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Biology subjects

Pachter, L.

Publications and source records attributed to Pachter, L..

At least 19 recordsLinked to original sources

Dimensionality Reduction for Cyclic Compositional Data

The cyclic nature of many biological systems is frequently reflected in periodic gene expression patterns. Standard dimensionality reductions can reveal such variation but do not link a sample's position on a cycle to an ordering of the molecular features that define it. We introduce clrcycle, which applies a centered log-ratio transformation, learns a cyclic feature ordering by maximizing variance captured by the first Fourier mode, and projects samples onto the corresponding cosine and sine coordinates. The resulting two-dimensional geometry assigns each sample a phase and a radius measuring the magnitude of its projection onto the learned Fourier circle. Across a mouse circadian atlas, clrcycle recovers known temporal organization and reveals tissue-specific differences in rhythmic timing. In FUCCI-profiled single cells, clrcycle represents independently measured cell-cycle phase in two-dimensional coordinates more reliably than conventional two-dimensional embeddings. In human endometrium, a circular coordinate learned from 250 highly variable genes recovers menstrual-stage organization and can stage unseen endometrial biopsies. Together, these results establish clrcycle as powerful, explicitly circular dimensionality reduction technique to quantify high-dimensional cyclic variation across biological cycles.

bioinformatics↗

Biological and technical spatial variation in spatial transcriptomics

Spatial transcriptomics technologies enable the identification of spatially variable genes that can shed light on tissue structure and function. However, spatial variability of gene expression can be due not only to biological function but also technical artifact. We develop a model for spatially variable capture that forms part of a bursty transcription model, allowing us to distinguish biological variability from spatially variable technical noise. Our approach is implemented in a program called stipple, which we demonstrate can be useful for identifying and interpreting spatial gene expression variation from spatial transcriptomics assays.

bioinformatics↗

fastQpick: scalable bootstrap and subsampling of FASTQ reads

Summary: fastQpick is a command-line tool and Python library for sampling FASTQ reads with replacement. Sampling with replacement turns a single FASTQ file into an arbitrary number of bootstrap replicates, which enables uncertainty quantification and statistical analysis at the level of raw reads. This process answers questions such as how much an abundance estimate would change if the library were resequenced, or whether a low-abundance call is robust to the particular reads that were sequenced. fastQpick works efficiently on large libraries by streaming files in two passes by default: first to count reads and create a hash-based counter, and then to write the sample. It generates a full-size bootstrap replicate of a 500-million-read library in under 30 minutes with 9.4 GB of peak memory, with a low-memory mode that reduces the peak to 1.4 GB. A single-pass mode draws samples in a single read through the file, using O(1) working memory and producing an output size that is exact in expectation but not fixed. In a real yeast RNA-seq experiment, bootstrap replicates generated by fastQpick recover the sampling uncertainty of transcript abundance estimates, matching the analytic multinomial standard errors to within a few percent. Availability and implementationfastQpick is open source and freely available under the MIT license on GitHub at https://github.com/pachterlab/fastQpick and on PyPI (pip install fastQpick).

bioinformatics↗

The Rayleigh Quotient and Contrastive Principal Component Analysis II

Contrastive principal component analysis (PCA) methods are effective approaches to dimensionality reduction where variance of a target dataset is maximized while variance of a background dataset is minimized. We previously described how contrastive PCA problems can be written as solutions to generalized eigenvalue problems that maximize particular instantiations of the Rayleigh quotient. Here, we discuss two extensions of contrastive PCA: we use kernel weighting from spatial PCA (k-{rho}PCA) to contrast spatial and non-spatial axes of variation, and separately solve the Rayleigh quotient in the space of basis function coefficients (f-{rho}PCA) to find modes of variation in functional data. Together, these extensions expand the scope of contrastive PCA while unifying disparate fields of spatial and functional methods within a single conceptual and mathematical framework. We showcase the utility of these extensions with several examples drawn from genomics, analyzing gene expression in cancer and immune response to vaccination.

bioinformatics↗

Spectral requirements for cooperation

We introduce a spectral existence criterion for the evolution of cooperation in the form of the inequality{lambda} maxb > c, where{lambda} max is the leading eigenvalue of an interaction operator encoding population structure, and b and c represent benefit and cost tradeoffs, respectively. Nowaks five rules for the evolution of cooperation correspond to cases in which the cooperation condition reduces to a scalar assortment coefficient. These results follow from the Price equation, which sheds light on a long-standing debate on the role of inclusive fitness and evolutionary dynamics in explaining the evolution of cooperation.

evolutionary biology↗

Hybrid crosses reveal a cell-type-specific landscape of mouse regulatory variation

