lead up to 1997 all these things were broadly seen as being in Beijing’s interests to preserve.

But these days, the one thing that few said in 1997 has come to pass – the People’s Republic has maintained one-party rule as a political system, but become one of the world’s great economies. It has been so far a huge success.

Hong Kong therefore has diminished in importance over the years to the point that maintaining at least some semblance of one country, two systems is almost like an act of charity. It has been nibbled at, compromised and seems to grow weaker by the day. Most in Hong Kong would say there is a system: one country, one system. That’s the deal.

The central government’s deepening involvement in the territory’s politics is a subject of growing controversy in Hong Kong. Does the Chinese government need to alter its approach?

Not particularly. It doesn’t want to see Hong Kong fail as an economy. That doesn’t suit its interests at all. But nor does it want a truculent, disobedient polity that is meant to be part of its sovereign territory.

So it has increasingly set political parameters. Hong Kong can have its unique system – as long as it is obedient. And on the whole, that is the deal that all of the city’s chief executives until now have internally understood perfectly.

As part of the 1997 handover, the UK has ‘a continuing moral and political obligation’ to Hong Kong. How will this relationship play out as Brexit shifts Britain’s place in the world?

The Foreign Office offers a six-monthly report to Parliament, updating on how the handover deal is going. As the years go on, however, it becomes increasingly illusive how the UK has any real locus to say much about the situation on Hong Kong. It did say, rightly, that the detainment of one of the booksellers taken in in 2015 was a violation of the treaty because he was British. This was the strongest wording that has ever appeared from an official British source. But with dependence on creating a new kind of relationship with China now foremost in people’s minds because of Brexit and other economic pressures, it is not surprising that the priority increasingly lies elsewhere.

With direct management of Hong Kong gone, the UK was always going to be more and more irrelevant. That has happened. And in any case, relations with China have had to become more complex and multifaceted. Hong Kong was always the tail wagging the dog for the UK relations with China. Now there has been a rebalancing, the calculation always has to be how much unilaterally supporting Hong Kong will damage relations with Beijing. This has become an increasingly asymmetrical question: in a playoff, preserving links with Beijing will always prevail. That’s just the reality of the new world we are seeing come into being.

Hong Kong has played an important financial role for China over the past 20 years, but where will it fit as markets and financial institutions on the mainland mature?

It maintains is role as a major RMB hub, and as a finance centre. But it is surrounded by competition. Singapore, and Sydney, and other places in the region have RMB deals. Shanghai and Tianjin aspire to be portals for entry to the domestic Chinese market. Hong Kong every day has to think of new ways to maintain its relevance and beat back competition. So far, it has done well. But this is an issue it can never be complacent about.

What has been the most significant change in Hong Kong society since 1997?

The rising cultural and linguistic influence of the mainland on Hong Kong. Hong Kong has maintained its difference – but it has had to change. It is clear that Hong Kong is now in the hands of Hong Kongese – they cannot rely on others to stick up for them now. The culture, identity and future of the territory are in their hands. In that sense, they have autonomy.

Thomas O. Gallouët, Andrea Natale and Gabriele Todeschi
Math. Comp. 93 (), 2769-2810.
Abstract, references and article information

Imelda Flores Vazquez from Econometrica, Inc. explains how economists use mathematics to evaluate the efficacy of health care policies. When a hospital or government wants to adjust their health policies — for instance, by encouraging more frequent screenings for certain diseases — how do they know whether their program will work or not? If the service has already been implemented elsewhere, researchers can use that data to estimate its effects. But if the idea is brand-new, or has only been used in very different settings, then it's harder to predict how well the new program will work. Luckily, a tool called a microsimulation can help researchers make an educated guess.

Dr. Abiy Tasissa of Tufts University, discusses the mathematics he and colleagues used to study particle collider data, including optimal transport and optimization. Collider physics often result in distributions referred to as jets. Dr. Tasissa and his team used "Earth Mover's Distance" and other mathematical tools to study the shape of jets. "It is interesting for me to see how mathematics can be applied to study these fundamental problems answering fundamental equations in physics, not only at the level of formulating new ideas, which is, in this particular case, a notion of distance, but also how the importance of designing fast optimization algorithms to be able to actually compute these distances," says Dr. Tasissa.

