Unravelling the roles of a small membrane protein in DNA-damage repair and tolerance in Caulobacter crescentus

Carrion Sanabria, Alejandro (2023) Unravelling the roles of a small membrane protein in DNA-damage repair and tolerance in Caulobacter crescentus. Doctoral thesis, University of East Anglia.

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Abstract

Antibiotic resistance has become a major concern in public health in recent years. Bacteria harbour numerous mechanisms to bypass and resist antimicrobial compounds. DNA damage response in bacteria has been widely studied in species such as Escherichia coli but poorly investigated in other bacterial species. We aimed to study the DNA damage response in a different class of alphaproteobacteria with a distinct chromosome organisation and segregation pattern than E. coli. In this study, we report the discovery of a novel 37-amino-acid protein with an essential effect on maintaining cell fitness under DNA-damaging conditions in C. crescentus. CalP is a small amphipathic transmembrane protein with Nout-Cin topology that polymerises, generating a homooligomer in the inner cell membrane. Sensitivity assays confirmed the ΔcalP deficient phenotype in antibiotic-treated cells such as mitomycin C (MMC), norfloxacin, ciprofloxacin, or MMS. Complementation analysis confirmed the restoration of the WT phenotype in MMC or norfloxacin-exposed cells. DNA damage-induced cells with MMC caused calP downregulation in ΔrecA. In contrast, the calP expression in WT upregulated calP following the induction of DNA damage with MMC. Immunoblot assays showed that CalP production increased following cellular DNA damage with MMC, norfloxacin, or MMS. Nevertheless, despite MMC-induced DNA damage in the mutant cells, CalP production did not increase in ΔrecA. ΔcalP mutants were more permeable to MMC-TRC (MMC linked to the Texas Red Cadaverine fluorophore) than WT, suggesting that CalP may have an efflux pump-like function in the cell. Our results indicate that CalP could be involved in DNA damage tolerance/response in C. crescentus. However, elucidating the role and molecular mechanisms of CalP requires further investigation.

Item Type:	Thesis (Doctoral)
Faculty \ School:	Faculty of Science > School of Biological Sciences
Depositing User:	Chris White
Date Deposited:	20 Dec 2023 13:43
Last Modified:	20 Dec 2023 13:43
URI:	https://ueaeprints.uea.ac.uk/id/eprint/94021
DOI:

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