Biology subjects

Nolan, B.

Publications and source records attributed to Nolan, B..

5 recordsLinked to original sources

bioRxiv2026

Guanine content in the CTCF motif encodes a vulnerability to persistent, alcohol-induced redistribution

Alcohol consumption is a leading contributor to the global disease burden, yet its molecular consequences remain incompletely understood. Ethanols first metabolite, acetaldehyde, modifies guanine to form DNA adducts and DNA-protein crosslinks that block elongating RNA polymerase II. Because acetaldehyde acts specifically on guanine, we hypothesized that chromatin features would be disrupted in a sequence-dependent rather than stochastic manner. Here we confirm that alcohol exposure displaces elongating RNA polymerase II (RNAPIIs2p) from gene bodies, relocating to promoters. In addition to this confirmation, we found that RNAPIIs2p also relocates to discrete guanine-rich intergenic CTCF sites. Loci with high guanine content at the CTCF motifs zinc finger 5 (ZF5) binding position showed the largest increase in both CTCF and cohesin (RAD21), with CTCF occupancy changes persisting even six days after alcohol withdrawal. Surprisingly, using Hi-C, we found that these changes to CTCF and cohesin do not impact the pronounced structural CTCF loops, but are instead primarily associated with altered guanine-rich enhancer-promoter contacts that connect to lasting changes to gene expression. Beyond our cell-line model, the same sequence rule accounts for altered CTCF occupancy in a mouse starvation (metabolic stress) model and in the placentae after paternal alcohol exposure. Together these findings link alcohol exposure to sequence-directed changes to chromatin whose rules are conserved in alcohol-associated disease.

molecular biology↗

bioRxiv2026

Kilobase-scale compartments enabled by CRUSH reveal regulatory programs across cell types, single-cells, and ancient mammoths

How chromatin is spatially organized in the nucleus has long been studied through the lens of large-scale A/B compartments, but whether these sizes reflect true biological units or analytical artifacts has remained unclear. We find that limits imposed by the conventional eigenvector-based compartment calling have required extreme sequencing depth and coarse resolution, obscuring regulatory-scale organization. We developed CRUSH to iteratively refine compartments to 1 kb resolution without the need for extreme sequencing depth, we show that kilobase-scale A/B segregation (micro-compartments) is evident across cell types. Across these maps, we demonstrate that RNA polymerase II pausing contributes to a sub-genic compartment signature at the transcription start site and that active enhancers almost universally occupy the A compartment. We then show that fine-scale compartment maps can resolve cancer subtype-specific regulatory programs, single-cell tissue identity, and cold-adaptation regulomes in a 52,000-year-old woolly mammoth. These findings establish chromatin compartmentalization as a gene-scale regulatory feature with broad implications for development, disease, and genome evolution. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=176 SRC="FIGDIR/small/719469v1_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@15c1992org.highwire.dtl.DTLVardef@192f541org.highwire.dtl.DTLVardef@123a737org.highwire.dtl.DTLVardef@1b7292f_HPS_FORMAT_FIGEXP M_FIG C_FIG CRUSH uses iterative resolution walking to identify and correct A/B compartment measurements. This allows identification of A/B compartments at kilobase resolution with low sequencing depth and consistency across methodologies. Kilobase-scale compartments reveal sub-genic compartment organization, compartment identities in single-cells, and a distinct cold-adapted regulome in mammoths.