Frederic Heihoff
Proc. Amer. Math. Soc. 152 (), 5229-5247.
Abstract, references and article information

Neelarnab Raha
Proc. Amer. Math. Soc. 152 (), 5067-5081.
Abstract, references and article information

The Math Community Educational Outreach (Math CEO) program at the University of California, Irvine (UCI) will receive the 2025 AMS Award for an Exemplary Program or Achievement in a Mathematics Department.

Founded in 2014, UCI’s Math CEO is an after-school math enrichment program aimed at increasing the number of talented students in STEM from diverse backgrounds by fostering mathematical exploration, mentor development, and community engagement.

From the citation

The University of California, Irvine (UCI) Math CEO program is recognized for its exceptional contributions to the mathematics community and society at large. Established in 2014 by professors Alessandra Pantano and Li-Sheng Tseng, Math CEO targets students from Title I middle schools, providing them with a high-quality after-school math enrichment program. This program brings middle-school students to the UCI campus to work in small groups with undergraduate mentors, many of whom are also from historically marginalized groups, to engage in challenging mathematical tasks and encourage exploration. 

From September 2019 to June 2024, Math CEO engaged a total of 1,221 youth, with 48.6% identifying as female. The ethnic background of the participants was predominantly Latinx (93.5%), with smaller representations of Asian, white, and multiethnic students. In the same five-year period, Math CEO engaged 553 undergraduate mentors, 62.2% of whom were female. The mentors’ ethnic backgrounds were diverse, with significant representation of Asian (52%) and Latinx (30%) students. The undergraduate mentors, many of whom pursue careers in education, receive training in culturally responsive teaching practices and equity in education, significantly impacting their professional development. In a post-survey, 52.3% of the undergraduate mentors expressed interest in teaching or working in education and 45.9% were likely to pursue professions working with children or families.

Recognizing the central role of families in supporting Latinx youth, Math CEO involves parents through bilingual workshops that enhance community awareness of college pathways and financial opportunities. 

Math CEO has been the foundation for numerous research projects in mathematics education, supported by NSF grants, leading to publications and program growth. The program’s success is evident in its expansion to high schools and other regions in Southern California, including a new branch at California State University, Dominguez Hills. Math CEO continues to make a substantial impact on underserved youth, demonstrating a model of systemic, reproducible change that can be implemented by others.

Response of Alessandra Pantano, UCI Math CEO

I am deeply honored to receive the AMS Award for an Exemplary Program in a Mathematics Department on behalf of the UCI Math CEO team. This wonderful award recognizes the work of many colleagues, graduate students, and undergraduate students in developing and delivering the UCI Math Community Educational Outreach (Math CEO) program. For over a decade, Math CEO has provided creative and culturally responsive math enrichment opportunities for hundreds of underprivileged middle-school students, many of which have since “graduated” to high school or even college. Leading this exceptional and dedicated team of volunteers has been the highest pride of my professional life. A special thanks to my partners-in-crime, Prof. Li-Sheng Tseng, codirector of Math CEO, and former graduate student Andres Forero Cuervo, academic coordinator for Math CEO: We could have never done this without you. I look forward to pushing this activity forward and continuing to dedicate my energy to help kids in our county find the way to express their potential – in math and in life! A big thanks to the colleagues who nominated us and to the AMS for recognizing our efforts.  

History of the program

The UC Irvine Math Community Educational Outreach (Math CEO) program was founded in 2014 by math faculty Alessandra Pantano and Li-Sheng Tseng in collaboration with Santa Ana Unified math teacher Jasmina Matasovic. The founders shared a belief that low standardized test scores in underserved communities do not reflect students’ interest and potential to succeed in STEM. Math CEO runs free, weekly, after-school math enrichment sessions, welcoming all youth regardless of math achievement. Starting with only 25 students from one middle school, the program has grown and engaged nearly two thousand students in all, from multiple school districts in Southern California.

