Mechanisms underpinning recruitment of the HUSH-MORC2 corepressor to transposable elements in neural progenitor cells

Abstract

The HUSH-MORC2 corepressor has emerged as a key player in epigenetic silencing of L1 retrotransposons and other mobile genetic elements during human development. In the ‘canonical’ mode of its recruitment to chromatin, HUSH makes interactions with nascent transcripts from target elements, leading to the assembly of SETDB1, which deposits the repressive heterochromatin mark H3K9me3, and chromatin remodeler MORC2. Alternatively, it has been reported that in certain contexts, HUSH and its effectors can be recruited ‘non-canonically’ by DNA-binding transcription factor ZNF638. However, the function of ZNF638 and the mechanism underpinning recruitment of the HUSH-MORC2 corepressor during human brain development remain poorly understood. To address these gaps, webfirst made two technical advances: we optimized a CUT&RUN protocol to profile MORC2 chromatin binding and implemented a new bioinformatic analysis pipeline to efficiently analyze CUT&RUN epigenome profiling data. Using these tools, we assessed the ‘canonical’ recruitment mode by measuring MORC2 chromatin binding in the presence and absence of DNA methyltransferase DNMT1, known to control nascent transcription of L1 retrotransposons. Our results indicate that MORC2 targets young, full-length L1s following loss of DNA methylation. To investigate the ‘non-canonical’ recruitment of HUSH-MORC2 by ZNF638, and test whetherZNF638 has a role in the silencing of endogenous elements, we used CUT&RUN to study changes in chromatin marks H3K4me3 and H3K9me3 following CRISPRi knockdown of the ZNF638gene. These results suggest that ZNF638 is active at a subset of HUSH-MORC2-regulated retrotransposons and thus acts as a functional epigenetic regulator in neural progenitor cells. These findings increase our functional understanding of MORC2 and ZNF638 in human brain development.

Popular Abstract

HUSH little transposon, don’t say a word: how the Human Silencing Hub defends us from ‘jumping genes’

Approximately 50% of our DNA is made from transposons, sequences that can jump around the genome like parasites. When they do so, they can disrupt the function of genes, thus leading to disease. For example, transposons have been associated with various brain disorders. Therefore, our cells have developed a variety of ways to fight against these threats and protect our genomes. In our work, we are particularly interested in the defense against these elements in the developing human brain.

One of the defenders against transposons is the human silencing hub (HUSH) protein complex. To protect us, HUSH has to be recruited to the parasitic sequences, but it is unknown how this happens in the brain. This is important, because changes in HUSH function have been linked to neurodevelopmental disorders, indicating an important function in brain health. Two modes of recruitment have been proposed. In the first, the complex senses when the transposons are switched on and are producing a ‘transcript’ molecule. This in turn leads to the recruitment of other proteins, including one called MORC2, to switch the sequence off again. In the second, HUSH is recruited by a DNA-binding protein called ZNF638. This protein is known to be important in defending against viruses, but it has an unknown role in the brain.

In this project, we set to unravel how HUSH is recruited to fight against transposons in neural progenitor cells, i.e., cells that can give rise to neurons, that shape our brain. To this end, we implemented a computational analysis pipeline that allowed us to analyze data obtained from the experiments included in the project. We employed a technique called CUT&RUN, that helps us investigate the DNA regions that proteins interact with. The optimization of a CUT&RUN protocol allowed us to profile the genomic locations where MORC2 and HUSH are recruited. We also investigated how the absence of DNA methylation, a chemical change of the DNA associated with the repression of transposons, affects this recruitment. Our observations suggest that when DNA lacks this chemical modification, young elements a family of transposons known as LINE1 are reactivated and targeted by HUSH-MORC2. To study the role of ZNF638 in the fight of HUSH and MORC2 against transposons during human brain development, we inhibited its expression in the neural progenitor cells. We then used CUT&RUN to look for regions in the DNA where specific chemical changes in the histones, proteins that DNA wraps around, occurred. Our preliminary results showed that histone changes that are indicative of gene activation or repression occurred following the inhibition of ZNF638. This suggests that the role of ZNF638 could be associated with regulation of genes during brain development. Notably, we also observed that it is active at a subset of regions that HUSH and MORC2 target and could thus work along with them to attack transposons. These results shed light to the way HUSH is recruited to repress transposons and open the way for future work aiming to fully understand how this intricate system safeguards our genome.


Master’s Thesis Project in Molecular Biology, 60 credits, September 2023
Department of Biology, Lund University
Supervisor: Dr. Christopher Douse, Laboratory of Epigenetics and Chromatin Dynamics, Department of Experimental Medical Science, Lund University

• author: Koutounidou, Symela
• supervisor: Christopher Douse
• organization: Degree Projects in Molecular Biology
• course: MOBN03 20222
• year: 2022
• type: H2 - Master's Degree (Two Years)
• subject: Biology and Life Sciences
• language: English
• id: 9140404
• date added to LUP: 2023-10-24 17:09:44
• date last changed: 2023-10-24 17:09:44