Alcohol is the most consumed and abused psychoactive drug globally, but the molecular mechanisms driving alcohol action and its associated behaviors in the brain remain enigmatic. Here, we have discovered a transmembrane protein TMEM132B that is a GABAA receptor (GABAAR) auxiliary subunit. Functionally, TMEM132B promotes GABAAR expression at the cell surface, slows receptor deactivation, and enhances the allosteric effects of alcohol on the receptor. In TMEM132B knockout (KO) mice or TMEM132B I499A knockin (KI) mice in which the TMEM132B-GABAAR interaction is specifically abolished, GABAergic transmission is decreased and alcohol-induced potentiation of GABAAR-mediated currents is diminished in hippocampal neurons. Behaviorally, the anxiolytic and sedative/hypnotic effects of alcohol are markedly reduced, and compulsive, binge-like alcohol consumption is significantly increased. Taken together, these data reveal a GABAAR auxiliary subunit, identify the TMEM132B-GABAAR complex as a major alcohol target in the brain, and provide mechanistic insights into alcohol-related behaviors.

Pyrenoids are subcompartments of algal chloroplasts that increase the efficiency of Rubisco-driven CO2 fixation. Diatoms fix up to 20% of global CO2, but their pyrenoids remain poorly characterized. Here, we used in vivo photo-crosslinking to identify pyrenoid shell (PyShell) proteins, which we localized to the pyrenoid periphery of model pennate and centric diatoms, Phaeodactylum tricornutum and Thalassiosira pseudonana. In situ cryo-electron tomography revealed that pyrenoids of both diatom species are encased in a lattice-like protein sheath. Single-particle cryo-EM yielded a 2.4-Å-resolution structure of an in vitro TpPyShell1 lattice, which showed how protein subunits interlock. T. pseudonana TpPyShell1/2 knockout mutants had no PyShell sheath, altered pyrenoid morphology, and a high-CO2 requiring phenotype, with reduced photosynthetic efficiency and impaired growth under standard atmospheric conditions. The structure and function of the diatom PyShell provide a molecular view of how CO2 is assimilated in the ocean, a critical ecosystem undergoing rapid change.

The SWR1 chromatin remodeling complex is recruited to +1 nucleosomes downstream of transcription start sites of eukaryotic promoters, where it exchanges histone H2A for the specialized variant H2A.Z. Here, we use cryoelectron microscopy (cryo-EM) to resolve the structural basis of the SWR1 interaction with free DNA, revealing a distinct open conformation of the Swr1 ATPase that enables sliding from accessible DNA to nucleosomes. A complete structural model of the SWR1-nucleosome complex illustrates critical roles for Swc2 and Swc3 subunits in oriented nucleosome engagement by SWR1. Moreover, an extended DNA-binding α helix within the Swc3 subunit enables sensing of nucleosome linker length and is essential for SWR1-promoter-specific recruitment and activity. The previously unresolved N-SWR1 subcomplex forms a flexible extended structure, enabling multivalent recognition of acetylated histone tails by reader domains to further direct SWR1 toward the +1 nucleosome. Altogether, our findings provide a generalizable mechanism for promoter-specific targeting of chromatin and transcription complexes.

Widespread sequencing has yielded thousands of missense variants predicted or confirmed as disease causing. This creates a new bottleneck: determining the functional impact of each variant-typically a painstaking, customized process undertaken one or a few genes and variants at a time. Here, we established a high-throughput imaging platform to assay the impact of coding variation on protein localization, evaluating 3,448 missense variants of over 1,000 genes and phenotypes. We discovered that mislocalization is a common consequence of coding variation, affecting about one-sixth of all pathogenic missense variants, all cellular compartments, and recessive and dominant disorders alike. Mislocalization is primarily driven by effects on protein stability and membrane insertion rather than disruptions of trafficking signals or specific interactions. Furthermore, mislocalization patterns help explain pleiotropy and disease severity and provide insights on variants of uncertain significance. Our publicly available resource extends our understanding of coding variation in human diseases.