About the award

The annual AMS Award for an Exemplary Program or Achievement in a Mathematics Department was established in 2004 and first given in 2006. This award recognizes a department which has distinguished itself by undertaking an unusual or particularly effective program of value to the mathematics community, internally or in relation to the rest of society. Departments of mathematical sciences in North America that offer at least a bachelor’s degree in mathematical sciences are eligible. The award amount is currently $5,000. The award will be presented at the 2025 Joint Mathematics Meetings in Seattle.

Learn more about the award and previous recipients.

Contact: AMS Communications.

*****

The American Mathematical Society is dedicated to advancing research and connecting the diverse global mathematical community through our publications, meetings and conferences, MathSciNet, professional services, advocacy, and awareness programs.
 

Animals can sense the presence of microbes in their tissues and mobilize their own defenses by recognizing and responding to conserved microbial structures (often called microbe-associated molecular patterns (MAMPs)). Successful host defenses may kill the invaders, yet the host animal may fail to restore homeostasis if the stimulatory microbial structures are not silenced. Although mice have many mechanisms for limiting their responses to lipopolysaccharide (LPS), a major Gram-negative bacterial MAMP, a highly conserved host lipase is required to extinguish LPS sensing in tissues and restore homeostasis. We review recent progress in understanding how this enzyme, acyloxyacyl hydrolase (AOAH), transforms LPS from stimulus to inhibitor, reduces tissue injury and death from infection, prevents prolonged post-infection immunosuppression, and keeps stimulatory LPS from entering the bloodstream. We also discuss how AOAH may increase sensitivity to pulmonary allergens. Better appreciation of how host enzymes modify LPS and other MAMPs may help prevent tissue injury and hasten recovery from infection.

Infiltration of peripheral immune cells after blood-brain barrier dysfunction causes severe inflammation after a stroke. Although the endothelial glycocalyx, a network of membrane-bound glycoproteins and proteoglycans that covers the lumen of endothelial cells, functions as a barrier to circulating cells, the relationship between stroke severity and glycocalyx dysfunction remains unclear. In this study, glycosaminoglycans, a component of the endothelial glycocalyx, were studied in the context of ischemic stroke using a photochemically induced thrombosis mouse model. Decreased levels of heparan sulfate and chondroitin sulfate and increased activity of hyaluronidase 1 and heparanase (HPSE) were observed in ischemic brain tissues. HPSE expression in cerebral vessels increased after stroke onset and infarct volume greatly decreased after co-administration of N-acetylcysteine + glycosaminoglycan oligosaccharides as compared with N-acetylcysteine administration alone. These results suggest that the endothelial glycocalyx was injured after the onset of stroke. Interestingly, scission activity of proHPSE produced by immortalized endothelial cells and HEK293 cells transfected with hHPSE1 cDNA were activated by acrolein (ACR) exposure. We identified the ACR-modified amino acid residues of proHPSE using nano LC–MS/MS, suggesting that ACR modification of Lys139 (6-kDa linker), Lys107, and Lys161, located in the immediate vicinity of the 6-kDa linker, at least in part is attributed to the activation of proHPSE. Because proHPSE, but not HPSE, localizes outside cells by binding with heparan sulfate proteoglycans, ACR-modified proHPSE represents a promising target to protect the endothelial glycocalyx.

RNase J enzymes are metallohydrolases that are involved in RNA maturation and RNA recycling, govern gene expression in bacteria, and catalyze both exonuclease and endonuclease activity. The catalytic activity of RNase J is regulated by multiple mechanisms which include oligomerization, conformational changes to aid substrate recognition, and the metal cofactor at the active site. However, little is known of how RNase J paralogs differ in expression and activity. Here we describe structural and biochemical features of two Staphylococcus epidermidis RNase J paralogs, RNase J1 and RNase J2. RNase J1 is a homodimer with exonuclease activity aided by two metal cofactors at the active site. RNase J2, on the other hand, has endonuclease activity and one metal ion at the active site and is predominantly a monomer. We note that the expression levels of these enzymes vary across Staphylococcal strains. Together, these observations suggest that multiple interacting RNase J paralogs could provide a strategy for functional improvisation utilizing differences in intracellular concentration, quaternary structure, and distinct active site architecture despite overall structural similarity.

