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There is substantial evidence for extensive nonvesicular sterol transport in cells. For example, lipid transfer by the steroidogenic acute regulator-related proteins (StarD) containing a StarT domain has been shown to involve several pathways of nonvesicular trafficking. Among the soluble StarT domain-containing proteins, StarD4 is expressed in most tissues and has been shown to be an effective sterol transfer protein. However, it was unclear whether the lipid composition of donor or acceptor membranes played a role in modulating StarD4-mediated transport. Here, we used fluorescence-based assays to demonstrate a phosphatidylinositol phosphate (PIP)-selective mechanism by which StarD4 can preferentially extract sterol from liposome membranes containing certain PIPs (especially, PI(4,5)P2 and to a lesser degree PI(3,5)P2). Monophosphorylated PIPs and other anionic lipids had a smaller effect on sterol transport. This enhancement of transport was less effective when the same PIPs were present in the acceptor membranes. Furthermore, using molecular dynamics (MD) simulations, we mapped the key interaction sites of StarD4 with PIP-containing membranes and identified residues that are important for this interaction and for accelerated sterol transport activity. We show that StarD4 recognizes membrane-specific PIPs through specific interaction with the geometry of the PIP headgroup as well as the surrounding membrane environment. Finally, we also observed that StarD4 can deform membranes upon longer incubations. Taken together, these results suggest a mechanism by which PIPs modulate cholesterol transfer activity via StarD4.

The motor protein myosin-15 is necessary for the development and maintenance of mechanosensory stereocilia, and mutations in myosin-15 cause hereditary deafness. In addition to transporting actin regulatory machinery to stereocilia tips, myosin-15 directly nucleates actin filament ("F-actin") assembly, which is disrupted by a progressive hearing loss mutation (p.D1647G, "Jordan"). Here, we present cryo-electron microscopy structures of myosin-15 bound to F-actin, providing a framework for interpreting the impacts of deafness mutations on motor activity and actin nucleation. Rigor myosin-15 evokes conformational changes in F-actin yet maintains flexibility in actin's D-loop, which mediates inter-subunit contacts, while the Jordan mutant locks the D-loop in a single conformation. Adenosine diphosphate-bound myosin-15 also locks the D-loop, which correspondingly blunts actin-polymerization stimulation. We propose myosin-15 enhances polymerization by bridging actin protomers, regulating nucleation efficiency by modulating actin's structural plasticity in a myosin nucleotide state-dependent manner. This tunable regulation of actin polymerization could be harnessed to precisely control stereocilium height.

The Omicron variant of SARS-CoV-2 infected many vaccinated and convalescent individuals(1-)(3). Despite the reduced protection from infection, individuals who received three doses of an mRNA vaccine were highly protected from more serious consequences of infection(4). Here we examine the memory B cell repertoire in a longitudinal cohort of individuals receiving three mRNA vaccine doses(5,6). We find that the third dose is accompanied by an increase in, and evolution of, receptor-binding domain (RBD)-specific memory B cells. The increase is due to expansion of memory B cell clones that were present after the second dose as well as the emergence of new clones. The antibodies encoded by these cells showed significantly increased potency and breadth when compared with antibodies obtained after the second dose. Notably, the increase in potency was especially evident among newly developing clones of memory cells, which differed from persisting clones in targeting more conserved regions of the RBD. Overall, more than 50% of the analysed neutralizing antibodies in the memory compartment after the third mRNA vaccine dose neutralized the Omicron variant. Thus, individuals receiving three doses of an mRNA vaccine have a diverse memory B cell repertoire that can respond rapidly and produce antibodies capable of clearing even diversified variants such as Omicron. These data help to explain why a third dose of a vaccine that was not specifically designed to protect against variants is effective against variant-induced serious disease.

