Boyes, J. | bioRxiv Science

bioRxiv2023

YY1 Binding to Regulatory Elements that Lack Enhancer Activity Promotes Locus Folding and Gene Activation

Enhancers activate their cognate promoters over huge distances but how enhancer/promoter interactions become established is not completely understood. There is strong evidence that cohesin-mediated loop extrusion is involved but this does not appear to be a universal mechanism. Here, we identify an element within the mouse immunoglobulin lambda (Ig{lambda}) light chain locus, HSC{lambda}1, that has characteristics of active regulatory elements but lacks intrinsic enhancer or promoter activity. Remarkably, knock-out of the YY1 binding site from HSC{lambda}1 reduces Ig{lambda} transcription significantly and disrupts enhancer/promoter interactions, even though these elements are >10 kb from HSC{lambda}1. Genome-wide analyses of mouse embryonic stem cells identified 3503 similar YY1-bound, putative genome organizing elements that lie within CTCF/cohesin loop boundaries but that lack intrinsic enhancer activity. We suggest that such elements play a fundamental role in locus folding and in facilitating enhancer/promoter interactions. HighlightsHow long-range enhancer-promoter interactions are established is not fully understood An element in the lambda light chain locus, HSC{lambda}1, lacks intrinsic enhancer activity Removal of YY1 binding from HSC{lambda}1 disrupts neighbouring enhancer/promoter contacts Genome-wide analyses detect similar elements that lack enhancer or promoter activity We propose these elements aid locus folding and nearby enhancer-promoter interactions O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=64 SRC="FIGDIR/small/554459v1_ufig1.gif" ALT="Figure 1"> View larger version (14K): org.highwire.dtl.DTLVardef@562875org.highwire.dtl.DTLVardef@7a1a85org.highwire.dtl.DTLVardef@634652org.highwire.dtl.DTLVardef@19ddfda_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2022

An Ultralong Bovine CDRH3 that Targets a Conserved, Cryptic Epitope on SARS-CoV and SARS-CoV-2

The ability of broadly neutralising antibodies to target conserved epitopes gives them huge potential as antibody-based therapeutics, particularly in the face of constant viral antigen evolution. Certain bovine antibodies are highly adept at binding conserved, glycosylated epitopes, courtesy of their ultralong complementarity determining region (CDR)H3. Here, we used a SARS-naive, bovine ultralong CDRH3 library and mammalian cell display, to isolate a bovine paratope that engages the SARS-CoV and SARS-CoV-2 receptor-binding domain (RBD). This neutralises viruses pseudo-typed with SARS-CoV Spike protein but not by competition with RBD binding to ACE2. Instead, using differential hydrogen-deuterium exchange mass spectrometry and site-directed mutagenesis, we demonstrate that this ultralong CDRH3 recognises a rarely identified, conserved, cryptic epitope that overlaps the target of pan-sarbecovirus antibodies (7D6/6D6). The epitope is glycan-shielded and becomes accessible only transiently via inter-domain movements. This represents the first bovine anti-sarbecovirus paratope and highlights the power of this approach in identifying novel tools to combat emerging pathogens.

molecular biology↗