Structural Mechanism for GSDMD Targeting by Autoprocessed Caspases in Pyroptosis.

The pyroptosis execution protein GSDMD is cleaved by inflammasome-activated caspase-1 and LPS-activated caspase-11/4/5. The cleavage unmasks the pore-forming domain from GSDMD-C-terminal domain. How the caspases recognize GSDMD and its connection with caspase activation are unknown. Here, we show site-specific caspase-4/11 autoprocessing, generating a p10 product, is required and sufficient for cleaving GSDMD and inducing pyroptosis. The p10-form autoprocessed caspase-4/11 binds the GSDMD-C domain with a high affinity. Structural comparison of autoprocessed and unprocessed capase-11 identifies a ß sheet induced by the autoprocessing. In caspase-4/11-GSDMD-C complex crystal structures, the ß sheet organizes a hydrophobic GSDMD-binding interface that is only possible for p10-form caspase-4/11. The binding promotes dimerization-mediated caspase activation, rendering a cleavage independently of the cleavage-site tetrapeptide sequence. Crystal structure of caspase-1-GSDMD-C complex shows a similar GSDMD-recognition mode. Our study reveals an unprecedented substrate-targeting mechanism for caspases. The hydrophobic interface suggests an additional space for developing inhibitors specific for pyroptotic caspases.

Shigella evades pyroptosis by arginine ADP-riboxanation of caspase-11.

Mouse caspase-11 and human caspase-4 and caspase-5 recognize cytosolic lipopolysaccharide (LPS) to induce pyroptosis by cleaving the pore-forming protein GSDMD1-5. This non-canonical inflammasome defends against Gram-negative bacteria6,7. Shigella flexneri, which causes bacillary dysentery, lives freely within the host cytosol where these caspases reside. However, the role of caspase-11-mediated pyroptosis in S. flexneri infection is unknown. Here we show that caspase-11 did not protect mice from S. flexneri infection, in contrast to infection with another cytosolic bacterium, Burkholderia thailandensis8. S. flexneri evaded pyroptosis mediated by caspase-11 or caspase 4 (hereafter referred to as caspase-11/4) using a type III secretion system (T3SS) effector, OspC3. OspC3, but not its paralogues OspC1 and 2, covalently modified caspase-11/4; although it used the NAD+ donor, this modification was not ADP-ribosylation. Biochemical dissections uncovered an ADP-riboxanation modification on Arg314 and Arg310 in caspase-4 and caspase-11, respectively. The enzymatic activity was shared by OspC1 and 2, whose ankyrin-repeat domains, unlike that of OspC3, could not recognize caspase-11/4. ADP-riboxanation of the arginine blocked autoprocessing of caspase-4/11 as well as their recognition and cleavage of GSDMD. ADP-riboxanation of caspase-11 paralysed pyroptosis-mediated defence in Shigella-infected mice and mutation of ospC3 stimulated caspase-11- and GSDMD-dependent anti-Shigella humoral immunity, generating a vaccine-like protective effect. Our study establishes ADP-riboxanation of arginine as a bacterial virulence mechanism that prevents LPS-induced pyroptosis.

Whole genome sequencing based analysis of inflammation biomarkers in the Trans-Omics for Precision Medicine (TOPMed) consortium.

Inflammation biomarkers can provide valuable insight into the role of inflammatory processes in many diseases and conditions. Sequencing based analyses of such biomarkers can also serve as an exemplar of the genetic architecture of quantitative traits. To evaluate the biological insight, which can be provided by a multi-ancestry, whole-genome based association study, we performed a comprehensive analysis of 21 inflammation biomarkers from up to 38 465 individuals with whole-genome sequencing from the Trans-Omics for Precision Medicine (TOPMed) program (with varying sample size by trait, where the minimum sample size was n = 737 for MMP-1). We identified 22 distinct single-variant associations across 6 traits-E-selectin, intercellular adhesion molecule 1, interleukin-6, lipoprotein-associated phospholipase A2 activity and mass, and P-selectin-that remained significant after conditioning on previously identified associations for these inflammatory biomarkers. We further expanded upon known biomarker associations by pairing the single-variant analysis with a rare variant set-based analysis that further identified 19 significant rare variant set-based associations with 5 traits. These signals were distinct from both significant single variant association signals within TOPMed and genetic signals observed in prior studies, demonstrating the complementary value of performing both single and rare variant analyses when analyzing quantitative traits. We also confirm several previously reported signals from semi-quantitative proteomics platforms. Many of these signals demonstrate the extensive allelic heterogeneity and ancestry-differentiated variant-trait associations common for inflammation biomarkers, a characteristic we hypothesize will be increasingly observed with well-powered, large-scale analyses of complex traits.

