Biology subjects

Ray, J. P.

Publications and source records attributed to Ray, J. P..

8 recordsLinked to original sources

bioRxiv2026

Atlas of HIV cis-regulatory elements reveals extensive transcriptional variation across clades, isolates, and within individuals

Human immunodeficiency virus (HIV) replication, persistence, and reactivation depend on transcription from integrated proviruses. Despite extensive sequence variation, how viral genetic diversity influences transcriptional regulation remains poorly understood. Here, we generate a functional regulatory atlas of HIV-1 and HIV-2 by combining tiling and saturation mutagenesis massively parallel reporter assays (MPRAs) with comparative sequence analysis and predictive modeling. By profiling thousands of HIV isolates in Jurkat and human primary CD4+T cells, we reveal extensive variation in baseline and stimulus-induced long terminal repeat (LTR) activity across and within clades, driven by distinct transcription factor configurations. These activities frequently differ among proviruses from the same individual and shift over infection and transmission without consistent selection for activity. Beyond the LTR, we identify conserved intragenic cis-regulatory elements, revealing regulatory architectures that complement LTR activity. Finally, we develop sequence-based models that accurately predict transcriptional activity, enabling scalable functional annotation of viral diversity and evolution.

microbiology↗

bioRxiv2025

Expanding the DNA Motif Lexicon of the Transcriptional Regulatory Code

Transcriptional regulatory sequences in metazoans contain intricate combinations of transcription factor (TF) motifs. Stereospecific arrangements of simple motifs constitute composite elements (CEs) that enhance DNA-protein interaction specificity and enable combinatorial regulatory logic. Despite their importance, CEs remain underexplored. We advance CE discovery and functional characterization by developing an integrated framework that combines computational prediction, experimental testing and deep learning. The extended TF motif catalog comprises both synergistic and counteracting CEs, which are supported by evidence of TF binding in vivo and in vitro. A deep learning model GRACE trained on customized massively parallel reporter assays learns the lexicon of CEs at single-nucleotide resolution. Comparative analysis with a neural network model trained on chromatin accessibility demonstrates striking convergence and distinctions within the expanded regulatory lexicon, enabling joint predictions of motif contributions and the impact of variants on chromatin structure and transcriptional activity in diverse cellular contexts.

genomics↗

bioRxiv2024

Linking candidate causal autoimmune variants to T cell networks using genetic and epigenetic screens in primary human T cells.

Genetic variants associated with autoimmune diseases are highly enriched within putative cis-regulatory regions of CD4+ T cells, suggesting that they alter disease risk via changes in gene regulation. However, very few genetic variants have been shown to affect T cell gene expression or function. We tested >18,000 autoimmune disease-associated variants for allele-specific expression using massively parallel reporter assays in primary human CD4+ T cells. The 545 expression-modulating variants (emVars) identified greatly enrich for likely causal variants. We provide evidence that many emVars are mediated by common upstream regulatory conduits, and that putative target genes of primary T cell emVars are highly enriched within a lymphocyte activation network. Using bulk and single-cell CRISPR-interference screens, we confirm that emVar-containing T cell cis-regulatory elements modulate both known and novel target genes that regulate T cell proliferation, providing plausible mechanisms by which these variants alter autoimmune disease risk.

genetics↗

bioRxiv2024

A haplotype-resolved view of human gene regulation

Diploid human cells contain two non-identical genomes, and differences in their regulation underlie human development and disease. We present Fiber-seq Inferred Regulatory Elements (FIRE) and show that FIRE provides a more comprehensive and quantitative snapshot of the accessible chromatin landscape across the 6 Gbp diploid human genome, overcoming previously unrecognized biases in existing regulatory element catalogs. FIRE enables comprehensive detection of haplotype-selective chromatin accessibility (HSCA), exposing novel imprinted elements lacking underlying parent-of-origin CpG methylation differences, and gene regulatory modules that permit genes to escape X chromosome inactivation. We uncover that the human leukocyte antigen (HLA) locus harbors the most HSCA in immune cells, where we resolve specific transcription factor (TF) binding events disrupted by disease-associated variants. Finally, we demonstrate that the regulatory landscape of a cell is littered with autosomal somatic chromatin epimutations that are propagated by clonal expansions to create mitotically stable and non-genetically deterministic chromatin alterations.

