Cell genomics最新文献

Long interspersed nuclear element 1 (L1) retrotransposons represent a vast source of genetic variability. However, mechanistic analysis of whether and how L1s contribute to human developmental programs is lacking, in part due to the challenges associated with specific profiling and manipulation of human L1 expression. Here, we show that thousands of hominoid-specific L1 integrants are expressed in human induced pluripotent stem cells and cerebral organoids. The activity levels of individual L1 promoters vary widely and correlate with an active epigenetic state. Efficient on-target CRISPR interference (CRISPRi) silencing of L1s revealed nearly a hundred co-opted L1-derived chimeric transcripts, and L1 silencing resulted in changes in neural differentiation programs and reduced cerebral organoid size. Together, these data implicate L1s and L1-derived transcripts in hominoid-specific CNS developmental processes.

The NPIP gene family is among the most positively selected gene families in humans/apes and drives independent duplication in primate lineages. These duplications promote genetic instability, leading to recurrent disease-associated microduplication and microdeletion syndromes. Despite its importance, little is known about its function or variation in humans, as short-read sequencing cannot distinguish high-identity duplications. Using long-read assemblies of 169 human haplotypes, we find extreme variation in the content and organization of NPIP loci. We identify fixed and polymorphic paralogs and observe ongoing positive selection. With long-read RNA sequencing (RNA-seq), we create paralog-specific gene models, the majority of which were not previously documented, and observe paralog-specific tissue specificity. This analysis of an exceptionally dynamic gene family provides candidates for future functional study.

The Yellow and Yangtze river basins in China are among the world's oldest independent agricultural centers, known for the domestication of millet and rice, respectively, yet their genetic history is poorly understood. Here, we present genome-wide data from 74 Middle Neolithic genetic samples from these regions, showing marked genetic differentiation but bidirectional gene flow, supporting a demic diffusion model of mixed farming. Yellow River populations exhibit distinct genetic substructures resulting from interactions with surrounding groups during the mid-Neolithic expansion of millet agriculture. Upper Yellow River populations are genetically linked to Tibetan Plateau populations and possess the earliest adaptive EPAS1 haplotype (∼5,800 BP) among modern humans. Meanwhile, Yangtze River rice farmers show genetic affinity with Neolithic to present-day southeast coastal China and Austronesian populations, tracing the origins of proto-Austronesians farther north to the Yangtze River. These findings offer new insights into the impact of mid-Neolithic agricultural expansion on human genetic history.

Urine contains fragments of cell-free DNA (cfDNA) that offer molecular insights into processes within the urinary system and the body. It remains unclear whether these fragments exist as chromatin and retain chromatin modifications from their cells of origin. Here, we employ cell-free chromatin immunoprecipitation followed by sequencing (cfChIP-seq) on human urine to address this issue. We show that cf-nucleosomes can be captured from urine and preserve histone modifications associated with gene activation and repression. Analysis in healthy individuals reveals distinct tissue contributions to urine cf-nucleosomes, including a kidney-derived population not detected in matched exfoliated cells or plasma. This suggests that kidney filtration largely excludes plasma cf-nucleosomes. In patients with bladder cancer, urine cf-nucleosomes reflect tumor-associated transcriptional programs and immune responses. These findings highlight the utility of urine cf-nucleosomes as accessible, non-invasive biomarkers for studying renal physiology and monitoring urinary pathologies.

Psoriasis vulgaris (PsV) is an immune-mediated inflammatory skin disorder with complex genetic architecture. Most genome-wide association studies (GWASs) of PsV have been limited to analyzing common single-nucleotide variants in Europeans, lacking diversity in the variant spectrum and ancestral background. To investigate the contribution of rare variants (RVs) and structural variants (SVs), we perform a whole-genome sequencing study involving 1,415 PsV cases and 3,968 controls in Japanese. A GWAS signal at IFNLR1 is fine-mapped to a 3.3-kb deletion SV disrupting an epithelium-specific putative enhancer, which is validated by PacBio long-read sequencing. Gene-based RV analyses identify two susceptibility genes: IFIH1 (p = 9.8 × 10^-6) and CERCAM (p = 4.1 × 10^-7). Notably, IL36RN, a causative gene for generalized pustular psoriasis, a rare and lethal multi-systemic inflammatory disorder, is associated with common PsV (p = 1.2 × 10^-4). Finally, Cercam knockout (Cercam^-/-) in an imiquimod-induced psoriasis mouse model aggravates dermatitis with elevated T cell retention in the subepidermis. Our study elucidates the overlooked genetic basis of PsV.

