Role of homologous recombination genesRAD51,RAD52, andRAD59in the repair of lesions caused by γ-radiation to cycling and G2/M-arrested cells ofCandida albicans

Abstract

We have analysed the role of homologous recombination (HR) genes on the repair of double-strand breaks induced by γ-ionising radiation in Candida albicans. Depletion of either CaRad51 or CaRad52 caused a dramatic drop in the number of survivors compared with wild type, whereas depletion of CaRad59 caused a moderate decrease. Besides, compared with Saccharomyces cerevisiae, C. albicans relies more on HR proteins for repair of ionising radiation lesions. Pulse-field electrokaryotypes of survivors identified genetic alterations mainly in the form of aneuploidy in HR mutants and chromosome length polymorphism and ectopic translocation in wild type. Increasing irradiation (4 to 80 krad) of both cycling and nocodazole-treated (G2/M-arrested) cells revealed a gradual loss of chromosomes, larger chromosomes being more affected than smaller ones. For cycling wild-type cells, shattered chromosomes were progressively restored following incubation in yeast extract, peptone, dextrose medium, but not in phosphate-buffered saline, and this accompanied by a moderate increase in colony-forming units, suggesting that repair was followed by replication of survivors. Irradiated G2/M arrested cells from wild type but not from HR mutants partially restored the chromosome ladder following incubation (4-8 hr) in yeast peptone dextrose-nocodazole. However, HR mutants showed a chromosome shattering pattern similar to wild type, an indication that lesions other than double-strand breaks, likely single-strand break, are responsible for their drastically reduced survivability.

RAD51–WSS1-dependent genetic pathways are essential for DNA–protein crosslink repair and pathogenesis in Candida albicans

“…Rad51 plays a central role in the homology search and catalyzes strand exchange between the recombination partners ( 14 , 15 , 16 ). As compared with the parental strain, rad51 null strain of C. albicans exhibits a filamentous morphology, decreased growth rate, and weak sensitivity to UV light, oxidizing agents, and compounds that cause double-strand breaks ( 17 ). Except HR, other DNA repair pathways such as BER and NER in C. albicans do not seem to be actively operating to take care of DNA lesions, and other backup pathways could be more pronounced in C. albicans than in S. cerevisiae .…”

Heterologous Single-Strand DNA-Annealing and Binding Protein Enhance CRISPR-Based Genome Editing Efficiency in Komagataella phaffii

“…14,15 We introduced SSAPs and SSB in pairs into K. phaffii to overcome the limitations of the endogenous DNA repair pathway mediated by eukaryotic proteins, such as RAD51, to promote genome editing in K. phaffii. 16,17 In this study, we aimed to develop a host-independent and highly efficient gene editing tool for K. phaffii using short homology arms by introducing microbial SSAPs and SSBs. Initially, we evaluated the effects of various microbial SSAPs, including LBet (from Escherichia phage λ), gp2.5 (from Bacteriophage T7), PapRecT (from a Pseudomonas aeruginosa phage), and CspRecT (from a Collinsella stercoris phage), on genome integration efficiency and colony forming unit numbers.…”

“…phaffii to overcome the limitations of the endogenous DNA repair pathway mediated by eukaryotic proteins, such as RAD51 , to promote genome editing in K. phaffii. , …”

Transcriptional Profiling of the Candida albicans Response to the DNA Damage Agent Methyl Methanesulfonate

“…Moreover, DNA damage repair-related genes also contribute to the pathogenicity of C. albicans cells. Losing the HR element Rad52 and the NHEJ ligase Lig4 impair the pathogenicity of C. albicans cells [ 27 , 28 ]. As well, the protein phosphatase Pph3 contributes to the dephosphorylation of checkpoint Rad53 and plays a negative role in virulence, probably depending on its role in checkpoint-related morphology regulation [ 29 , 30 ].…”