Rare variants in long non-coding RNAs are associated with blood lipid levels in the TOPMed whole-genome sequencing study.

Long non-coding RNAs (lncRNAs) are known to perform important regulatory functions in lipid metabolism. Large-scale whole-genome sequencing (WGS) studies and new statistical methods for variant set tests now provide an opportunity to assess more associations between rare variants in lncRNA genes and complex traits across the genome. In this study, we used high-coverage WGS from 66,329 participants of diverse ancestries with measurement of blood lipids and lipoproteins (LDL-C, HDL-C, TC, and TG) in the National Heart, Lung, and Blood Institute (NHLBI) Trans-Omics for Precision Medicine (TOPMed) program to investigate the role of lncRNAs in lipid variability. We aggregated rare variants for 165,375 lncRNA genes based on their genomic locations and conducted rare-variant aggregate association tests using the STAAR (variant-set test for association using annotation information) framework. We performed STAAR conditional analysis adjusting for common variants in known lipid GWAS loci and rare-coding variants in nearby protein-coding genes. Our analyses revealed 83 rare lncRNA variant sets significantly associated with blood lipid levels, all of which were located in known lipid GWAS loci (in a ±500-kb window of a Global Lipids Genetics Consortium index variant). Notably, 61 out of 83 signals (73%) were conditionally independent of common regulatory variation and rare protein-coding variation at the same loci. We replicated 34 out of 61 (56%) conditionally independent associations using the independent UK Biobank WGS data. Our results expand the genetic architecture of blood lipids to rare variants in lncRNAs.

A multi-dimensional integrative scoring framework for predicting functional variants in the human genome.

Attempts to identify and prioritize functional DNA elements in coding and non-coding regions, particularly through use of in silico functional annotation data, continue to increase in popularity. However, specific functional roles can vary widely from one variant to another, making it challenging to summarize different aspects of variant function with a one-dimensional rating. Here we propose multi-dimensional annotation-class integrative estimation (MACIE), an unsupervised multivariate mixed-model framework capable of integrating annotations of diverse origin to assess multi-dimensional functional roles for both coding and non-coding variants. Unlike existing one-dimensional scoring methods, MACIE views variant functionality as a composite attribute encompassing multiple characteristics and estimates the joint posterior functional probabilities of each genomic position. This estimate offers more comprehensive and interpretable information in the presence of multiple aspects of functionality. Applied to a variety of independent coding and non-coding datasets, MACIE demonstrates powerful and robust performance in discriminating between functional and non-functional variants. We also show an application of MACIE to fine-mapping and heritability enrichment analysis by using the lipids GWAS summary statistics data from the European Network for Genetic and Genomic Epidemiology Consortium.

A framework for detecting noncoding rare-variant associations of large-scale whole-genome sequencing studies.

Large-scale whole-genome sequencing studies have enabled analysis of noncoding rare-variant (RV) associations with complex human diseases and traits. Variant-set analysis is a powerful approach to study RV association. However, existing methods have limited ability in analyzing the noncoding genome. We propose a computationally efficient and robust noncoding RV association detection framework, STAARpipeline, to automatically annotate a whole-genome sequencing study and perform flexible noncoding RV association analysis, including gene-centric analysis and fixed window-based and dynamic window-based non-gene-centric analysis by incorporating variant functional annotations. In gene-centric analysis, STAARpipeline uses STAAR to group noncoding variants based on functional categories of genes and incorporate multiple functional annotations. In non-gene-centric analysis, STAARpipeline uses SCANG-STAAR to incorporate dynamic window sizes and multiple functional annotations. We apply STAARpipeline to identify noncoding RV sets associated with four lipid traits in 21,015 discovery samples from the Trans-Omics for Precision Medicine (TOPMed) program and replicate several of them in an additional 9,123 TOPMed samples. We also analyze five non-lipid TOPMed traits.

Cortical thickness reveals sex differences in verbal and visuospatial memory.