genomics↗

bioRxiv2021

Prioritization of autoimmune disease-associated genetic variants that perturb regulatory element activity in T cells

Genome-wide association studies have uncovered hundreds of autoimmune disease-associated loci; however, the causal genetic variant(s) within each locus are mostly unknown. Here, we perform high-throughput allele-specific reporter assays to prioritize disease-associated variants for five autoimmune diseases. By examining variants that both promote allele-specific reporter expression and are located in accessible chromatin, we identify 60 putatively causal variants that enrich for statistically fine-mapped variants by up to 57.8-fold. We introduced the risk allele of a prioritized variant (rs72928038) into a human T cell line and deleted the orthologous sequence in mice, both resulting in reduced BACH2 expression. Naive CD8 T cells from mice containing the deletion had reduced expression of genes that suppress activation and maintain stemness. Our results represent an example of an effective approach for prioritizing variants and studying their physiologically relevant effects.

genetics↗

bioRxiv2020

Genome-wide maps of enhancer regulation connect risk variants to disease genes

Genome-wide association studies have now identified tens of thousands of noncoding loci associated with human diseases and complex traits, each of which could reveal insights into biological mechanisms of disease. Many of the underlying causal variants are thought to affect enhancers, but we have lacked genome-wide maps of enhancer-gene regulation to interpret such variants. We previously developed the Activity-by-Contact (ABC) Model to predict enhancer-gene connections and demonstrated that it can accurately predict the results of CRISPR perturbations across several cell types. Here, we apply this ABC Model to create enhancer-gene maps in 131 cell types and tissues, and use these maps to interpret the functions of fine-mapped GWAS variants. For inflammatory bowel disease (IBD), causal variants are >20-fold enriched in enhancers in particular cell types, and ABC outperforms other regulatory methods at connecting noncoding variants to target genes. Across 72 diseases and complex traits, ABC links 5,036 GWAS signals to 2,249 unique genes, including a class of 577 genes that appear to influence multiple phenotypes via variants in enhancers that act in different cell types. Guided by these variant-to-function maps, we show that an enhancer containing an IBD risk variant regulates the expression of PPIF to tune mitochondrial membrane potential. Together, our study reveals insights into principles of genome regulation, illuminates mechanisms that influence IBD, and demonstrates a generalizable strategy to connect common disease risk variants to their molecular and cellular functions.

genetics↗

bioRxiv2020

Genome-wide discovery of lupus genetic risk variant allelic regulatory activity

Genome-wide association studies of Systemic Lupus Erythematosus (SLE) nominate 3,073 genetic variants at 91 risk loci. To systematically screen these variants for allelic transcriptional enhancer activity, we constructed a massively parallel reporter assay (MPRA) library comprising 12,396 DNA oligonucleotides containing the genomic context around every allele of each SLE variant. Transfection into the Epstein-Barr virus-transformed B cell line GM12878 revealed 482 variants with enhancer activity, with 51 variants showing genotype-dependent (allelic) enhancer activity at 27 risk loci. Comparison of MPRA results in GM12878 and Jurkat T cell lines highlights shared and unique allelic transcriptional regulatory mechanisms at SLE risk loci. In-depth analysis of allelic transcription factor (TF) binding at and around allelic variants identifies one class of TFs whose DNA-binding motif tends to be directly altered by the risk variant and a second, larger class of TFs that bind allelically without direct alteration of their motif by the variant. Collectively, our approach provides a blueprint for the discovery of allelic gene regulation at risk loci for any disease and offers insight into the transcriptional regulatory mechanisms underlying SLE.

genomics↗

bioRxiv2019

MAUDE: Inferring expression changes in sorting-based CRISPR screens

Pooled CRISPR screens allow high-throughput interrogation of genetic elements that alter expression of a reporter gene readout. New computational methods are needed to model these data. We created MAUDE (Mean Alterations Using Discrete Expression) for quantifying the impact of guide RNAs on a target genes expression in a pooled, sorting-based expression screen. MAUDE quantifies guide-level effects by modeling the distribution of cells across sorting expression bins. It then combines guides to estimate the statistical significance and effect size of targeted genetic elements. We show that MAUDE significantly improves over previous approaches and provide experimental design guidelines to best leverage MAUDE.

bioinformatics↗