7 - Huge thanks to my colleague and friend @cmoene.bsky.social and Marantha Kaagman for their major contributions to this work, and to @bracciolilab.bsky.social and @dewitlab.bsky.social for their support.
6 - These were just some highlights of our recent preprint, if you want to know more or dig deeper you can find the paper on bioarchive.
5 - We also looked at the effect of inserting CBS sites (without promoter) in the locus. Between gene and enhancer, CBS insertions consistently reduce Sox2 expression in an orientation-dependent manner, further suggesting a polarity in the loop extrusion process.
4 - Inverting only the CBS upstream of the reporter did not recapitulate the effects that we observed when the entire CBS–promoter construct was inverted, indicating that reporter activation depends on the combined orientation of the CBS and the promoter.
3 - We found that a CBS that is naturally located upstream of the Sox2 promoter endows this P with a strong orientation-dependent activation, anywhere within the gene-enhancer interval. Adding three CBSs enhances the orientation bias, suggesting a quantitative effect of CTCF on regulatory output.
2 - We previously designed a Sox2 reporter containing the Sox2 promoter followed by a blue fluorescent protein (mTurquoise2). To test the effect of CBSs on the reporter, we added CBSs upstream of the promoter in either the forward or reverse motif orientation.
1 - Using our recently developed “hopping” approach, we relocate CBS-containing reporters and CBSs to thousands of genomic positions and reconstruct detailed regulatory landscapes.
🔥 How does CTCF shape enhancer–promoter communication?
In our new preprint, we systematically test how the position and orientation of CTCF binding sites (CBSs) influence gene regulation at the mouse Sox2 locus.
🧪 full paper can be found here: www.biorxiv.org/content/10.6...