Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer characterized by profound desmoplasia and cellular heterogeneity, which cannot be fully resolved using traditional bulk sequencing approaches. To understand how this heterogeneity contributes to PDAC biology, we analyzed a large cohort of primary tumors (n = 62) profiled using single-cell RNA sequencing (443,451 single cells), including a subset of 11 tumors with matched spatial transcriptomics (53,236 spots), to map clonal, transcriptional and spatial diversity. Moreover, our sample set includes a unique cohort of matched samples from six tumors collected before and after treatment with chemotherapy. Our analysis revealed notable intratumoral heterogeneity, with multiple genetically distinct neoplastic clones coexisting within individual tumors. We observed clone-specific ligand-receptor programs that are modulated by genomic dosage: amplifications of ligands or receptors enriched corresponding interactions, while deletions suppressed them, with genetically encoded differences supported by spatial transcriptomics and multiplex immunofluorescence. Phylogenetic integration identified focal 8q24 (MYC) amplification as recurrently associated with classical-to-basal transitions occurring within tumor lineages at a frequency above chance; these transitions were accompanied by coordinated depletion of immune-response programs. Functional perturbation experiments in murine and primary human models showed that MYC knockdown reactivates classical transcriptional programs and restores immune-response pathways, supporting a role for MYC in maintaining a basal, immune-depleted state. Spatial analyses further revealed dispersed clones enriched for epithelial-to-mesenchymal transition activity and immune suppression, correlating with metastatic potential and colonization of lymph node niches. These dispersed clones shifted toward a basal phenotype, contributing to disease progression. Our findings highlight the critical role of clonal diversity, transcriptional plasticity and tumor microenvironment interactions in shaping human PDAC biology. This work provides a detailed view of the molecular and spatial heterogeneity of PDAC and offers potential avenues for therapeutic intervention targeting clonal evolution and the mechanisms driving metastasis.

Extrachromosomal DNA (ecDNA) contributes to cancer genome instability by enabling high-copy oncogene amplification and genetic intratumoral heterogeneity. Micronuclei (MNs) are common in chromosomally unstable cancers, yet their role in ecDNA-driven tumors remains unclear. Here we investigate how ecDNA damage, mitotic segregation and MN formation are linked. We find that ecDNAs are a major source of MNs following their damage, particularly under replication stress. Replication stress promotes mitotic ecDNA clustering, asymmetric inheritance, mis-segregation into MNs and reduced transcriptional output. Single-MN sequencing reveals that multiple distinct ecDNA species coalesce within individual MNs. Cells inheriting ecDNA[+] MNs display impaired proliferative capacity and are more likely to undergo cell death. In patients with MYCN-amplified neuroblastoma, higher frequencies of ecDNA[+] MNs at diagnosis are associated with improved overall survival. Together, these findings identify ecDNA mis-segregation as a mechanism that limits tumor fitness, highlighting mitotic ecDNA retention as a potential therapeutic vulnerability.

Population sequencing studies are uncovering millions of missense rare variants (RVs) implicated in the 'missing heritability' of complex diseases, yet distinguishing functional from neutral variation remains challenging. Here we present rare variant heritability-optimized scores (RovHer), a heritability-optimized method for inferring a variant's probability of being functional (that is, its contribution to trait variance). RovHer leverages exome-wide association statistics across 4,927,334 RVs and uses multivariate adaptive regression splines to integrate 75 variant- and gene-level features without a priori constraints. Across 21 traits in up to 357,086 UK Biobank European individuals, the top 1% of RovHer-prioritized missense RVs (m = 13,410) explained, on average, 16.1% of total RV heritability, a 10-fold gain over 7 alternative methods (average 1.5%; range 1.2-2.3%). RovHer's predictions are independent of allele frequency, generalize in a multi-ancestry replication and effectively identify high-risk carriers of severe clinical outcomes across 17 monogenic gene-trait pairs. Our tool showcases the value of missense RVs in population risk stratification.

Studying the genetics of intelligence provides insight into the neurobiology of cognitive function and rare neurodevelopmental conditions. Here we integrated fluid intelligence (FI) tests across different UK Biobank measures and imputed FI for unmeasured individuals, increasing the sample size from ~270,000 to >455,000. We show through extensive validation that the imputed phenotype genetically resembles measured FI, although some noncognitive signal likely remains. Combining measured and imputed FI increases the number of independent significant common SNP associations from 390 to 550 and reduces the ascertainment bias introduced by FI test participants being, on average, more educated. Rare variant analyses identify 26 significantly associated genes (false discovery rate < 1%), including 8 without strong prior evidence of involvement in intelligence or neurodevelopmental conditions, which replicate in aggregate in external cohorts. Our results demonstrate that imputed intelligence can boost power for genetic discovery and support its use to help discover new neurodevelopmental condition-associated genes.

