The β-catenin destruction complex (BDC) regulates WNT-β-catenin signaling and is a prime therapeutic target in colorectal cancer, yet its biochemical complexity has hindered mechanistic understanding. We mapped the seque...The β-catenin destruction complex (BDC) regulates WNT-β-catenin signaling and is a prime therapeutic target in colorectal cancer, yet its biochemical complexity has hindered mechanistic understanding. We mapped the sequence-function landscape of the BDC using tiled base editor screens across its components CTNNB1, AXIN1, APC and GSK3B. Amongst ~150 previously unreported mutations that affected WNT signaling, we discovered gain-of-function and separation-of-function alleles that reveal mechanisms of complex assembly, including a β-catenin region regulating TCF/LEF transcription factor binding. Critically, we found that the AXIN1-β-catenin interface controls signaling flux through the oncogenic BDC found in APC-mutant cancers. In cells expressing truncated APC, β-catenin itself scaffolds BDC assembly, establishing a substrate-assisted autoregulatory mechanism. This architecture represents an unexploited therapeutic vulnerability: strengthening the AXIN1-β-catenin interaction restores destruction complex function and impairs the growth of colorectal cancer cells. Our mutational resource provides a foundation for mechanistic understanding and therapeutic targeting of the WNT pathway.
Han X, Ren K, Wang P
… +13 more, Bi H, Li E, Aydemir I, Ji A, Cai W, Soleimanisardoo L, Wai CM, Schipma MJ, Liu Y, Goldstein J, Sukhanova M, Yang J, Ji P
Erythroid cells require specialized microenvironments called erythroblastic islands (EBIs), niches comprising a central macrophage surrounded by developing erythroid precursors, to complete their maturation. Understandin...Erythroid cells require specialized microenvironments called erythroblastic islands (EBIs), niches comprising a central macrophage surrounded by developing erythroid precursors, to complete their maturation. Understanding EBI composition and function has been limited by two-dimensional in vitro models and the unclear composition of EBIs in human hematopoietic tissues. Using spatial transcriptomic mapping in mouse and human hematopoietic tissues during development and under stress conditions, we show that EBI architecture is unexpectedly species-specific. In mice, C1q-expressing macrophages serve as a hallmark of EBI central macrophages and mediate clearance of ejected erythroid nuclei. In humans, however, EBIs are characterized by macrophage-independent erythroid clusters in fetal liver and bone marrow, whose integrity depends critically on the adhesion molecule ICAM4. These human erythroid clusters are disrupted in myeloid diseases but can be restored with therapy. These findings redefine conventional models of erythroid niche biology and establish a framework for understanding niche dynamics across species.
Heritability enrichment analysis using data from genome-wide association studies is often used to understand the functional basis of genetic architecture. Stratified linkage disequilibrium score regression (S-LDSC) is a...Heritability enrichment analysis using data from genome-wide association studies is often used to understand the functional basis of genetic architecture. Stratified linkage disequilibrium score regression (S-LDSC) is a widely used method-of-moments estimator for heritability enrichment, but S-LDSC has low statistical power compared with likelihood-based approaches. We introduce graphREML, a precise and powerful likelihood-based heritability partitioning and enrichment analysis method. It utilizes summary statistics from genome-wide association studies and sparse linkage disequilibrium graphical models, which make likelihood calculations tractable. We validate our method using extensive simulations and in analyses of a wide range of real traits. On average across traits, graphREML produces enrichment estimates that are concordant with S-LDSC, indicating that both methods are unbiased; however, graphREML identifies 2.5 times more significant trait-annotation enrichments, demonstrating greater power compared with the moment-based S-LDSC approach. Furthermore, graphREML flexibly models the relationship between the annotations of an SNP and its heritability, producing well-calibrated estimates of per-SNP heritability.
The mechanisms by which mutations of splicing factor gene U2AF1 contribute to lung adenocarcinoma pathogenesis are not well understood. Here we used prime editing to modify the endogenous U2AF1 gene in lung adenocarcinom...The mechanisms by which mutations of splicing factor gene U2AF1 contribute to lung adenocarcinoma pathogenesis are not well understood. Here we used prime editing to modify the endogenous U2AF1 gene in lung adenocarcinoma cells and assessed the impact on alternative splicing. One specific KRAS mutation, G12S, led to skipping of KRAS exon 2 and generation of a nonfunctional KRAS transcript. However, expression of the U2AF1 mutant reverted this exon skipping and restored KRAS function, leading to enrichment of U2AF1 mutations in KRAS-mutant lung adenocarcinomas. Comprehensive analysis of splicing factor-oncogene mutation co-occurrence in cancer genomes also revealed significant coenrichment of KRAS and U2AF1 mutations. Experimentally, KRAS mutation led to KRAS exon 3 skipping, which in turn could be rescued by expression of U2AF1. Our findings provide evidence that splicing factor mutations can rescue splicing defects caused by oncogenic mutations in a dynamic process of cascading selection.