The repair of a DNA double-strand break (DSB) by non-homologous end joining (NHEJ) generally leaves an intact or minimally modified sequence. Resection exposes single-stranded DNA and directs repair toward homology-dependent pathways and away from NHEJ. Here, we report that in Saccharomyces cerevisiae, the Cdc13/Stn1/Ten1 (CST) complex, characterized for its telomeric functions, acts after resection initiation to mediate a back-up NHEJ repair. We found a CST-specific mutation signature after repair characterized by deletions of 5-85 bp that were mostly dependent on NHEJ, with a subset dependent on microhomology-mediated end joining (MMEJ). The interaction between CST and Polα-primase is critical for these intermediate-size deletions, suggesting a role for fill-in synthesis, thus limiting extensive resection, which would otherwise lead to MMEJ-dependent deletions of several kilobases. Collectively, these results depict a complex picture of repair pathway choice where CST facilitates post-resection NHEJ repair, promoting local deletions but guarding against larger and potentially more deleterious deletions and rearrangements.

Circulating cell-free DNA (cfDNA) is a promising molecular biomarker, but its role in severe infection is unclear. Here, we profile cfDNA from sepsis patients and controls, demonstrating a 41-fold increase during disease. Methylation-based deconvolution revealed similar cfDNA compositions in the two groups, suggesting that cfDNA accumulation during disease is due not to excess cell death but to impaired hepatic clearance. Fragmentation and end-motif patterns both support this hypothesis, suggesting prolonged exposure of cfDNA to circulating nucleases. In addition, we show that cfDNA retains nucleosome footprints informative of gene activity. By developing a novel method to quantify these footprints and integrate them with single-cell data, we report an increase in cfDNA from Kupffer cells and liver parenchyma in patients with liver dysfunction. Finally, we show that cfDNA contains pathogen-derived material, highlighting its diagnostic potential. This high-throughput, multimodal study provides a reference for understanding cfDNA's role in sepsis and critical illness.

Inherited genetic variants contribute to Barrett's esophagus (BE) and esophageal adenocarcinoma (EAC), but it is unknown which cell types are involved in this process. We performed single-cell RNA sequencing of BE, EAC, and paired normal tissues and integrated genome-wide association data to determine cell-type-specific genetic risk and cellular processes that contribute to BE and EAC. The analysis reveals that EAC development is driven to a greater extent by local cellular processes than BE development and suggests that one cell type of BE origin (intestinal metaplasia cells) and cellular processes that control the differentiation of columnar cells are of particular relevance for EAC development. Specific subtypes of fibroblasts and endothelial cells likely contribute to BE and EAC development, while dendritic cells and CD4⁺ memory T cells seem to contribute to BE development. The diagnostic value of markers characterizing the cell types and cellular processes should be explored for EAC prediction.

Enhancers are known to spatiotemporally regulate gene transcription, yet the identification of enhancers and their target genes is often indirect, low resolution, and/or assumptive. To identify and functionally perturb enhancers at their endogenous sites, we performed a pooled tiling CRISPR activation (CRISPRa) screen surrounding PHOX2B, a master regulator of neuronal cell fate and a key player in neuroblastoma, and found many CRISPRa-responsive elements (CaREs) that alter cellular growth. To determine CaRE target genes, we developed TESLA-seq (targeted single-cell activation), which combines CRISPRa screening with targeted single-cell RNA sequencing and enables the parallel readout of the effect of hundreds of enhancers on all genes in the locus. While most TESLA-revealed CaRE-gene relationships involved neuroblastoma-related regulatory elements, we found many CaREs and target connections normally active only in other tissues. This highlights the power of TESLA-seq to reveal gene regulatory networks, including edges active outside of a given experimental system.

Single-cell omics, named Method of the Year three times, have revolutionized biological research by enabling the high-resolution exploration of cellular heterogeneity and molecular processes. Initially centered on transcriptomics, this rapidly evolving field now ranges from multiomics to spatial analysis, with expanding customization options. The ubiquity of such analyses and the lack of a unified pipeline necessitate the development of scalable, flexible, and integrated tools and workflows. The Galaxy platform has responded to these technological advancements, extending its repertoire of freely accessible tools and workflows, backed by expert-reviewed and user-informed training resources to empower researchers to perform and interpret their own analyses. With more than 175 tools, 120 training resources, and 300,000 jobs running at the time of writing, this process has culminated in the development of Galaxy single-cell and spatial omics community (SPOC), designed to promote global collaboration in advancing usable, reproducible, accessible, and sustainable single-cell and spatial omics research.