Millions of genetic variants are linked to human disease but identifying underlying mechanisms is challenging because most variants are noncausal and lie within the noncoding genome. We developed a Micro Capture-C variant-to-function platform (MCCv) based on analysis of single-allele chromatin structure. This can identify changes in nanoscale chromatin architecture and link variants in cis-regulatory elements to target genes. Furthermore, MCCv can phase other heterozygous variants within a locus to link regulatory variants to allelically imbalanced gene expression and directly read out variant effects on chromatin interactions after genome editing. With this approach, we investigated 405 cis-regulatory elements linked to immune-mediated inflammatory disease in CD4[+] T cells. We uncovered a previously undescribed gain-of-function mechanism, which increases risk of autoimmunity through creation of a neo-CTCF motif that blocks super-enhancer contacts with the SESN3 promoter. We showed that SESN3 regulates mammalian target of rapamycin by sensing tryptophan and demonstrated its role in autoimmunity using mouse models.

Hemoglobinopathies are the most common inherited anemias worldwide. While most carriers are asymptomatic or mildly affected, severe forms of α-thalassemia, including severe hemoglobin H (HbH) disease and surviving individuals with Hb Bart's hydrops fetalis syndrome (BHFS), require lifelong transfusion support or stem cell transplantation. Reactivating the silenced embryonic α-like globin (ζ-globin) gene is a promising approach for treating these diseases. Here we identify the key cis-regulatory elements responsible for silencing the ζ-globin gene in definitive erythroid cells and subsequently develop gene-editing strategies that reactivate ζ-globin expression to therapeutically relevant levels. In a preclinical mouse model of BHFS, we show that this strategy rescues the otherwise prenatal lethal condition until late in development (E17.5). Using primary erythroid cells derived from patients with HbH disease and BHFS, we show that these approaches reactivate ζ-globin expression to levels exceeding those achieved in the mouse model, producing therapeutic levels of α-like globin.

Leveraging single-cell reference atlases to analyze new data has brought about a paradigm shift in single-cell data science akin to the first reference genome in genomics. However, methods for performing this mapping require computational expertise and, oftentimes, considerable compute power, limiting access for researchers who may benefit the most. Here ArchMap, a no-code query-to-reference mapping tool, removes this barrier by providing all-in-one automated mapping, cell-type annotation and collaborative features to analyze single-cell datasets from a wide range of integrated, often published, reference atlases and allows the extension of atlases with the growing Human Cell Atlas and related efforts. This paves the way for a democratization of reference mapping capabilities.

Alternative polyadenylation (APA) has a central role in regulation of the fate of mRNA and exhibits high variability in the brain and in neurons. However, brain APA regulation has not been mapped at cell type resolution, and the genetic control of APA in brain cell types and its contribution to genetic disease have remained unclear. Here we report a single-cell atlas of APA of the aged human brain across 2 million cells from 379 human postmortem brains across individuals with and without Alzheimer's disease (AD). We show that APA provides an independent mechanistic layer for understanding gene regulatory changes in AD, as genes with APA alterations are distinct from those with expression changes but often converge to similar pathways, including microglial activation and microtubule transport in neurons. We integrate APA variation with whole-genome sequencing to identify cell-type-resolved 3' untranslated region quantitative trait loci (3'aQTLs) for 4,288 genes. We find that 3'aQTLs preferentially colocalize with pQTLs over eQTLs. Across 17 brain traits and diseases, we identify 168 GWAS loci dependent on 3'aQTLs, of which only 17.5% are shared with eQTLs. These include PLEKHA1 and APOC2 for AD, PAK6 and AP3B2 for schizophrenia, NDUFA13 for bipolar disorder, MTCH2 for multiple brain traits, and SNCA, the top locus in Parkinson's disease. We show that 3' untranslated region choice alters SNCA mRNA localization within the cell, consistent with the observation that the SNCA risk locus acts in an eQTL-independent manner in neurons and oligodendrocytes. Our results provide a cell-type-specific foundation to interrogate the cis and trans regulation of APA and to understand the contribution to disease heritability of this key post-transcriptional layer.