The developmental history of a cell fundamentally defines its identity and function. Recent advances in cell lineage tracing now enable high-resolution reconstruction of cellular ancestries in vivo, illuminating how line...The developmental history of a cell fundamentally defines its identity and function. Recent advances in cell lineage tracing now enable high-resolution reconstruction of cellular ancestries in vivo, illuminating how lineage dictates fate in health and disease. This Review highlights recently developed tools, ranging from refined recombinase systems to advanced synthetic and natural barcoding approaches, that facilitate the investigation of pathological lineage programs in cancer, cardiovascular disease and aging. Moving forward, integrating permanent lineage records with single-cell multi-omics promises to provide a unified framework to decode how a cell's past shapes its present state and future potential, heralding a new era for precision medicine.
Jin M, Yan S, Wu Y
… +27 more, Zhai Z, Alseekh S, Chen Y, Huang W, Ma N, Tao K, Xiao K, Zhu Y, Yu Y, Shao Y, Jiang C, Liu X, Sun J, Qu Y, Wei W, Li W, Xu J, Luo J, Wang X, Zhuo L, Zhan J, Qiu F, Yang N, Xiao Y, Liu HJ, Fernie AR, Yan J
Maize primary metabolism drives complex agronomic traits, yet its genetic regulation remains difficult to resolve. Here we integrated genomic, transcriptomic and metabolomic data from 1,404 maize progenies derived from 2...Maize primary metabolism drives complex agronomic traits, yet its genetic regulation remains difficult to resolve. Here we integrated genomic, transcriptomic and metabolomic data from 1,404 maize progenies derived from 24 diverse founders to dissect the genetic architecture of primary metabolism. We constructed a high-confidence regulatory network that resolved causal genes underlying metabolic quantitative trait loci and successfully identified targets for improving maize nutritional quality. This systems-level framework further prioritized ZmAVT1A-1, encoding a putative amino acid transporter, as a key regulator of amino acid accumulation. Natural variation and transgenic analyses showed that ZmAVT1A-1 modulates nitrogen partitioning between vegetative tissues and kernels, revealing pleiotropic effects on agronomic traits. These findings illustrated the intricate trade-offs inherent in metabolic regulation. Together, our study provides a comprehensive multiomics resource for decoding metabolic networks and underscores the necessity of a systems approach to navigate the pleiotropic nature of crop improvement targets.
The theory of immunosurveillance posits that T cells eliminate clones harboring nonself-antigens generated by somatic mutations. Although a role of immunosurveillance in cancer is supported by evidence, whether it affect...The theory of immunosurveillance posits that T cells eliminate clones harboring nonself-antigens generated by somatic mutations. Although a role of immunosurveillance in cancer is supported by evidence, whether it affects precancerous expansions has not been well established. Here we studied the association between MHC-variant binding and risk of clonal hematopoiesis (CH), a blood precancer state, predicting MHC binding affinity toward CH hotspot variants in 380,000 UK Biobank participants and examining the relationship between predicted binding to each variant and its expansion risk. Despite the study being powered to detect subtle differences in selective pressure, we did not find associations between predicted binding and CH prevalence for any of the examined variants. In CH-affected individuals, we identified no relationship between predicted variant binding and clone size. Overall, we found no evidence that the MHC genotype affects which variants expand in CH, suggesting a limited role for immunosurveillance in shaping clonal expansions in the blood.
Medical genetics currently operates through a fragmented diagnostic cascade built around short-read sequencing technologies that carry well-documented blind spots, including regions of high sequence homology, tandem repe...Medical genetics currently operates through a fragmented diagnostic cascade built around short-read sequencing technologies that carry well-documented blind spots, including regions of high sequence homology, tandem repeats and segmental duplications, as well as large or complex structural variants, invisible base modifications and a lack of variant phasing. We propose that long-read genome sequencing should be considered as one pillar of a broader technological convergence encompassing diploid genome assembly, pangenome references and artificial intelligence-driven variant interpretation, termed near-perfect genome sequencing (NPGS). We further propose a Bayesian framework in which genomic completeness itself constitutes interpretive evidence for variant classification. This principle has direct implications for the interpretation of variants of uncertain significance in clinical practice. We highlight the potential of NPGS across postnatal, prenatal and oncological settings and outline a staged implementation roadmap toward the one-test paradigm. We also address real-world implementation challenges, including cost, computational demand, equity and ethical considerations.
