Abstract
dc:description.abstractCurrent methods for profiling histone modifications and protein localisation along chromatin suffer from critical limitations: the gold standard, ChIP-seq, relies on high-quality antibodies, while other antibody-free techniques like DamID are limited by the availability of specific DNA sequences. Here, I describe the development of ChILI-seq (Chromatin Interactions by Lentiviral Integration Sequencing), an antibody-free and sequence-independent method to profile the genomic localisation of proteins, as well as DNA and histone modifications. ChILI-seq exploits the dependency of the lentiviral integrase on LEDGF (Lens Epithelium-Derived Growth Factor). By fusing LEDGF to a chromatin-binding protein of interest, lentiviral integrations are redirected to the genomic targets of that protein, and can be identified through integration site mapping and next-generation sequencing. To validate ChILI-seq, I demonstrate its utility to profile histone and DNA modifications using known chromatin reader domains. I show that ChILI-seq is a cost-effective alternative to traditional techniques, providing superior resolution and signal-to-noise ratios. I then go on to show that ChILI-seq can reveal the binding preferences of many chromatin reader domains, allowing us to dissect the recruitment requirements of key epigenetic modifiers, including BRD4 and the HUSH complex. Finally, I utilise a high-throughput screening approach to identify chromatin-binding domains of unknown specificities and exploit ChILI-seq to reveal their binding preferences. This screen suggested an intriguing new role for the KAT14 acetyltransferase in LINE-1 element regulation. In summary, ChILI-seq expands our chromatin profiling toolkit, offering a versatile, antibody-free and sequence-independent method to probe interactions between epigenetic modifiers and chromatin.
Degree
thesis:*- Name dc:type.qualificationname
- Doctor of Philosophy (PhD)
- Level dc:type.qualificationlevel
- Doctoral
- Grantor dc:publisher.institution
- University of Cambridge
- Year dc:date.issued
- 2025
Author and committee
dc:creator, dc:contributor.*- Author dc:creator
-
- Qin, Chuyan
- Advisor dc:contributor.advisor
-
- Tchasovnikarova, Iva
Subjects
dc:subject × 4Rights
dc:rightsIdentifiers
dc:identifier.*- DOI dc:identifier.doi
- https://doi.org/10.17863/CAM.124872
- OAI identifier oai:identifier
- oai:www.repository.cam.ac.uk:1810/395327