Connexin (Cx) protein forms hemichannels and gap junctional channels, which play diverse and profound roles in human physiology and diseases. Gap junctions are arrays of intercellular channels formed by the docking of two hemichannels from adjacent cells. Each hexameric hemichannel contains the same or different Cx isoform. Although homomeric Cxs forms have been largely described functionally and structurally, the stoichiometry and arrangement of heteromeric Cx channels remain unknown. The latter, however, are widely expressed in human tissues and variation might have important implications on channel function. Investigating properties of heteromeric Cx channels is challenging considering the high number of potential subunit arrangements and stoichiometries, even when only combining two Cx isoforms. To tackle this problem, we engineered an HA tag onto Cx26 or Cx30 subunits and imaged hemichannels that were liganded by Fab-epitope antibody fragments via atomic force microscopy. For Cx26-HA/Cx30 or Cx30-HA/Cx26 heteromeric channels, the Fab-HA binding distribution was binomial with a maximum of three Fab-HA bound. Furthermore, imaged Cx26/Cx30-HA triple liganded by Fab-HA showed multiple arrangements that can be derived from the law of total probabilities. Atomic force microscopy imaging of ringlike structures of Cx26/Cx30-HA hemichannels confirmed these findings and also detected a polydisperse distribution of stoichiometries. Our results indicate a dominant subunit stoichiometry of 3Cx26:3Cx30 with the most abundant subunit arrangement of Cx26-Cx26-Cx30-Cx26-Cx30-Cx30. To our knowledge, this is the first time that the molecular architecture of heteromeric Cx channels has been revealed, thus providing the basis to explore the functional effect of these channels in biology.

Heme oxygenase-2 (HO2) and -1 (HO1) catalyze heme degradation to biliverdin, CO, and iron, forming an essential link in the heme metabolism network. Tight regulation of the cellular levels and catalytic activities of HO1 and HO2 is important for maintaining heme homeostasis. HO1 expression is transcriptionally regulated; however, HO2 expression is constitutive. How the cellular levels and activity of HO2 are regulated remains unclear. Here, we elucidate the mechanism of post-translational regulation of cellular HO2 levels by heme. We find that, under heme-deficient conditions, HO2 is destabilized and targeted for degradation, suggesting that heme plays a direct role in HO2 regulation. HO2 has three heme binding sites: one at its catalytic site and the others at its two heme regulatory motifs (HRMs). We report that, in contrast to other HRM-containing proteins, the cellular protein level and degradation rate of HO2 are independent of heme binding to the HRMs. Rather, under heme deficiency, loss of heme binding to the catalytic site destabilizes HO2. Consistently, an HO2 catalytic site variant that is unable to bind heme exhibits a constant low protein level and an enhanced protein degradation rate compared with the WT HO2. Finally, HO2 is degraded by the lysosome through chaperone-mediated autophagy, distinct from other HRM-containing proteins and HO1, which are degraded by the proteasome. These results reveal a novel aspect of HO2 regulation and deepen our understanding of HO2's role in maintaining heme homeostasis, paving the way for future investigation into HO2's pathophysiological role in heme deficiency response.