genomics↗

bioRxiv2024

HiCrayon reveals distinct layers of multi-state 3D chromatin organization

The co-visualization of chromatin conformation with 1D omics data is key to the multi-omics driven data analysis of 3D genome organization. Chromatin contact maps are often shown as 2D heatmaps and visually compared to 1D genomic data by simple juxtaposition. While common, this strategy is imprecise, placing the onus on the reader to align features with each other. To remedy this, we developed HiCrayon, an interactive tool that facilitates the integration of 3D chromatin organization maps and 1D datasets. This visualization method integrates data from genomic assays directly into the chromatin contact map by coloring interactions according to 1D signal. HiCrayon is implemented using R shiny and python to create a graphical user interface (GUI) application, available in both web or containerized format to promote accessibility. HiCrayon is implemented in R, and includes a graphical user interface (GUI), as well as a slimmed-down web-based version that lets users quickly produce publication-ready images. We demonstrate the utility of HiCrayon in visualizing the effectiveness of compartment calling and the relationship between ChIP-seq and various features of chromatin organization. We also demonstrate the improved visualization of other 3D genomic phenomena, such as differences between loops associated with CTCF/cohesin vs. those associated with H3K27ac. We then demonstrate HiCrayons visualization of organizational changes that occur during differentiation and use HiCrayon to detect compartment patterns that cannot be assigned to either A or B compartments, revealing a distinct 3rd chromatin compartment. Overall, we demonstrate the utility of co-visualizing 2D chromatin conformation with 1D genomic signals within the same matrix to reveal fundamental aspects of genome organization. Local version: https://github.com/JRowleyLab/HiCrayon Web version: https://jrowleylab.com/HiCrayon

genomics↗

bioRxiv2021

Mining underutilized whole-genome sequencing projects to improve 16S rRNA databases

Current approaches to interpreting 16S rDNA amplicon data are hampered by several factors. Among these are database inaccuracy or incompleteness, sequencing error, and biased DNA/RNA extraction. Existing 16S rRNA databases source the majority of sequences from deposited amplicon sequences, draft genomes, and complete genomes. Most of the draft genomes available are assembled from short reads. However, repeated ribosomal regions are notoriously difficult to assemble well from short reads, and as a consequence the short-read-assembled 16S rDNA region may be an amalgamation of different loci within the genome. This complicates high-resolution community analysis, as a draft genomes 16S rDNA sequence may be a chimera of multiple loci; in such cases, the draft-derived sequences in a database may not represent a 16S rRNA sequence as it occurs in biology. We present Focus16, a pipeline for improving 16S rRNA databases by mining NCBIs Sequence Read Archive for whole-genome sequencing runs that could be reassembled to yield additional 16S rRNA sequences. Using riboSeed (a genome assembly tool for correcting rDNA misassembly), Focus16 provides a way to augment 16S rRNA databases with high-quality re-assembled sequences. In this study, we augmented the widely-used SILVA 16S rRNA database with the novel sequences disclosed by Focus16 and re-processed amplicon sequences from several benchmarking datasets with DADA2. Using this augmented SILVA database increased the number of amplicon sequence variants that could be assigned taxonomic annotations. Further, fine-scale classification was improved by revealing ambiguities. We observed, for example, that amplicon sequence variants (ASVs) may be assigned to a specific genus where Focus16-correction would indicate that the ASV is represented in two or more genera. Thus, we demonstrate that improvements can be made to taxonomic classification by incorporating these carefully re-assembled 16S rRNA sequences, and we invite the community to expand our work to augment existing 16S rRNA reference databases such as SILVA, GreenGenes, and RDP.

bioinformatics↗

bioRxiv2018

KH-type splicing regulatory protein controls colorectal cancer cell growth and modulates the tumor microenvironment

KH-type splicing regulatory protein (KHSRP) is a multifunctional nucleic acid binding protein implicated in key aspects of cancer cell biology: inflammation and cell-fate determination. However, the role KHSRP plays in colorectal cancer (CRC) tumorigenesis remains largely unknown. Using a combination of in silico analysis of large datasets, ex vivo analysis of protein expression in patients, and mechanistic studies using in vitro models of CRC, we investigated the oncogenic role of KHSRP. We demonstrated KHSRP expression in the epithelial and stromal compartments of both primary and metastatic tumors. Elevated expression was found in tumor versus matched normal tissue, and we validated these findings in larger independent cohorts in silico. KHSRP expression was a prognostic indicator of worse overall survival (HR=3.74, 95% CI = 1.43-22.97, p=0.0138). Mechanistic data in CRC cell line models supported a role of KHSRP in driving epithelial cell proliferation in both a primary and metastatic setting, through control of the G1/S transition. Additionally, KHSRP promoted a pro-angiogenic extracellular environment by regulating the secretion of oncogenic proteins involved in diverse cellular processes such as migration and response to cellular stress. Our study provides novel mechanistic insight into the tumor-promoting effects of KHSRP in CRC.

cancer biology↗