mathiaseder.bsky.social

🔥 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...

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.

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.

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.

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.

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.

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.

1 - Using our recently developed “hopping” approach, we relocate CBS-containing reporters and CBSs to thousands of genomic positions and reconstruct detailed regulatory landscapes.

mathiaseder.bsky.social reposted

Jesse Engreitz

@jengreitz.bsky.social

New preprint from our lab! What can we learn about the properties of gene regulatory elements by CRISPR’ing a random set of accessible sites in human cells? Find out here: www.biorxiv.org/content/10.1... 👇 1/

science.orgFunctional maps of a genomic locus reveal confinement of an enhancer by its target geneGenes are often activated by enhancers located at large genomic distances, and the importance of this positioning is poorly understood. By relocating promoter-reporter constructs into thousands of alt...

✨Exciting news: the main story of my PhD is out in Science! Together with Christine Moene @cmoene.bsky.social, we explored what happens when you scramble the genome—revealing how Sox2’s position shapes enhancer activation. 📖 Read the full story here: www.science.org/doi/10.1126/...

6/ Beyond Sox2, this technology opens the door to studying how genomic location shapes the activity of many different DNA elements in a high-throughput way. We believe it will be broadly useful for diverse biological questions. 🚀

nature.comInteractions between the genome and the nuclear lamina are multivalent and cooperative - Nature Structural & Molecular BiologyUsing a transposon-based approach to create a set of large genomic rearrangements, Dauban et al. demonstrate that interactions of lamina-associated domains with the nuclear lamina involve multiple con...

For another cool example of what “hopping” can uncover—this time in the context of lamina interactions—check out Lise’s paper: www.nature.com/articles/s41...

@cmoene.bsky.social @nkinl.bsky.social @oncodeinstitute.bsky.social @basvansteensellab.bsky.social

4/ Strikingly, deleting the endogenous Sox2 gene expands this landscape—yet the original Sox2 site remains a “sweet spot” for activation. Genes can thus act as gatekeepers, restricting their enhancer’s influence.

5/ We found that this competition is partly encoded in the Sox2 coding sequence (CDS) – a 1kb exon. Addition of the CDS to the reporter increases reporter expression and competition with the Sox2 gene. This effect seems to be mediated by the CDS-DNA sequence rather than the produced RNA or protein.

2/ By “loading” our transposon with a fluorescent Sox2 reporter, relocating it, and sorting cells by reporter expression, we generated high-resolution activity maps.

3/ These maps depict a sharply confined activation landscape. Interestingly, reporter expression is highest at the enhancer and around the endogenous gene whereas it turns silent upstream of the gene and downstream of the enhancer.