Aspergillus terreus is an allergenic fungus, in addition to causing infections in both humans and plants. However, the allergens in this fungus are still unknown, limiting the development of diagnostic and therapeutic strategies. We used a proteomic approach to search for allergens, identifying 16 allergens based on two-dimensional immunoblotting with A. terreus susceptible patient sera. We further characterized triose-phosphate isomerase (Asp t 36), one of the dominant IgE (IgE)-reactive proteins. The gene was cloned and expressed in Escherichia coli. Phylogenetic analysis showed Asp t 36 to be highly conserved with close similarity to the triose-phosphate isomerase protein sequence from Dermatophagoides farinae, an allergenic dust mite. We identified four immunodominant epitopes using synthetic peptides, and mapped them on a homology-based model of the tertiary structure of Asp t 36. Among these, two were found to create a continuous surface patch on the 3D structure, rendering it an IgE-binding hotspot. Biophysical analysis indicated that Asp t 36 shows similar secondary structure content and temperature sensitivity with other reported triose-phosphate isomerase allergens. In vivo studies using a murine model displayed that the recombinant Asp t 36 was able to stimulate airway inflammation, as demonstrated by an influx of eosinophils, goblet cell hyperplasia, elevated serum Igs, and induction of Th2 cytokines. Collectively, our results reveal the immunogenic property of Asp t 36, a major allergen from A. terreus, and define a new fungal allergen more broadly. This allergen could serve as a potent candidate for investigating component resolved diagnosis and immunotherapy.

Pectins are a major dietary nutrient source for the human gut microbiota. The prominent gut microbe Bacteroides thetaiotaomicron was recently shown to encode the founding member (BT1017) of a new family of pectin methylesterases essential for the metabolism of the complex pectin rhamnogalacturonan-II (RG-II). However, biochemical and structural knowledge of this family is lacking. Here, we showed that BT1017 is critical for the metabolism of an RG-II–derived oligosaccharide ΔBT1017oligoB generated by a BT1017 deletion mutant (ΔBT1017) during growth on carbohydrate extract from apple juice. Structural analyses of ΔBT1017oligoB using a combination of enzymatic, mass spectrometric, and NMR approaches revealed that it is a bimethylated nonaoligosaccharide (GlcA-β1,4-(2-O-Me-Xyl-α1,3)-Fuc-α1,4-(GalA-β1,3)-Rha-α1,3-Api-β1,2-(Araf-α1,3)-(GalA-α1,4)-GalA) containing components of the RG-II backbone and its side chains. We showed that the catalytic module of BT1017 adopts an α/β-hydrolase fold, consisting of a central twisted 10-stranded β-sheet sandwiched by several α-helices. This constitutes a new fold for pectin methylesterases, which are predominantly right-handed β-helical proteins. Bioinformatic analyses revealed that the family is dominated by sequences from prominent genera of the human gut microbiota, including Bacteroides and Prevotella. Our re-sults not only highlight the critical role played by this family of enzymes in pectin metabolism but also provide new insights into the molecular basis of the adaptation of B. thetaiotaomicron to the human gut.

Visual Abstract

Visual Abstract

Alba Simats
Dec 1, 2020; 19:1921-1935
Research

Daniel J. Geiszler
Dec 1, 2020; 0:TIR120.002216v1-mcp.TIR120.002216
Technological Innovation and Resources

Mutations in the GUCY2D gene coding for the dimeric human retinal membrane guanylyl cyclase (RetGC) isozyme RetGC1 cause various forms of blindness, ranging from rod dysfunction to rod and cone degeneration. We tested how the mutations causing recessive congenital stationary night blindness (CSNB), recessive Leber's congenital amaurosis (LCA1), and dominant cone–rod dystrophy-6 (CORD6) affected RetGC1 activity and regulation by RetGC-activating proteins (GCAPs) and retinal degeneration-3 protein (RD3). CSNB mutations R666W, R761W, and L911F, as well as LCA1 mutations R768W and G982VfsX39, disabled RetGC1 activation by human GCAP1, -2, and -3. The R666W and R761W substitutions compromised binding of GCAP1 with RetGC1 in HEK293 cells. In contrast, G982VfsX39 and L911F RetGC1 retained the ability to bind GCAP1 in cyto but failed to effectively bind RD3. R768W RetGC1 did not bind either GCAP1 or RD3. The co-expression of GUCY2D allelic combinations linked to CSNB did not restore RetGC1 activity in vitro. The CORD6 mutation R838S in the RetGC1 dimerization domain strongly dominated the Ca2+ sensitivity of cyclase regulation by GCAP1 in RetGC1 heterodimer produced by co-expression of WT and the R838S subunits. It required higher Ca2+ concentrations to decelerate GCAP-activated RetGC1 heterodimer—6-fold higher than WT and 2-fold higher than the Ser838-harboring homodimer. The heterodimer was also more resistant than homodimers to inhibition by RD3. The observed biochemical changes can explain the dominant CORD6 blindness and recessive LCA1 blindness, both of which affect rods and cones, but they cannot explain the selective loss of rod function in recessive CSNB.

