FKBP52 and Its Role of in the DNA Damage Repair

Abstract

Damage to genetic material represents a constant threat to genomic stability, with tens of thousands of DNA lesions being produced per day per cell. If DNA damage is not repaired correctly, it can be detrimental to the cell, leading to alterations in the genetic material and disease states such as cancer and neurodegenerative disorders. To protect the integrity of the DNA, cells have evolved a global signalling network known as the DNA damage response. This network detects different types of genotoxic stress to raise a versatile and coordinated response, which includes control of the cell cycle transitions, transcriptional processes and stimulation of DNA repair and apoptosis.Recently, the Richard Lab discovered the human single strand DNA binding protein 1 (hSSB1) to be a crucial component of the DNA damage signalling pathway.To increase our understanding of the function of hSSB1 in the DNA damage pathway, a connectivity screen was performed on hSSB1. The connectivity screen is a bioinformatic tool used to identify mRNA transcripts that are regulated in a similar manner as the bait (hSSB1). In this study, I sought to identify proteins that may also be involved in the DNA damage response (DDR) pathway, utilising hSSB1 as the bait.One of the transcripts identified from this screen was FKBP52. Based on the previously published literature, this protein has been linked to numerous cancers and in the regulation of other proteins involved in genome maintenance, such as p53, hTERT and NF-κB, making it a highly interesting candidate for further study. This study aims to determine if FKBP52 plays a role in the DDR and if it interacts or regulates hSSB1. To determine if FKBP52 and hSSB1 interact co-immunoprecipitation studies were performed as well as in vitro direct interaction assays. These showed that FKBP52 and hSSB1 might co-localise in a complex in vivo but that is was unlikely to be a direct interaction, leading to the theory that FKBP52 was potentially acting as a cochaperone. Supporting this proposition, was the in vivo observation that when FKBP52 levels were depleted by siRNA the corresponding levels of hSSB1 in the nucleus were reduced and showed little response to IR.FKBP52 expression was observed to responded to IR induced DNA damage.FKBP52 levels were also found to increase in the nucleus in response to IR and was observed to become chromatin associated. found to be bound to the nucleus, this suggesting it was involved in the DNA damage response. FKBP52 depletion also led to altered DNA damage signalling in particular with the phosphorylation of key regulatory proteins of the DNA damage pathway. It was also found that reduced levels via siRNA led to reduced survival after IR via clonogenic assays and reduced the efficiency of HR via the HR assay. FKBP52 levels in the nucleus were shown to be regulated by ATM and ATR consistent with this FKBP52 was found to be phosphorylated following IR on a putative ATM/ATR phosphorylation site. It was FKBP52 and its role in DNA Damage Repair iv found that phosphor...