Nature Methods最新文献

Comparing chromatin contact maps is an essential step in quantifying how three-dimensional (3D) genome organization shapes development, evolution, and disease. However, methods often disagree, and no gold standard exists for comparing pairs of maps. Here, we evaluate 25 ways to compare contact maps using Micro-C and Hi-C data from two cell types and in silico-generated contact maps. We identify similarities and differences between the methods and quantify their robustness to common sources of biological and technical variation, including losses and gains of CTCF-binding sites, changes in contact intensity or patterns, and noise. We find that global comparison methods, such as mean squared error, are suitable for initial screening; however, biologically informed methods are necessary for identifying how maps diverge and for proposing specific functional hypotheses. We provide a reference guide, codebase, and thorough evaluation for rapidly comparing chromatin contact maps at scale to enable biological insights into 3D genome organization.

Advances in spatial profiling technologies are providing insights into how molecular programs are influenced by local signaling and environmental cues. However, cell fate specification and tissue patterning involve the interplay of biochemical and mechanical feedback. Here we develop a computational framework that enables the joint statistical analysis of transcriptional and mechanical signals in the context of spatial transcriptomics. To illustrate the application and utility of the approach, we use spatial transcriptomics data from the developing mouse embryo to infer the forces acting on individual cells, and use these results to identify mechanical, morphometric and gene expression signatures that are predictive of tissue compartment boundaries. In addition, we use geoadditive structural equation modeling to identify gene modules that predict the mechanical behavior of cells in an unbiased manner. This computational framework is easily generalized to other spatial profiling contexts, providing a generic scheme for exploring the interplay of biomolecular and mechanical cues in tissues.

Recent advances in single-cell RNA sequencing (scRNA-seq) techniques have provided unprecedented insights into the heterogeneity of various tissues. However, gene expression data alone often fails to capture and identify changes in cellular pathways and complexes, as they are more discernible at the protein level. Moreover, analyzing scRNA-seq data presents further challenges due to inherent characteristics such as high noise levels and zero inflation. In this study, we propose an approach to address these limitations by integrating scRNA-seq datasets with a protein-protein interaction network. Our method utilizes a unique dual-view architecture based on graph neural networks, enabling joint representation of gene expression and protein-protein interaction network data. This approach models gene-to-gene relationships under specific biological contexts and refines cell-cell relations using an attention mechanism. Next, through comprehensive evaluations, we demonstrate that scNET better captures gene annotation, pathway characterization and gene-gene relationship identification, while improving cell clustering and pathway analysis across diverse cell types and biological conditions.

The human genome contains instructions to transcribe more than 200,000 RNAs. However, many RNA transcripts are generated from the same gene, resulting in alternative isoforms that are highly similar and that remain difficult to quantify. To evaluate the ability to study RNA transcript expression, we profiled seven human cell lines with five different RNA-sequencing protocols, including short-read cDNA, Nanopore long-read direct RNA, amplification-free direct cDNA and PCR-amplified cDNA sequencing, and PacBio IsoSeq, with multiple spike-in controls, and additional transcriptome-wide N⁶-methyladenosine profiling data. We describe differences in read length, coverage, throughput and transcript expression, reporting that long-read RNA sequencing more robustly identifies major isoforms. We illustrate the value of the SG-NEx data to identify alternative isoforms, novel transcripts, fusion transcripts and N⁶-methyladenosine RNA modifications. Together, the SG-NEx data provide a comprehensive resource enabling the development and benchmarking of computational methods for profiling complex transcriptional events at isoform-level resolution.

The Xenium In Situ platform is a new spatial transcriptomics product commercialized by 10x Genomics, capable of mapping hundreds of genes in situ at subcellular resolution. Given the multitude of commercially available spatial transcriptomics technologies, recommendations in choice of platform and analysis guidelines are increasingly important. Herein, we explore 25 Xenium datasets generated from multiple tissues and species, comparing scalability, resolution, data quality, capacities and limitations with eight other spatially resolved transcriptomics technologies and commercial platforms. In addition, we benchmark the performance of multiple open-source computational tools, when applied to Xenium datasets, in tasks including preprocessing, cell segmentation, selection of spatially variable features and domain identification. This study serves as an independent analysis of the performance of Xenium, and provides best practices and recommendations for analysis of such datasets.

The availability of single-cell transcriptomics has allowed the construction of reference cell atlases, but their usefulness depends on the quality of dataset integration and the ability to map new samples. Previous benchmarks have compared integration methods and suggest that feature selection improves performance but have not explored how best to select features. Here, we benchmark feature selection methods for single-cell RNA sequencing integration using metrics beyond batch correction and preservation of biological variation to assess query mapping, label transfer and the detection of unseen populations. We reinforce common practice by showing that highly variable feature selection is effective for producing high-quality integrations and provide further guidance on the effect of the number of features selected, batch-aware feature selection, lineage-specific feature selection and integration and the interaction between feature selection and integration models. These results are informative for analysts working on large-scale tissue atlases, using atlases or integrating their own data to tackle specific biological questions.

The Human BioMolecular Atlas Program (HuBMAP) aims to construct a 3D Human Reference Atlas (HRA) of the healthy adult body. Experts from 20+ consortia collaborate to develop a Common Coordinate Framework (CCF), knowledge graphs and tools that describe the multiscale structure of the human body (from organs and tissues down to cells, genes and biomarkers) and to use the HRA to characterize changes that occur with aging, disease and other perturbations. HRA v.2.0 covers 4,499 unique anatomical structures, 1,195 cell types and 2,089 biomarkers (such as genes, proteins and lipids) from 33 ASCT+B tables and 65 3D Reference Objects linked to ontologies. New experimental data can be mapped into the HRA using (1) cell type annotation tools (for example, Azimuth), (2) validated antibody panels or (3) by registering tissue data spatially. This paper describes HRA user stories, terminology, data formats, ontology validation, unified analysis workflows, user interfaces, instructional materials, application programming interfaces, flexible hybrid cloud infrastructure and previews atlas usage applications.