Genotoxic stress causes DNA damage or stalled DNA replication and filamentous growth in the pathogenic fungus Candida albicans. The DNA checkpoint kinase Rad53 critically regulates by phosphorylation effectors that execute the stress response. Rad53 itself is activated by phosphorylation and inactivated by dephosphorylation. Previous studies have suggested that the phosphatase Pph3 dephosphorylates Rad53. Here, we used mass spectrometry and mutagenesis to identify Pph3 dephosphorylation sites on Rad53 in C. albicans. We found that serine residues 351, 461 and 477, which were dephosphorylated in wild-type cells during the recovery from DNA damage caused by methyl methanesulfonate (MMS), remained phosphorylated in pph3Δ/Δ cells. Phosphomimetic mutation of the three residues (rad53-3D) impaired Rad53 dephosphorylation, exit from cell cycle arrest, dephosphorylation of two Rad53 effectors Dun1 and Dbf4, and the filament-to-yeast growth transition during the recovery from MMS-induced DNA damage. The phenotypes observed in the rad53-3D mutant also occurred in the pph3Δ/Δ mutant. Together, our findings reveal a molecular mechanism by which Pph3 controls DNA damage response in C. albicans.
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Cover Image
Cover Image
Structure of O-acetylserine sulfhydrylase (OASS) from Brucella abortus compared with all known OASS structures. The high degree of variation is observed in N-terminal domain, which determined the size of active site cleft and responsible for interactions with Serine acetyl Transferase. The co-factor Pyridoxal phosphate (PLP) is shown in ball & stick model in the active site. For more information, please see study by Dharavath et al. in this issue, pages 1221–1239. Image provided by Samudrala Gourinath.
Characterization of Pph3-mediated dephosphorylation of Rad53 during methyl methanesulfonate-induced DNA damage repair in Candida albicans
Guangyin Yao, Junhua Wan, Qizheng Liu, Chunhua Mu, Yue Wang, Jianli Sang; Characterization of Pph3-mediated dephosphorylation of Rad53 during methyl methanesulfonate-induced DNA damage repair in Candida albicans. Biochem J 1 April 2017; 474 (7): 1293–1306. doi: https://doi.org/10.1042/BCJ20160889
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