Gene expression in mammalian cells is controlled by enhancers that are often dispersed across large cis-regulatory landscapes around a promoter. How enhancers determine transcription of their target genes and how this depends on their relative position within a cis-regulatory landscape remain unclear. Here we use live-cell imaging to track the activity of a promoter under the control of the same enhancer, but inserted at different positions across a simplified regulatory landscape with minimal complexity. Combined with mathematical modeling, this reveals that RNA production from the promoter occurs in clusters of transcriptional bursts, with enhancer position controlling the frequency at which clusters appear. This results in bursts being more frequent and occurring more uniformly across cells when the enhancer is genomically close to the promoter than when it is located at a large genomic distance. Mathematical modeling further indicates that the enhancer modulates the promoter's ability to transition from its basal transcriptional state to a regime in which clusters of bursts become more frequent. Our results reveal unexplored modes of mammalian promoter operation and show that enhancer position within a cis-regulatory landscape critically controls the timing and variability of transcriptional output in single cells.

The generalizability of polygenic scores (PGS) remains a major hurdle in the pursuit of equitable genomic medicine. Differences in disease prevalence across groups, potentially including social strata, influence the relationship between PGS and risk. Here we quantified the magnitude of PGS-by-context (PGS×C) interactions for seven human diseases and pairs of 75 contexts in the UK Biobank (n = 408,801). Across 24,198 PGS×C models, 746 (3.1%) had significant interactions by two criteria, up to fourfold more than expected by chance, improving predictive accuracy. The predominant mechanism for PGS×C is the amplification of genetic effects in adverse contexts, such as low polyunsaturated fatty acids or social determinants of ill health. We introduce the notion of the proportion needed to benefit as a metric that quantifies the expected effectiveness of interventions as a function of polygenic risk. Our results highlight the need for more comprehensive sampling across groups experiencing adverse exposures.

Musculoskeletal diseases are a major health burden. Development of bone-active therapies has been hindered by limited understanding of the cells and genes that regulate the skeleton. We exploited the value of cross-species analysis and developed single-cell methodologies in skeletal tissues to define the critical endosteal compartment that regulates bone turnover. Thirty-four distinct cell types were identified, and disease-relevant cells prioritized using enrichment for rare skeletal disorder genes and bone-mineral-density-associated genes in an extended UK Biobank genome-wide association study. Functional validation was undertaken in over 1,000 genetically modified mouse models. Endothelial cells and vascular smooth muscle cells were identified as new skeletal-disease-relevant cells alongside osteoblast, chondrocyte and osteoclast cell lineages. Hundreds of cell-specific genes with unappreciated roles in skeletal pathophysiology were identified. This comprehensive cellular and molecular framework underpins skeletal physiology and disease and will help prioritize new therapeutic targets to accelerate development of therapies to treat musculoskeletal disease.

Mung bean (Vigna radiata) is a globally important legume crop valued for its short growing cycle, nitrogen-fixing capacity and high nutritional value, particularly in developing countries. Here we report a comprehensive graph-based pan-genome assembled from 11 genetically diverse global accessions. The framework captures 75,268 gene families (50.86% core, 35.19% dispensable and 13.95% private) and 66,862 nonredundant structural variants. Integrating these structural variants and single nucleotide polymorphisms, genome-wide association studies across five environments identified candidate genes for 20 agronomic traits, underscoring the pivotal roles of these variants in driving mung bean domestication and improvement. Mechanistically, we demonstrate that a 68-bp promoter insertion in VrTIFY6B and a 136-bp promoter deletion in VrPGIP1 regulate flavonoid content and confer bruchid resistance, respectively. These genomic resources and actionable functional variants provide a powerful toolkit to accelerate mung bean improvement through marker-assisted breeding, genomic selection and genome editing to address global food security.

Phenotypically healthy cells frequently harbor somatic variants at cancer-associated genes, indicating that malignant transformation requires the selection of several alterations. Predicting which combinations of mutations, or co-mutations, exhibit oncogenic capacity requires identifying co-mutations that occur more or less frequently than expected. However, statistical frameworks to solve this problem are hampered by tumor heterogeneity and data availability. Here we curated putative oncogenic mutations in >70,000 human tumors from 119 subtypes, and designed a strategy to search for co-mutations based on in silico simulation of mutagenesis (SelectSim). Using this dataset and tool, we discovered and validated co-mutations across independent human cohorts, compared co-mutations across different tumor types and identified potential risk factors of metastatic progression. Notably, across several cohorts of phenotypically normal tissue samples, we show that, unlike individual oncogenic variants, significantly co-occurring mutations are largely cancer-specific and are observed rarely in healthy tissues, providing clues about the paths to tumorigenesis.

Metabolic dysfunction-associated steatotic liver disease (MASLD), especially its severe form, metabolic dysfunction-associated steatohepatitis (MASH), progresses to liver fibrosis, leading to cirrhosis and liver cancer. The cross-talk between spleen and liver in MASLD/MASH progression remains poorly understood. Here we found enlarged spleens in patients with MASLD and identified induced TRNP1[hi]CD8[+] T cells in spleens of MASLD/MASH mouse models and patients. These cells exhibited pro-fibrotic properties through secretion of INSR-α. Mechanistically, demethylated DNA and H3K27me3, increased H3K27ac and bolstered enhancer-promoter contact synergistically reorganized chromatin topologically associating domains spatially to initiate the expression of transcription factor TRNP1 in splenic CD8[+] T cells. TRNP1 then transcriptionally activated the expression of FURIN and CTSD, promoting the maturation and ectodomain shedding of INSR-α and facilitating its secretion to activate hepatic stellate cells. In vivo blockade of INSR-α using neutralizing antibodies alleviated MASLD/MASH-induced liver fibrosis. This study reveals splenic TRNP1[hi]CD8[+] T cells with pro-fibrotic properties and suggests a potential anti-fibrotic strategy.

Histone post-translational modifications are fundamental to genome regulation, yet dissecting the functions of individual histone marks in mammals remains challenging due to the presence of multiple histone gene copies. Here we develop a high-throughput clustered regularly interspaced short palindromic repeats (CRISPR) prime editing platform enabling precise, reversible and combinatorial mutagenesis of canonical and noncanonical histone H3 genes within their native genomic context. Using systematic lysine-to-arginine substitutions benchmarked against synonymous controls, we identify key residues, including H3K4, H3K9, H3K14, H3K18 and H3K79, whose mutation compromises fitness in mouse embryonic stem cells. We further show that H3K56, linked to genome stability in yeast and Drosophila, has a conserved role in mammalian cells. Through analysis of selected double mutants, we uncover functional crosstalk across residues, with combinations such as H3K27R + H3K36R impairing stem cell self-renewal and altering transcription. Altogether, this study establishes a functional map of histone H3 lysines in mammals and provides a broadly applicable platform for systematic dissection of chromatin regulation.