The kinesin-3 family contains the fastest and most processive motors of the three neuronal transport kinesin families, yet the sequence of states and rates of kinetic transitions that comprise the chemomechanical cycle and give rise to their unique properties are poorly understood. We used stopped-flow fluorescence spectroscopy and single-molecule motility assays to delineate the chemomechanical cycle of the kinesin-3, KIF1A. Our bacterially expressed KIF1A construct, dimerized via a kinesin-1 coiled-coil, exhibits fast velocity and superprocessivity behavior similar to WT KIF1A. We established that the KIF1A forward step is triggered by hydrolysis of ATP and not by ATP binding, meaning that KIF1A follows the same chemomechanical cycle as established for kinesin-1 and -2. The ATP-triggered half-site release rate of KIF1A was similar to the stepping rate, indicating that during stepping, rear-head detachment is an order of magnitude faster than in kinesin-1 and kinesin-2. Thus, KIF1A spends the majority of its hydrolysis cycle in a one-head-bound state. Both the ADP off-rate and the ATP on-rate at physiological ATP concentration were fast, eliminating these steps as possible rate-limiting transitions. Based on the measured run length and the relatively slow off-rate in ADP, we conclude that attachment of the tethered head is the rate-limiting transition in the KIF1A stepping cycle. Thus, KIF1A's activity can be explained by a fast rear-head detachment rate, a rate-limiting step of tethered-head attachment that follows ATP hydrolysis, and a relatively strong electrostatic interaction with the microtubule in the weakly bound post-hydrolysis state.

This article has been withdrawn by the authors as part of this review overlapped with the contents of Pietrangelo A and Ridgway ND. 2018. Cellular and Molecular Life Sciences. 75; 3079-98.

Photoreceptors have high energy-demands and a high density of mitochondria that produce adenosine triphosphate (ATP) through oxidative phosphorylation (OXPHOS) of fuel substrates. Although glucose is the major fuel for central nervous system (CNS) brain neurons, in photoreceptors (also CNS), most glucose is not metabolized through OXPHOS but is instead metabolized into lactate by aerobic glycolysis. The major fuel sources for photoreceptor mitochondria remained unclear for almost six decades. Similar to other tissues (like heart and skeletal muscle) with high metabolic rates, photoreceptors were recently found to metabolize fatty acids (palmitate) through OXPHOS. Disruption of lipid entry into photoreceptors leads to extracellular lipid accumulation, suppressed glucose transporter expression, and a duel lipid/glucose fuel shortage. Modulation of lipid metabolism helps restore photoreceptor function. However, further elucidation of the types of lipids used as retinal energy sources, the metabolic interaction with other fuel pathways, as well as the crosstalk among retinal cells to provide energy to photoreceptors is not yet known. In this review, we will focus on the current understanding of photoreceptor energy demand and sources, and potential future investigations of photoreceptor metabolism.

Vitamin A aldehyde covalently bound to opsin protein is embedded in a phospholipid-rich membrane that supports photon absorption and phototransduction in photoreceptor cell outer segments. Following absorption of a photon, the 11-cis-retinal chromophore of visual pigment in photoreceptor cells isomerizes to all-trans-retinal. To maintain photosensitivity 11-cis-retinal must be replaced. At the same time, however, all-trans-retinal has to be handled so as to prevent nonspecific aldehyde activity. Some molecules of retinaldehyde upon release from opsin are efficiently reduced to retinol. Other molecules are released into the lipid phase of the disc membrane where they form a conjugate (N-retinylidene-PE, NRPE) through a Schiff base linkage with phosphatidylethanolamine (PE). The reversible formation of NRPE serves as a transient sink for retinaldehyde that is intended to return retinaldehyde to the visual cycle. However, if instead of hydrolyzing to PE and retinaldehyde, NRPE reacts with a second molecule of retinaldehyde a synthetic pathway is initiated that leads to the formation of multiple species of unwanted bisretinoid fluorophores. We report on recently identified members of the bisretinoid family some of which differ with respect to the acyl chains associated with the glycerol backbone. We discuss processing of the lipid moieties of these fluorophores in lysosomes of retinal pigment epithelial (RPE) cells, their fluorescence characters and new findings related to light and iron-associated oxidation of bisretinoids.