Understanding the genetic architecture of gene expression is fundamental to evolutionary biology and medicine. As part of the IGVF Consortium, we present a single-nucleus RNA-seq resource of 5.3 million nuclei across eight tissue groups, featuring seven F1 hybrids from C57BL/6J dams crossed with the other Collaborative Cross founder strains for comparison against parental strains. We identify 25,864 genes (91% of those detected) exhibiting non-conserved regulatory behavior in at least one of 92 cell types in one or more crosses. Our results show that while cis-acting variation primarily drives divergence, trans-acting effects are substantially more cell-type specific and sensitive to tissue environment. Notably, bulk tissue analyses frequently mask these signals, particularly in smaller populations such as astrocytes. Furthermore, increasing genetic divergence primarily expands the landscape of cis-acting variation, while trans-acting effects remain stable across genetic distances within species. This atlas establishes a foundational framework for decoding the complex interplay between genetic variation and cell-type-specific regulation across the mammalian body.

genomics↗

Single-Cell Genomics Decontamination with CellSweep

Single-cell genomics technologies enable high-throughput cell profiling, but technical contamination remains an obstacle to accurate downstream analysis. Free-floating ambient molecules released from lysed cells and global bulk contamination introduced during library preparation can distort molecular profiles. These artifacts can obscure cellular identities and reduce the reliability of differential analysis or clustering results. We present an efficient and effective approach to removing ambient and bulk contamination that can be applied to data generated from a wide variety of technologies. We show that our tool, CellSweep, outperforms other methods to remove artifacts using numerous benchmarks.

bioinformatics↗

Differential analysis of genomics count data with edge

The edgeR Bioconductor package is one of the most widely used tools for differential expression analysis of count-based genomics data. Despite its popularity, the R-only implementation limits its integration with the Python-centric ecosystem that has become dominant in single-cell genomics. We present edgePython, a Python port of edgeR 4.8.2 that extends the framework with a negative binomial-gamma mixed model for multi-subject single-cell analysis and empirical Bayes shrinkage of cell-level dispersion.

bioinformatics↗

Science should be machine-readable

We develop a machine-automated approach for extracting results from papers, which we assess via a comprehensive review of the entire eLife corpus. Our method facilitates a direct comparison of machine and peer review, and sheds light on key challenges that must be overcome in order to facilitate AI-assisted science. In particular, the results point the way towards a machine-readable framework for disseminating scientific information. We therefore argue that publication systems should optimize separately for the dissemination of data and results versus the conveying of novel ideas, and the former should be machine-readable.

scientific communication and education↗

Computational array reconstruction for spatial transcriptomics

Several optics-free spatial transcriptomics assays that make use of diffusion of reads from beads have recently been developed, and provide a proof of principle for high-throughput, untargeted, spatial transcriptomics. These approaches leads to a challenging computational array reconstruction problem. We show that an statistical inference method, which we term accumap, outperforms the current approach which is to run UMAP on the bead count matrix. The accumap method makes use of predicted distances to reconstruct bead positions and can improve the accuracy of reconstruction by a factor of 3 in the diffusible Slide-Tag simulation framework.

bioinformatics↗

k-spaces: Mixtures of Gaussian latent variable models

Principal component analysis (PCA) and k-means clustering are two seemingly different methods for dimension reduction and clustering, respectively, but can be understood as special cases of inference in a Gaussian latent variable model framework. We leverage this insight to develop a probabilistic framework and methods for simultaneous dimension reduction, clustering, and latent space learning that are efficient and interpretable, and that can replace current ad hoc combinations of PCA and clustering. The algorithm, k-spaces, has broad applicability, which we demonstrate in several distinct genomic settings. In particular, we show how k-spaces can be used to model gene expression in quantitative hybridization chain reaction (qHCR) images, for inference in epigenomics, and for dimension reduction of single-cell RNA-sequencing data.

bioinformatics↗

Determining gene specificity from multivariate single-cell RNA sequencing data

An important application of single-cell genomics experiments is to identify genes specific to biological categories or experimental conditions. Although numerous approaches have been proposed to identify such genes, we consider an axiomatic approach based on defining properties that a specificity measure should have. This leads us to develop ember (Entropy Metrics for Biological ExploRation), which we show is the only method satisfying four key desired properties for a specificity measure. Applying ember to eight tissues from eight founder mouse strains, we find that gene specificity is often unintuitive: canonical markers can be supplanted, housekeeping genes are context-dependent, and mouse strain can drive unexpected cell type switching. Unsupervised learning on entropy metrics uncovers shared genes specialized to male gonads and kidney, as well as genes specific to non-consecutive developmental stages in the kidney. To facilitate further exploration of gene specificity in mice, we have also developed a comprehensive specificity database, along with a web interface, API and MCP server. Extending ember to a human PBMC dataset collected from 255 diverse individuals, we find that variation in PBMCs is largely localized to classical monocytes. We also find genes with unique specificity by sex, age and ancestral background. Together, these applications establish ember as a powerful tool and provide a roadmap for elucidating the impact of human genetic variation using the murine model.

bioinformatics↗

Biophysical constraints on mRNA decay rates shape macroevolutionary divergence in steady-state abundances