RNA alternative splicing is a fundamental post-transcriptional mechanism whose dysregulation drives various human diseases. Predicting splicing outcomes is therefore a central challenge in precision medicine. This Review...RNA alternative splicing is a fundamental post-transcriptional mechanism whose dysregulation drives various human diseases. Predicting splicing outcomes is therefore a central challenge in precision medicine. This Review traces the evolution of computational approaches from early statistical heuristics to modern artificial intelligence frameworks. We dissect the methodologies that shape predictive performance, including training data scale, output resolution, splicing event quantification and model complexity. Among these factors, quantitative assessment of splicing events and the increasing complexity of model architectures represent particularly critical axes that define both biological interpretability and computational feasibility. We further describe how these models empower translational applications, from annotating variant effects to guiding antisense oligonucleotide development. Nonetheless, persistent challenges remain, including the interpretation of deep-intronic mutations, isoform-level reconstruction and integration of multimodal data. Together, these perspectives define both the progress achieved and the opportunities ahead for splicing prediction in genomics and medicine.
Most genetic variants associated with complex heritability phenotypes lie in non-coding regions and are thought to influence disease risk by regulating gene expression. However, most transcriptome-wide association approa...Most genetic variants associated with complex heritability phenotypes lie in non-coding regions and are thought to influence disease risk by regulating gene expression. However, most transcriptome-wide association approaches primarily model local (cis) genetic effects, leaving much of gene regulation unexplained. Here, we show that incorporating distal (trans) regulatory effects improves the prediction of gene expression and the identification of disease-associated genes. Using RNA sequencing data from six human post-mortem brain regions, we developed INGENE and MODULE, two models capturing the combined influence of candidate trans-acting variants within gene coexpression networks. Integrating these models with conventional cis-based predictors improved gene expression imputation (maximum likelihood estimation, α = 0.05) for 18,744 genes across regions. Applying this framework to Psychiatric Genomics Consortium wave 3 genotypes identified 766 genes associated with schizophrenia (P < 0.01), including 641 not previously reported by transcriptome-wide analyses. These findings highlight the contribution of distal regulatory mechanisms and gene network interactions to schizophrenia risk.
Nomura M, Raviram R, Schiffman JS
… +19 more, Bussema L, Lu V, Wheeler N, Lee JJY, Fan Y, Zheng MH, Ruiz F, Danish H, Kellett S, Nusrat L, Chaligne R, Huse JT, Yung WKA, Tanaka S, Saito N, Das S, Potenski C, Landau DA, Suvà ML
The progression of isocitrate dehydrogenase-mutant glioma (IDH-G) from slow-growing tumor to fatal disease is associated with transcriptional and DNA methylation changes that remain poorly understood. Here, we profiled a...The progression of isocitrate dehydrogenase-mutant glioma (IDH-G) from slow-growing tumor to fatal disease is associated with transcriptional and DNA methylation changes that remain poorly understood. Here, we profiled a longitudinal cohort of 36 IDH-G samples from 19 patients by joint-capture multi-omic single-nucleus DNA methylation, single-nucleus RNA sequencing and bulk exome sequencing. We show that IDH-G progression is associated with an increase in malignant stem-like states, decreased differentiation and methylation loss, which marks tumors with worse clinical outcome. Methylation loss was uniformly observed across malignant cells within individual tumors, suggesting that it may underlie rather than result from the increase in stem-like states. Analysis of cell-state heritability and plasticity using high-resolution phylogenetic trees links DNA methylation loss to alterations in glioma cell-state encoding and heritability. Our study offers insights into how DNA methylation loss reshapes cellular transitions and how it may mark clinically more aggressive tumors across IDH-G subsets.
