Biology subjects

Elgood Hunt, E.

Publications and source records attributed to Elgood Hunt, E..

3 recordsLinked to original sources

bioRxiv2026

Co-option of a mouse-specific retrotransposon rewires Ash2l isoform usage to prime developmental promoters

How pluripotent stem cells pre-configure the chromatin landscape to enable timely activation of developmental gene programs remains a central question in developmental biology. Here we demonstrate that the core chromatin regulator ASH2L undergoes a developmentally regulated transcription start site (TSS) switch that generates two protein isoforms with distinct developmental functions: a full-length ASH2L containing an N-terminal intrinsically disordered region (IDR), and a conserved, truncated protein isoform lacking this region. In mouse pluripotent stem cells, a lineage-specific ERV-K retrotransposon drives the upstream TSS, which suppresses the downstream TSS through transcription-coupled SETD2-mediated histone H3 lysine 36 methylation. The resulting stem cell-specific truncated ASH2L assembles into COMPASS complexes and enhances histone H3 lysine 4 trimethylation at promoters governing early development. Despite being restricted to pluripotent stem cells, truncated ASH2L is required for proper axial patterning in gastruloids and for the transition of neural progenitors to motor neurons. This temporal decoupling, wherein a chromatin state established during pluripotency executes its function only after the establishing isoform disappears, defines an epigenetic priming mechanism for developmental timing. Thus, lineage-specific retrotransposon co-option reveals how a dedicated protein isoform configures the pluripotent epigenome for developmental transitions after its own expression has ceased.

molecular biology↗

bioRxiv2025

Mapping and quantifying nascent transcript start sites using TT-TSS-seq

Transcription initiation is a highly dynamic and tightly regulated process involving the coordinated action of transcription factors, chromatin remodelers, and RNA polymerase which determine where and when transcription begins. Accurately mapping and quantifying transcription start sites (TSSs) from nascently transcribed RNAs remains a key area of interest, as it provides critical insights into transcription dynamics. Here, we combined transient transcriptome sequencing with transcription start site sequencing (TT-TSS-seq) to accurately map and quantify transcription initiation sites from nascent transcripts. Since transient metabolic labelling yields low-input RNA, we optimized the TSS-seq protocol to enhance sensitivity and accuracy. Specifically, we refined enzymatic reactions for decapping and RNA ligation and incorporated 5 oligonucleotides containing unique molecular identifiers (UMIs) and barcodes to enable accurate quantification and sample multiplexing. The TT-TSS-seq approach detected transcription initiation of unstable transcripts, such as enhancer RNAs. Moreover, we identified that a large fraction of genes use multiple transcription initiation sites, yet often produce only a single stable transcript. Overall, TT-TSS-seq provides precise mapping and quantification of transcription initiation sites, offering new insights into transcriptional dynamics and expanding the toolkit for studying gene regulation.

molecular biology↗

bioRxiv2025

DNA supercoiling impacts alternative transcription start site selection in yeast

Most genes are transcribed from multiple transcription start sites (TSSs), defined as alternative TSSs, which are highly regulated and can lead to various gene regulatory outcomes including changes in translation efficiency and protein isoform expression. Transcription factors and chromatin regulators control alternative TSS selection. DNA supercoiling affects multiple aspects of transcription including transcription initiation; however, its regulatory effect on genes with multiple TSSs is not known. Here, we investigated how DNA supercoiling impacts alternative TSS usage in Saccharomyces cerevisiae. We depleted topoisomerases during early meiosis, where alternative TSS usage is prevalent, and applied an improved TSS sequencing protocol. We show that supercoiling affects alternative TSS usage at almost 600 genes. Increased alternative and aberrant TSS usage was observed near and within open reading frames, likely resulting from transcription-induced supercoiling originating from upstream alternative TSSs. DNA supercoiling had the greatest impact on genes with a dominant alternative TSS, significant spacing between alternative TSSs, and greater overall gene length. Our results establish that DNA supercoiling release during transcription is critical for correct TSS selection.

molecular biology↗