Interspecific comparisons of cell-type-specific gene expression levels can provide information about the evolutionary processes that drove divergence between species. From these comparisons, it is now evident that the predominant mode of gene expression evolution has been stabilizing selection, both on the steady-state (mean) protein levels as well as on the mRNA levels with additional lineage-specific shifts resulting from directional selection. However, as all previous work has used bulk RNA measurements, it has been impossible to determine which of the many cellular processes that contribute to mean abundances are highly constrained and which are more evolutionary labile. Assessing this is further complicated by the expectation that components of complex systems will evolve over time independent of changes in the selective regime so long as the net output of a system (i.e., mean expression) remains near the evolutionary optima. This process, known as evolutionary systems drift (ESD), has been frequently invoked as a non-adaptive explanation for changes in cellular phenotypes but has never been quantitatively tested or accounted for in any statistical test for selective constraints. Here, we develop a new paradigm that addresses both of these open problems simultaneously. Using single-cell expression data and biophysical models, we estimate mRNA transcriptional bursting rates, splicing rates, and decay rates across multiple vertebrate species. We then derive new mathematical results that describe how these various biophysical parameters are expected to co-evolve under ESD and then test whether we need additional evolutionary constraints to explain the divergences in these parameters. We find evidence that the biophysical parameters are indeed evolving in a coordinated manner as predicted by ESD and that there are additional strong constraints on transcriptional bursting, likely as a consequence of selection to reduce noise in expression. More broadly, this work opens up a whole new approach for studying the evolutionary dynamics of complex cellular systems.

evolutionary biology↗

The Rayleigh Quotient and Principal Component Analysis I

Contrastive learning methods can be powerful tools for genomics, enabling the identification of signals in an experiment via dimension reduction while reducing noise using a control. One such popular approach is contrastive PCA, which, despite being used in a variety of settings, does not scale to large datasets. We show that the contrastive PCA objective is an approximation of a Rayleigh quotient, analogous in form to Fishers linear discriminant analysis and the common spatial patterns method. The Rayleigh quotient is{rho} PCA, satisfies numerous desirable properties, and provides an interpretable form of dimension reduction via generalized eigenvectors. We demonstrate that{rho} PCA is more accurate than contrastive PCA and much more efficient. We also show how it can be used not only for dimension reduction of data with respect to a control, but also for contrasting conditions via an analysis of single-nucleus transcriptomics data. Finally, we discuss probabilistic interpretations of{rho} PCA that provide further insight into its effective performance.

bioinformatics↗

Joint Biophysical Modeling of Paired Single-Cell RNA and Protein Measurements

Surface protein measurements can supplement gene expression information from single-cell RNA sequencing to provide a more complete assessment of cell identity and function. Recently developed multiomic assays facilitate such measurements, and can, in principle, be utilized to understand the dynamics of transcription and translation. We develop a framework for biophysical modeling of transcription jointly with translation from single-cell data, along with a suitable technical noise model for sequencing data. We demonstrate its efficacy using simulations, and illustrate how it can be useful in practice with 10x multiomic data. Our proof-of-principle highlights the potential for jointly modeling transcription and translation as data quality and measurement accuracy improves.

biophysics↗

Reference-based variant detection with varseek

Variant detection from sequencing data is fundamental for genomics and is the first step in a wide range of applications, ranging from genome-wide association studies to disease diagnosis. Widely used tools for variant detection utilize a de novo approach that is based on a combination of read mapping algorithms and statistical methods for identifying genetic variation from error-prone sequencing data. This approach has been successful, although the detection of insertion and deletion variants, as well as the detection of variants from low-coverage data, remain challenging problems. We introduce varseek, a reference-based approach to variant detection that provides large improvements in performance in these challenging cases. The varseek approach utilizes a k-mer pseudoalignment approach, which provides the ability to identify variants at single-cell resolution in single-cell transcriptomics data. We showcase the versatility and performance of varseek for detecting tumor-specific COSMIC variants in glioblastoma single-cell sequencing.

bioinformatics↗

Transcriptomic responses to endurance exercise training in rats

The bio-molecular changes of exercise, and how to best optimize them for improved performance, are an important human health research question. A recent study by the Molecular Transducers of Physical Activity Consortium (MoTrPAC) used a cohort of Rattus norvegicus to produce a whole-organism molecular map of the temporal effects of endurance exercise training. This dataset, encompassing hundreds of assays across 19 different tissues, can be used to evaluate the predictive power of gene expression, understand isoform-level changes in response to exercise, and with modern tools can be examined for associations with viral infection. Our analysis of the RNA-seq data reveals that gene expression can accurately predict the amount of exercise a rat was trained in. Additionally, we find biologically relevant isoform-level differences in expression that are masked in gene-level analysis. Finally, we find a potential novel virus that may negatively impact physiological measurements. This more comprehensive analysis provides a blueprint for directing similar analyses with respect to physiological perturbations across organisms.

genomics↗

Pseudoassembly of k-mers

We introduce a pseudoassembly approach to identifying variation in sets of genomic sequences via colored de Bruijn graphs. Our pseudoassembly method is implemented in a program called klue that assembles k-mers into sequences compatible with a variant-aware extension of pseudoalignment. We show that this approach can be used to identify cell-type specific de novo variants from single-cell RNA-seq in a mouse melanoma model.

bioinformatics↗