Complex structural variations (cxSVs) are often overlooked in genome analyses due to detection challenges. We developed ARC-SV, a probabilistic and machine-learning-based method that enables accurate detection and reconstruction of cxSVs from standard datasets. By applying ARC-SV across 4,262 genomes representing all continental populations, we identified cxSVs as a significant source of natural human genetic variation. Rare cxSVs have a propensity to occur in neural genes and loci that underwent rapid human-specific evolution, including those regulating corticogenesis. By performing single-nucleus multiomics in postmortem brains, we discovered cxSVs associated with differential gene expression and chromatin accessibility across various brain regions and cell types. Additionally, cxSVs detected in brains of psychiatric cases are enriched for linkage with psychiatric GWAS risk alleles detected in the same brains. Furthermore, our analysis revealed significantly decreased brain-region- and cell-type-specific expression of cxSV genes, specifically for psychiatric cases, implicating cxSVs in the molecular etiology of major neuropsychiatric disorders.

The capability to spatially explore RNA biology in formalin-fixed paraffin-embedded (FFPE) tissues holds transformative potential for histopathology research. Here, we present pathology-compatible deterministic barcoding in tissue (Patho-DBiT) by combining in situ polyadenylation and computational innovation for spatial whole transcriptome sequencing, tailored to probe the diverse RNA species in clinically archived FFPE samples. It permits spatial co-profiling of gene expression and RNA processing, unveiling region-specific splicing isoforms, and high-sensitivity transcriptomic mapping of clinical tumor FFPE tissues stored for 5 years. Furthermore, genome-wide single-nucleotide RNA variants can be captured to distinguish malignant subclones from non-malignant cells in human lymphomas. Patho-DBiT also maps microRNA regulatory networks and RNA splicing dynamics, decoding their roles in spatial tumorigenesis. Single-cell level Patho-DBiT dissects the spatiotemporal cellular dynamics driving tumor clonal architecture and progression. Patho-DBiT stands poised as a valuable platform to unravel rich RNA biology in FFPE tissues to aid in clinical pathology evaluation.

In this high-throughput proteomic study of autosomal dominant Alzheimer's disease (ADAD), we sought to identify early biomarkers in cerebrospinal fluid (CSF) for disease monitoring and treatment strategies. We examined CSF proteins in 286 mutation carriers (MCs) and 177 non-carriers (NCs). The developed multi-layer regression model distinguished proteins with different pseudo-trajectories between these groups. We validated our findings with independent ADAD as well as sporadic AD datasets and employed machine learning to develop and validate predictive models. Our study identified 137 proteins with distinct trajectories between MCs and NCs, including eight that changed before traditional AD biomarkers. These proteins are grouped into three stages: early stage (stress response, glutamate metabolism, neuron mitochondrial damage), middle stage (neuronal death, apoptosis), and late presymptomatic stage (microglial changes, cell communication). The predictive model revealed a six-protein subset that more effectively differentiated MCs from NCs, compared with conventional biomarkers.

Phage therapy is gaining increasing interest in the fight against critically antibiotic-resistant nosocomial pathogens. However, the narrow host range of bacteriophages hampers the development of broadly effective phage therapeutics and demands precision approaches. Here, we combine large-scale phylogeographic analysis with high-throughput phage typing to guide the development of precision phage cocktails targeting carbapenem-resistant Acinetobacter baumannii, a top-priority pathogen. Our analysis reveals that a few strain types dominate infections in each world region, with their geographical distribution remaining stable within 6 years. As we demonstrate in Eastern Europe, this spatiotemporal distribution enables preemptive preparation of region-specific phage collections that target most local infections. Finally, we showcase the efficacy of phage cocktails against prevalent strain types using in vitro and animal infection models. Ultimately, genomic surveillance identifies patients benefiting from the same phages across geographical scales, thus providing a scalable framework for precision phage therapy.

Many mammals can temporally uncouple conception from parturition by pacing down their development around the blastocyst stage. In mice, this dormant state is achieved by decreasing the activity of the growth-regulating mTOR signaling pathway. It is unknown whether this ability is conserved in mammals in general and in humans in particular. Here, we show that decreasing the activity of the mTOR signaling pathway induces human pluripotent stem cells (hPSCs) and blastoids to enter a dormant state with limited proliferation, developmental progression, and capacity to attach to endometrial cells. These in vitro assays show that, similar to other species, the ability to enter dormancy is active in human cells around the blastocyst stage and is reversible at both functional and molecular levels. The pacing of human blastocyst development has potential implications for reproductive therapies.