Trafficking of M-protein (Mprt) from the cytosol of Group A Streptococcus pyogenes (GAS) occurs via Sec translocase membrane channels that associate with Sortase A (SrtA), an enzyme that catalyzes cleavage of Mprt at the proximal C-terminal [-LPST(355*)GEAA-] motif and subsequent transpeptidation of the Mprt-containing product to the cell wall (CW). These steps facilitate stable exposure of the N-terminus of Mprt to the extracellular milieu where it interacts with ligands. Previously, we found that inactivation of SrtA in GAS cells eliminated Mprt CW transpeptidation but effected little reduction in its cell surface exposure, indicating that the C-terminus of Mprt retained in the cytoplasmic membrane (CM) extends its N-terminus to the cell surface. Herein, we assessed the effects of mutating the Thr(355) residue in the WT SrtA consensus sequence (LPST(355*)GEAA-) in a specific Mprt, PAM. In vitro, we found that synthetic peptides with mutations (LPSX(355)GEAA) in the SrtA cleavage site displayed slower cleavage activities with rSrtA than the WT peptide. Aromatic residues at X had the lowest activities. Nonetheless, PAM/[Y(355)G] still transpeptidated the CW in vivo. However, when using isolated CMs from srtA-inactivated GAS cells, rapid cleavage of PAM/[LPSY(355)GEAA] occurred at E-357* but transpeptidation did not take place. These results show that another CM-resident enzyme nonproductively cleaved PAM/[LPSYGE(357*)AA]. However, SrtA associated with the translocon channel in vivo cleaved and transpeptidated PAM/ [LPSX(355*)GEAA] variants. These CM features allow diverse cleavage site variants to covalently attach to the CW despite the presence of other potent nonproductive CM proteases.

PD-1 blockade (nivolumab) efficacy remains modest for metastatic sarcoma. In this paper, we present an open-label, non-randomized, non-comparative pilot study of bempegaldesleukin, a CD122-preferential interleukin-2 pathway agonist, with nivolumab in refractory sarcoma at Memorial Sloan Kettering/MD Anderson Cancer Centers (NCT03282344). We report on the primary outcome of objective response rate (ORR) and secondary endpoints of toxicity, clinical benefit, progression-free survival, overall survival, and durations of response/treatment. In 84 patients in 9 histotype cohorts, all patients experienced >= 1 adverse event and treatment-related adverse event; 1 death was possibly treatment-related. ORR was highest in angiosarcoma (3/8) and undifferentiated pleomorphic sarcoma (2/10), meeting predefined endpoints. Results of our exploratory investigation of predictive biomarkers show: CD8 + T cell infiltrates and PD-1 expression correlate with improved ORR; upregulation of immune-related pathways correlate with improved efficacy; Hedgehog pathway expression correlate with resistance. Exploration of this combination in selected sarcomas, and of Hedgehog signaling as a predictive biomarker, warrants further study in larger cohorts. The activity of PD-1 blockade in patients with sarcoma has been modest so far. Here, the authors report the results of a pilot clinical trial to assess the efficacy and safety of bempegaldesleukin, a CD122-preferential interleukin-2 (IL-2) pathway agonist, in combination with the PD1 blockade (nivolumab) in patients with locally advanced or metastatic high-grade sarcoma.

Microbiome data have several specific characteristics (sparsity and compositionality) that introduce challenges in data analysis. The integration of prior information regarding the data structure, such as phylogenetic structure and repeated-measure study designs, into analysis, is an effective approach for revealing robust patterns in microbiome data. Past methods have addressed some but not all of these challenges and features: for example, robust principal-component analysis (RPCA) addresses sparsity and compositionality; compositional tensor factorization (CIF) addresses sparsity, compositionality, and repeated measure study designs; and UniFrac incorporates phylogenetic information. Here we introduce a strategy of incorporating phylogenetic information into RPCA and CTF. The resulting methods, phylo-RPCA, and phylo-CTF, provide substantial improvements over state-of-the-art methods in terms of discriminatory power of underlying clustering ranging from the mode of delivery to adult human lifestyle. We demonstrate quantitatively that the addition of phylogenetic information improves effect size and classification accuracy in both data-driven simulated data and real microbiome data. IMPORTANCE Microbiome data analysis can be difficult because of particular data features, some unavoidable and some due to technical limitations of DNA sequencing instruments. The first step in many analyses that ultimately reveals patterns of similarities and differences among sets of samples (e.g., separating samples from sick and healthy people or samples from seawater versus soil) is calculating the difference between each pair of samples. We introduce two new methods to calculate these differences that combine features of past methods, specifically being able to take into account the principles that most types of microbes are not in most samples (sparsity), that abundances are relative rather than absolute (compositionality), and that all microbes have a shared evolutionary history (phylogeny). We show using simulated and real data that our new methods provide improved classification accuracy of ordinal sample clusters and increased effect size between sample groups on beta-diversity distances.

