Biology subjects

Mawer, J. S. P.

Publications and source records attributed to Mawer, J. S. P..

2 recordsLinked to original sources

bioRxiv2026

Protocol for in vivo DNA-RNA hybrid immunoprecipitation sequencing and analysis from frozen mammalian tissues

DNA-RNA hybrids (R-loops) form transiently on the genome and regulate cellular homeostasis. They also influence genome editing outcomes, highlighting their therapeutic potential in vivo. This protocol enables high-resolution mapping of DNA-RNA hybrids directly from frozen mouse tissues. Following tissue homogenisation and lysis, genomic DNA is extracted, digested and DNA-RNA hybrids are isolated using the hybrid-specific S9.6 monoclonal antibody. The purified hybrids are then processed for whole-genome sequencing to generate R-loop profiles. For complete details on the use and execution of this protocol, please refer to Puzzo, Crossley et al1. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=188 SRC="FIGDIR/small/716701v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@3d8f86org.highwire.dtl.DTLVardef@199cd84org.highwire.dtl.DTLVardef@83c51eorg.highwire.dtl.DTLVardef@1024332_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOgraphical abstractC_FLOATNO C_FIG

molecular biology↗

bioRxiv2020

The RNA-binding protein Puf5 buffers mRNA levels against chromatin-mediated changes in nascent transcription

Gene expression is a dynamic process regulated at all stages, starting with opening of chromatin, transcription, and continuing with mRNA export, translation and, finally, degradation. While there are feedback mechanisms within the system, it is not clear whether these extend to crosstalk between chromatin architecture and mRNA decay. Here, we show that changes in nascent transcription, mediated by mutating H3K56 to alanine, are post-transcriptionally buffered by the Pumilio protein Puf5, which stabilizes transcripts in a context-dependent manner. Depleting Puf5 in an H3K56A background leads to synthetic lethality. This genetic interaction can be explained by a decrease in translation due to downregulation of its direct mRNA targets, largely consisting of ribosomal protein genes. Importantly, we show that this post-transcriptional buffering is not only linked to H3K56A, but may be a more widespread phenomenon that also buffers against an increase in nascent RNA transcription in order to maintain physiological mRNA levels and cellular homeostasis.

molecular biology↗