Despite the long history of consumption of fermented dairy, little is known about how the fermented microbes were utilized and evolved over human history. Here, by retrieving ancient DNA of Bronze Age kefir cheese (∼3,500 years ago) from the Xiaohe cemetery, we explored past human-microbial interactions. Although it was previously suggested that kefir was spread from the Northern Caucasus to Europe and other regions, we found an additional spreading route of kefir from Xinjiang to inland East Asia. Over evolutionary history, the East Asian strains gained multiple gene clusters with defensive roles against environmental stressors, which can be a result of the adaptation of Lactobacillus strains to various environmental niches and human selection. Overall, our results highlight the role of past human activities in shaping the evolution of human-related microbes, and such insights can, in turn, provide a better understanding of past human behaviors.

We report the 1-year results from one patient as the preliminary analysis of a first-in-human phase I clinical trial (ChiCTR2300072200) assessing the feasibility of autologous transplantation of chemically induced pluripotent stem-cell-derived islets (CiPSC islets) beneath the abdominal anterior rectus sheath for type 1 diabetes treatment. The patient achieved sustained insulin independence starting 75 days post-transplantation. The patient's time-in-target glycemic range increased from a baseline value of 43.18% to 96.21% by month 4 post-transplantation, accompanied by a decrease in glycated hemoglobin, an indicator of long-term systemic glucose levels at a non-diabetic level. Thereafter, the patient presented a state of stable glycemic control, with time-in-target glycemic range at >98% and glycated hemoglobin at around 5%. At 1 year, the clinical data met all study endpoints with no indication of transplant-related abnormalities. Promising results from this patient suggest that further clinical studies assessing CiPSC-islet transplantation in type 1 diabetes are warranted.

Interpretation of disease-causing genetic variants remains a challenge in human genetics. Current costs and complexity of deep mutational scanning methods are obstacles for achieving genome-wide resolution of variants in disease-related genes. Our framework, saturation mutagenesis-reinforced functional assays (SMuRF), offers simple and cost-effective saturation mutagenesis paired with streamlined functional assays to enhance the interpretation of unresolved variants. Applying SMuRF to neuromuscular disease genes FKRP and LARGE1, we generated functional scores for all possible coding single-nucleotide variants, which aid in resolving clinically reported variants of uncertain significance. SMuRF also demonstrates utility in predicting disease severity, resolving critical structural regions, and providing training datasets for the development of computational predictors. Overall, our approach enables variant-to-function insights for disease genes in a cost-effective manner that can be broadly implemented by standard research laboratories.

Long COVID, a type of post-acute sequelae of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) (PASC) defined by medically unexplained symptoms following infection with SARS-CoV-2, is a newly recognized infection-associated chronic condition that causes disability in some people. Substantial progress has been made in defining its epidemiology, biology, and pathophysiology. However, there is no cure for the tens of millions of people believed to be experiencing long COVID, and industry engagement in developing therapeutics has been limited. Here, we review the current state of knowledge regarding the biology and pathophysiology of long COVID, focusing on how the proposed mechanisms explain the physiology of the syndrome and how they provide a rationale for the implementation of a broad experimental medicine and clinical trials agenda. Progress toward preventing and curing long COVID and other infection-associated chronic conditions will require deep and sustained investment by funders and industry.

Pharmaceuticals can directly inhibit the growth of gut bacteria, but the degree to which such interactions manifest in complex community settings is an open question. Here, we compared the effects of 30 drugs on a 32-species synthetic community with their effects on each community member in isolation. While most individual drug-species interactions remained the same in the community context, communal behaviors emerged in 26% of all tested cases. Cross-protection during which drug-sensitive species were protected in community was 6 times more frequent than cross-sensitization, the converse phenomenon. Cross-protection decreased and cross-sensitization increased at higher drug concentrations, suggesting that the resilience of microbial communities can collapse when perturbations get stronger. By metabolically profiling drug-treated communities, we showed that both drug biotransformation and bioaccumulation contribute mechanistically to communal protection. As a proof of principle, we molecularly dissected a prominent case: species expressing specific nitroreductases degraded niclosamide, thereby protecting both themselves and sensitive community members.

Diet is a major determinant of gut microbiome composition, and variation in diet-microbiome interactions may contribute to variation in their health consequences. To mechanistically understand these relationships, here we map interactions between ∼150 small-molecule dietary xenobiotics and the gut microbiome, including the impacts of these compounds on community composition, the metabolic activities of human gut microbes on dietary xenobiotics, and interindividual variation in these traits. Microbial metabolism can toxify and detoxify these compounds, producing emergent interactions that explain community-specific remodeling by dietary xenobiotics. We identify the gene and enzyme responsible for detoxification of one such dietary xenobiotic, resveratrol, and demonstrate that this enzyme contributes to interindividual variation in community remodeling by resveratrol. Together, these results systematically map interactions between dietary xenobiotics and the gut microbiome and connect toxification and detoxification to interpersonal differences in microbiome response to diet.