Antiretroviral therapy is highly effective in suppressing human immunodeficiency virus (HIV)(1). However, eradication of the virus in individuals with HIV has not been possible to date(2). Given that HIV suppression requires life-long antiretroviral therapy, predominantly on a daily basis, there is a need to develop clinically effective alternatives that use long-acting antiviral agents to inhibit viral replication(3). Here we report the results of a two-component clinical trial involving the passive transfer of two HIV-specific broadly neutralizing monoclonal antibodies, 3BNC117 and 10-1074. The first component was a randomized, double-blind, placebo-controlled trial that enrolled participants who initiated antiretroviral therapy during the acute/early phase of HIV infection. The second component was an open-label single-arm trial that enrolled individuals with viraemic control who were naive to antiretroviral therapy. Up to 8 infusions of 3BNC117 and 10-1074, administered over a period of 24 weeks, were well tolerated without any serious adverse events related to the infusions. Compared with the placebo, the combination broadly neutralizing monoclonal antibodies maintained complete suppression of plasma viraemia (for up to 43 weeks) after analytical treatment interruption, provided that no antibody-resistant HIV was detected at the baseline in the study participants. Similarly, potent HIV suppression was seen in the antiretroviral-therapy-naive study participants with viraemia carrying sensitive virus at the baseline. Our data demonstrate that combination therapy with broadly neutralizing monoclonal antibodies can provide long-term virological suppression without antiretroviral therapy in individuals with HIV, and our experience offers guidance for future clinical trials involving next-generation antibodies with long half-lives.

Subacute sclerosing panencephalitis, a late complication of measles, is still present during epidemics of this disease due to insufficient vaccination. After a historical review, the importance of the diagnostic criteria and the pathophysiology of SSEP are discussed. Numerous studies on the parameters of innate immunity and interferon responses tend to show a decrease in the activity of cellular immunity. Several hypotheses are formulated based on the publications of the different forms of the disease: Congenital, perinatal, forms with short incubation similar to acute inclusion encephalitis (AIE), rapidly evolving forms, forms of the immunocompromised, or even adults. Familial forms have been identified, suggesting a genetic cause. Based on the duration of the latency period, two groups have been individualized, prompting a retrospective and prospective analysis of the exomes of these patients. The knowledge of the genes involved should be useful for the understanding of the pathophysiology of SSPE and other late neurological RNA virus infections.

Background: Psoriasis, a chronic inflammatory disease dependent on the IL-23/T(H)17 pathway, is initiated through plasmacytoid dendritic cell activation and type I IFN induction in the skin. Deucravacitinib, a selective tyrosine kinase 2 (TYK2) inhibitor, blocks IL-23, IL-12, and type I IFN signaling in cellular assays. Objective: We investigated changes in IL-23/T(H)17 and type I IFN pathway biomarkers and gene responses as well as measures of selectivity for TYK2 over Janus kinases (JAKs) 1-3 in patients with moderate to severe psoriasis receiving deucravacitinib. Methods: Deucravacitinib was evaluated in a randomized, placebo-controlled, dose-ranging trial. Biopsy samples from nonlesional (day 1) and lesional skin (days 1, 15, and 85) were assessed for changes in IL-23/IL-12 and type I IFN pathway biomarkers by quantitative reverse-transcription polymerase chain reaction, RNA sequencing, and immunohistochemistry. Laboratory markers were measured in blood. Percentage change from baseline in Psoriasis Area and Severity Index (PASI) score was assessed. Results: IL-23 pathway biomarkers in lesional skin returned toward nonlesional levels dose-dependently with deucravacitinib. IFN and IL-12 pathway genes were normalized. Markers of keratinocyte dysregulation, keratin-16, and beta-defensin genes approached nonlesional levels with effective doses. Select laboratory parameters affected by JAK1-3 inhibition were not affected by deucravacitinib. Greater improvements in PASI scores, correlated with biomarker changes, were seen with the highest doses of deucravacitinib versus lower doses or placebo. Conclusion: Robust clinical efficacy with deucravacitinib treatment was associated with decreases in IL-23/TH17 and IFN pathway biomarkers. The lack of effect seen on biomarkers specific to JAK1-3 inhibition supports selectivity of deucravacitinib for TYK2; larger confirmatory studies are needed.