Interaction of TRF2-Rap1 with telomeric chromatin in the context of aging and cancer

• Main Author: Woo, Jia Qi
• Other Authors: Lars Nordenskiöld
• Format: Final Year Project
• Language: English
• Published: Nanyang Technological University 2024
• Subjects: Medicine, Health and Life Sciences
• Online Access: https://hdl.handle.net/10356/178191
• Institution: Nanyang Technological University

Telomeric DNA form the ends of eukaryotic DNA. It is a tandem array of [TTAGGG]n, with a 3’ G-rich overhang. Telomeric nucleosomes form unique nucleoprotein structures with the shelterin complex consisting of 6 components; TRF1, TRF2, POT1, RAP1, TIN2, TPP1. Shelterin plays a crucial role in preventing inappropriate detection of telomeres as DNA breaks, and subsequent activation of damage repair pathways. Liquid-liquid phase separation induced by shelterin is proposed to be a mechanism underlying the dynamic behaviour of telomeres. Telomeres can switch between a protected and unprotected state to regulate specific access of factors to telomeres. Although the dynamic behaviour is integral to telomere integrity, little is understood about the behaviour. Here, we characterise the dynamic behaviour of biomolecular condensates formed by telomeric nucleosomes under the influence of the TRF2-Rap1 complex. Full length hTRF2 and hRap1 were purified to derive the TRF-Rap1 complex. Nucleosome arrays were reconstituted with 10 x 157 base pairs of [TTAGGG]n and human histone octamers. Confocal microscopy was employed to study condensate behaviour. Condensates exhibit dynamicity up to TRF2-Rap1: Nucleosome = 2. Condensates formed with increasing TRF2-Rap1 display progressive rigidity. Our studies demonstrate that varying stoichiometries of TRF2-Rap1 complex at telomeric nucleosomes mediate condensate dynamicity.