Phytochrome B (phyB) and phytochrome-interacting factors (PIFs) constitute a well-established signaling module critical for plants adapting to ambient light. However, mechanisms underlying phyB photoactivation and PIF binding for signal transduction remain elusive. Here, we report the cryo-electron microscopy (cryo-EM) structures of the photoactivated phyB or the constitutively active phyBY276H mutant in complex with PIF6, revealing a similar trimer. The light-induced configuration switch of the chromophore drives a conformational transition of the nearby tongue signature within the phytochrome-specific (PHY) domain of phyB. The resulting α-helical PHY tongue further disrupts the head-to-tail dimer of phyB in the dark-adapted state. These structural remodelings of phyB facilitate the induced-fit recognition of PIF6, consequently stabilizing the N-terminal extension domain and a head-to-head dimer of activated phyB. Interestingly, the phyB dimer exhibits slight asymmetry, resulting in the binding of only one PIF6 molecule. Overall, our findings solve a key question with respect to how light-induced remodeling of phyB enables PIF signaling in phytochrome research.

Duchenne muscular dystrophy (DMD) is a progressive muscle-wasting disease caused by mutations in the DMD gene. Muscle fibers rely on the coordination of multiple cell types for repair and regenerative capacity. To elucidate the cellular and molecular changes in these cell types under pathologic conditions, we generated a rhesus monkey model for DMD that displays progressive muscle deterioration and impaired motor function, mirroring human conditions. By leveraging these DMD monkeys, we analyzed freshly isolated muscle tissues using single-cell RNA sequencing (scRNA-seq). Our analysis revealed changes in immune cell landscape, a reversion of lineage progressing directions in fibrotic fibro-adipogenic progenitors (FAPs), and TGF-β resistance in FAPs and muscle stem cells (MuSCs). Furthermore, MuSCs displayed cell-intrinsic defects, leading to differentiation deficiencies. Our study provides important insights into the pathogenesis of DMD, offering a valuable model and dataset for further exploration of the underlying mechanisms, and serves as a suitable platform for developing and evaluating therapeutic interventions.

m6A modification is best known for its critical role in controlling multiple post-transcriptional processes of the mRNAs. Here, we discovered elevated levels of m6A modification on centromeric RNA (cenRNA) in cancerous cells compared with non-cancerous cells. We then identified CENPA, an H3 variant, as an m6A reader of cenRNA. CENPA is localized at centromeres and is essential in preserving centromere integrity and function during mitosis. The m6A-modified cenRNA stabilizes centromeric localization of CENPA in cancer cells during the S phase of the cell cycle. Mutations of CENPA at the Leu61 and the Arg63 or removal of cenRNA m6A modification lead to loss of centromere-bound CENPA during S phase. This in turn results in compromised centromere integrity and abnormal chromosome separation and hinders cancer cell proliferation and tumor growth. Our findings unveil an m6A reading mechanism by CENPA that epigenetically governs centromere integrity in cancer cells, providing potential targets for cancer therapy.

Sharing genetic and other study results with the communities who participate in research falls under benefit-sharing and capacity-building initiatives that underpin a more equitable biomedical research relationship. Yet, which results to return and how remain fundamental challenges that persist in the absence of practical guidance and institutional policies. Here, we discuss how the return of results can be implemented across different geographies, study designs, and project budgets.

Eukaryotic cell function and survival rely on the use of a mitochondrial H+ electrochemical gradient (Δp), which is composed of an inner mitochondrial membrane (IMM) potential (ΔΨmt) and a pH gradient (ΔpH). So far, ΔΨmt has been assumed to be composed exclusively of H+. Here, using a rainbow of mitochondrial and nuclear genetic models, we have discovered that a Na+ gradient equates with the H+ gradient and controls half of ΔΨmt in coupled-respiring mammalian mitochondria. This parallelism is controlled by the activity of the long-sought Na+-specific Na+/H+ exchanger (mNHE), which we have identified as the P-module of complex I (CI). Deregulation of this mNHE function, without affecting the canonical enzymatic activity or the assembly of CI, occurs in Leber's hereditary optic neuropathy (LHON), which has profound consequences in ΔΨmt and mitochondrial Ca2+ homeostasis and explains the previously unknown molecular pathogenesis of this neurodegenerative disease.