Although previous studies have reported the sex differences in behavior/cognition and the brain, the sex difference in the relationship between memory abilities and the underlying neural basis in the aging process remains unclear. In this study, we used a machine learning model to estimate the association between cortical thickness and verbal/visuospatial memory in females and males and then explored the sex difference of these associations based on a community-elderly cohort (n = 1153, age ranged from 50.42 to 86.67 years). We validated that females outperformed males in verbal memory, while males outperformed females in visuospatial memory. The key regions related to verbal memory in females include the medial temporal cortex, orbitofrontal cortex, and some regions around the insula. Further, those regions are more located in limbic, dorsal attention, and default-model networks, and are associated with face recognition and perception. The key regions related to visuospatial memory include the lateral prefrontal cortex, anterior cingulate gyrus, and some occipital regions. They overlapped more with dorsal attention, frontoparietal and visual networks, and were associated with object recognition. These findings imply the memory performance advantage of females and males might be related to the different memory processing tendencies and their associated network.

FAVOR: functional annotation of variants online resource and annotator for variation across the human genome.

Large biobank-scale whole genome sequencing (WGS) studies are rapidly identifying a multitude of coding and non-coding variants. They provide an unprecedented resource for illuminating the genetic basis of human diseases. Variant functional annotations play a critical role in WGS analysis, result interpretation, and prioritization of disease- or trait-associated causal variants. Existing functional annotation databases have limited scope to perform online queries and functionally annotate the genotype data of large biobank-scale WGS studies. We develop the Functional Annotation of Variants Online Resources (FAVOR) to meet these pressing needs. FAVOR provides a comprehensive multi-faceted variant functional annotation online portal that summarizes and visualizes findings of all possible nine billion single nucleotide variants (SNVs) across the genome. It allows for rapid variant-, gene- and region-level queries of variant functional annotations. FAVOR integrates variant functional information from multiple sources to describe the functional characteristics of variants and facilitates prioritizing plausible causal variants influencing human phenotypes. Furthermore, we provide a scalable annotation tool, FAVORannotator, to functionally annotate large-scale WGS studies and efficiently store the genotype and their variant functional annotation data in a single file using the annotated Genomic Data Structure (aGDS) format, making downstream analysis more convenient. FAVOR and FAVORannotator are available at https://favor.genohub.org.

Sex pheromone communication in an insect parasitoid, Campoletis chlorideae Uchida.

Sex pheromones are pivotal for insect reproduction. However, the mechanism of sex pheromone communication remains enigmatic in hymenopteran parasitoids. Here we have identified the sex pheromone and elucidated the olfactory basis of sex pheromone communication in Campoletis chlorideae (Ichneumonidae), a solitary larval endoparasitoid of over 30 lepidopteran pests. Using coupled gas chromatography-electroantennogram detection, we identified two female-derived pheromone components, tetradecanal (14:Ald) and 2-heptadecanone (2-Hep) (1:4.6), eliciting strong antennal responses from males but weak responses from females. We observed that males but not females were attracted to both single components and the blend. The hexane-washed female cadavers failed to arouse males, and replenishing 14:Ald and 2-Hep could partially restore the sexual attraction of males. We further expressed six C. chlorideae male-biased odorant receptors in Drosophila T1 neurons and found that CchlOR18 and CchlOR47 were selectively tuned to 14:Ald and 2-Hep, respectively. To verify the biological significance of this data, we knocked down CchlOR18 and CchlOR47 individually or together in vivo and show that the attraction of C. chlorideae to their respective ligands was abolished. Moreover, the parasitoids defective in either of the receptors were less likely to court and copulate. Finally, we show that the sex pheromone and (Z)-jasmone, a potent female attractant, can synergistically affect behaviors of virgin males and virgin females and ultimately increase the parasitic efficiency of C. chlorideae. Our study provides new insights into the molecular mechanism of sex pheromone communication in C. chlorideae that may permit manipulation of parasitoid behavior for pest control.

Non-bone-derived exosomes: a new perspective on regulators of bone homeostasis.

Accumulating evidence indicates that exosomes help to regulate bone homeostasis. The roles of bone-derived exosomes have been well-described; however, recent studies have shown that some non-bone-derived exosomes have better bone targeting ability than bone-derived exosomes and that their performance as a drug delivery vehicle for regulating bone homeostasis may be better than that of bone-derived exosomes, and the sources of non-bone-derived exosomes are more extensive and can thus be better for clinical needs. Here, we sort non-bone-derived exosomes and describe their composition and biogenesis. Their roles and specific mechanisms in bone homeostasis and bone-related diseases are also discussed. Furthermore, we reveal obstacles to current research and future challenges in the practical application of exosomes, and we provide potential strategies for more effective application of exosomes for the regulation of bone homeostasis and the treatment of bone-related diseases. Video Abstract.