The roles of YBX1 in intracellular and intercellular DNA damage response

Thesis

RNA-dependent DNA damage response (DDR) is an emerging field, focusing on the influence of RNA species on DNA repair. A multifunctional RNA and DNA-binding protein, YBX1 is under study as a player in RNA-dependent DDR which localises to DNA breaks and interacts with repair machinery. YBX1 also acts as a regulator of packaging small RNAs into exosomes for intercellular communication. Exosomes secreted from ionising radiation (IR)-treated cells have altered protein and RNA content which, upon transfer to neighbouring cells, can stimulate bystander effects and influence population-wide responses to damage. Using exosome transfer experiments, I observed reduced expression of DNA damage markers in irradiated recipient cells that were pre-incubated with radiation-derived exosomes, signifying a radioprotective bystander response which is dependent on YBX1. The analysis of small RNA sequencing of exosomes derived from wild-type and YBX1 knockout cells revealed that specific 30 nucleotide-long fragments derived from Y1 and Y3 RNA (ysRNA) are significantly upregulated in radiation-derived exosomes, in a YBX1-dependent manner. Treating cells with a combination of these fragments can recapitulate the radioprotective bystander phenotype observed via exosome transfer, suggesting they play a presently uncharacterised role in DNA repair. I further investigated their molecular mechanism and suggest that ysRNA localises to the nucleus and becomes modified by NSUN2 and PARP1 upon DNA damage, facilitated by ysRNA binding to YBX1. This may aid in DNA repair by influencing PARP1 auto-modification, thus modulating the occupancy of PARP1 at DSBs and highlighting a new role for these under-studied species of small non-coding RNA in DNA repair. I further show that modulation of YBX1 may influence cell sensitivity to PARP inhibitors, thus representing a potential therapeutic target. Overall, I present a dual role for YBX1 in intracellular and intercellular DDR that may have therapeutic implications for cancer treatment.

Authors: Shaw, A

• DOI: 10.5287/ora-8nd1k0kme
• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford
• Language: English
• Keywords: exosomes; DNA damage; bystander effect; small non-coding RNA; YBX1
• Subjects: DNA Damage; DNA Repair