Marderstein AR, Kundu S, Padhi EM
… +16 more, Deshpande S, Wang A, Robb E, Sun Y, Yun CM, Pomales-Matos D, Xie Y, Chang SH, Chin IM, Shah AJ, Gardell ZA, Corces MR, Nachun D, Jessa S, Kundaje A, Montgomery SB
Interpreting how noncoding variants act in specific cell types across human development is a major challenge. Here we generated 3 billion predictions from deep learning sequence models of chromatin accessibility across d...Interpreting how noncoding variants act in specific cell types across human development is a major challenge. Here we generated 3 billion predictions from deep learning sequence models of chromatin accessibility across diverse fetal and adult cellular contexts. These prioritized functional variants and revealed a dichotomy: common variants are more cell-type-specific, whereas ultra-rare variants had larger and broader effects across cell types, with the strongest evidence of purifying selection in fetal neurons. Leveraging these insights, we developed FLARE (Functional Lasso Analysis of Regulatory Evolution), which integrates evolutionary constraint to prioritize noncoding variants with extreme regulatory effects. FLARE provided a general framework for studying regulatory variation, from de novo mutations in childhood disorders to rare variants underlying outlier adult brain expression and common variants enriched for schizophrenia heritability. Together, these results demonstrate how integrating single-cell chromatin accessibility, population genetics and deep learning can identify regulatory variants that influence human development and disease.
Krzak M, Alegbe T, Taylor DL
… +35 more, Jones GR, Ghouraba M, Strickland M, Harris BT, Satti R, Arestang K, Ramirez-Navarro L, Nishad N, Cheam KAX, Tutert M, Ozols M, Noell G, Leonard S, Przybilla MJ, Suastegui CR, Khabirova E, Deng T, Najgebauer H, Petrova V, Jones CP, Wana N, Hu MX, Skelton J, Ostermayer J, Gu Y, Garri W, Brezina B, Caballes CQ, Corridoni D, Parkes M, Iyer V, Martin CC, McIntyre RE, Raine T, Anderson CA
Crohn's disease (CD) is a chronic inflammatory bowel disease exhibiting substantial heterogeneity in clinical presentation and response to therapy. To explore its molecular basis, we developed IBDverse, a large single-ce...Crohn's disease (CD) is a chronic inflammatory bowel disease exhibiting substantial heterogeneity in clinical presentation and response to therapy. To explore its molecular basis, we developed IBDverse, a large single-cell RNA sequencing (scRNA-seq) dataset of terminal ileal biopsies, profiling over 1.1 million cells from 111 patients with CD and 232 healthy controls. This resource integrates discovery and replication cohorts for the robust identification of CD-associated cell types, genes and pathways. We uncovered epithelial changes marked by interferon-driven upregulation of major histocompatibility complex class I molecules that persisted in progenitor cells after macroscopic inflammation resolution. ITGA4 macrophages were identified as key inflammatory drivers, showing enriched JAK-STAT signaling and cytokine expression (interleukin-6 (IL-6), IL-12 and IL-23). Heritability analysis linked inflammatory monocytes and macrophages to CD susceptibility, implicating resident and recruited immune cells in pathogenesis. These findings establish a comprehensive cellular and molecular framework for CD, offering insights into disease mechanisms and therapeutic opportunities.
Fujii M, Abeto N, Kishimoto S
… +13 more, Shiraishi K, Hashimoto T, Hiraoka N, Nonaka S, Kojima M, Matano M, Takahashi S, Colozza G, Koo BK, Yatabe Y, Kato M, Sato T, Sekine S
The majority of intestinal tumors harbor mutations in canonical Wnt pathway genes such as APC, whereas the lack of such alterations in a subset of tumors implies alternative tumorigenic routes. Here we identify recurrent...The majority of intestinal tumors harbor mutations in canonical Wnt pathway genes such as APC, whereas the lack of such alterations in a subset of tumors implies alternative tumorigenic routes. Here we identify recurrent in-frame deletion in COPA, frequently co-occurring with USP9X-truncating mutation, in small intestinal adenoma and adenocarcinoma. Patient-derived and CRISPR-engineered small intestinal organoids carrying COPA in-frame deletions exhibit R-spondin-independent yet Wnt ligand-dependent growth, maintaining LGR5 expression without canonical Wnt drivers. Mechanistically, COPA mutation stabilizes the Frizzled coreceptor LRP6 irrespective of R-spondin, sustaining Wnt pathway activation under growth factor-restricted conditions. USP9X loss further potentiates this phenotype. Unlike canonical Wnt pathway members, COPA encodes the α-subunit of coatomer complex I, which engages in vesicle trafficking with little prior linkage to intestinal tumorigenesis. Our findings establish COPA mutation as a unique and atypical intestinal tumor driver and implicate USP9X loss as a cooperating lesion.