Driven by the energy of a photon, the visual pigments in rod and cone photoreceptor cells isomerize 11-cis-retinal to the all-trans configuration. This photochemical reaction initiates the signal transduction pathway that eventually leads to the transmission of a visual signal to the brain and leaves the opsins insensitive to further light stimulation. For the eye to restore light sensitivity, opsins require recharging with 11-cis-retinal. This trans–cis back conversion is achieved through a series of enzymatic reactions composing the retinoid (visual) cycle. Although it is evident that the classical retinoid cycle is critical for vision, the existence of an adjunct pathway for 11-cis-retinal regeneration has been debated for many years. Retinal pigment epithelium (RPE)–retinal G protein-coupled receptor (RGR) has been identified previously as a mammalian retinaldehyde photoisomerase homologous to retinochrome found in invertebrates. Using pharmacological, genetic, and biochemical approaches, researchers have now established the physiological relevance of the RGR in 11-cis-retinal regeneration. The photoisomerase activity of RGR in the RPE and Müller glia explains how the eye can remain responsive in daylight. In this review, we will focus on retinoid metabolism in the eye and visual chromophore regeneration mediated by RGR.  

The field of phosphoinositide signaling has expanded significantly in recent years. Phosphoinositides (PIs) are universal signaling molecules that directly interact with membrane proteins or with cytosolic proteins containing domains that directly bind phosphoinositides and are recruited to cell membranes. Through the activities of PI kinases and PI phosphatases, seven distinct phosphoinositide lipid molecules are formed from the parent molecule phosphatidylinositol. PI signals regulate a wide range of cellular functions, including cytoskeletal assembly, membrane binding and fusion, ciliogenesis, vesicular transport, and signal transduction. Given the many excellent reviews on phosphoinositide kinases, phosphoinositide phosphatases, and PIs in general, in this review, we discuss recent studies and advances in PI lipid signaling in the retina. We specifically focus on PI lipids from vertebrate (e.g. bovine, rat, mice, toad, and zebrafish) and invertebrate (e.g. drosophila, horseshoe crab, and squid) retinas. We also discuss the importance of PIs revealed from animal models and human diseases, and methods to study PI levels both in vitro and in vivo. We propose that future studies should investigate the function and mechanism of activation of PI-modifying enzymes/phosphatases and further unravel PI regulation and function in the different cell types of the retina.

     Lens and tear film lipids are as unique as the systems they reside in. The major lipid of the human lens is dihydrosphingomylein, found in quantity only in the lens. The lens contains a cholesterol to phospholipid molar ratio as high as 10:1, more than anywhere in the body. Lens lipids contribute to maintaining lens clarity, and alterations in lens lipid composition due to age are likely to contribute to cataract. Lens lipid composition reflects adaptations to the unique characteristics of the lens: no turnover of lens lipids or proteins; the lowest amount of oxygen than any other tissue and contains almost no intracellular organelles. The tear film lipid layer (TFLL) is also unique. The TFLL is a thin, 100 nm layer of lipid on the surface of tears covering the cornea that contributes to tear film stability. The major lipids of the TFLL are wax esters and cholesterol esters that are not found in the lens. The hydrocarbon chains associated with the esters are longer than those found anywhere in the body, as long as 32 carbons, and many are branched. Changes in the composition and structure of the 30,000 different moieties of TFLL contribute to the instability of tears. The focus of the current review is how spectroscopy has been used to elucidate the relationships between lipid composition, conformational order and function and the etiology of cataract and dry eye.