Genetic interactions between protein phosphatases CaPtc2p and CaPph3p in response to genotoxins and rapamycin inCandida albicans

Abstract

In Saccharomyces cerevisiae cells, both of the two PP2C protein phosphatases ScPtc2p and ScPtc3p and the PP4 protein phosphatase ScPph3 are responsible for ScRad53p dephosphorylation after the DNA methylation agent methylmethane sulfonate (MMS)-induced DNA damage. In this study, we show that CaPtc2p is not required for the CaRad53p dephosphorylation during the recovery from DNA damage, as is CaPph3p in Candida albicans. However, deletion of CaPPH3 has an additive effect on the sensitivity of C. albicans cells lacking CaPTC2 to MMS and the DNA synthesis inhibitor hydroxyurea (HU). In addition, deletion of CaPPH3 promotes in vitro filamentation of C. albicans cells. Furthermore, mutation of CaPTC2 is epistatic to that of CaPPH3 in the sensitivity of C. albicans cells to rapamycin. Therefore, CaPtc2p and CaPph3p might play a role in the target of rapamycin (TOR) signaling in C. albicans cells.

Hof1 plays a checkpoint-related role in MMS-induced DNA damage response in Candida albicans

“…Given the Hof1-Rad53 association, we checked Rad53 behavior in a HOF1 deletion background. Previously, we reported that Rad53 is phosphorylated in response to MMS, resulting in a slower migration rate in SDS-PAGE (Feng et al, 2013). Here, we found that Rad53 was phosphorylated after MMS exposure in both WT and HOF1 deletion strains (Figure 6A).…”

“…To construct the PPH3 HOF1 double deletion strain, a previous PPH3 deletion strain (Feng et al, 2013) was used as a background strain and a similar CRISPR/Cas9 system for HOF1 was used. Similarly, a MMS21 deletion strain constructed before (Islam et al, 2019) was used as a background strain to construct the MMS21 HOF1 double deletion strain.…”

“…C. albicans yeast cells treated with either the DNA-replication inhibitor hydroxyurea (HU) or the DNA methylation agent MMS exhibit activation of the checkpoint kinase Rad53 accompanied by significant filamentous growth (Shi et al, 2007). Similarly, deletion of RAD52 causes a strong sensitivity to MMS and activates filamentous growth (Andaluz et al, 2006), and blocking the deactivation of Rad53 by deletion of PPH3 promotes filamentous growth and increased virulence (Sun et al, 2011;Feng et al, 2013Feng et al, , 2017. Deletion of RTT109, which plays critical roles in maintaining genome stability, results in fungal cells that are significantly less pathogenic in mice and more susceptible to killing by macrophages in vitro than wild-type (WT) cells (Lopes da Rosa et al, 2010).…”

The putative transcription factor CaRtg3 is involved in tolerance to cations and antifungal drugs as well as serum-induced filamentation inCandida albicans

“…All strains were routinely grown at 30°C in YPD medium (2% glucose, 2% peptone, 1% yeast extract) or SD medium (0.67% yeast nitrogen base without amino acids, 2% glucose, and auxotrophic amino acids as needed; Jiang et al, 2004). The following growth conditions were used to induce filamentation for C. albicans cells, YPD plus 10% fetal bovine serum (FBS; Sango Biotech, Shanghai, China), SLAD (0.17% yeast nitrogen base without amino acids and ammonium sulfate, 0.05 mM (NH4) 2 SO 4 , 2% glucose, pH 7.0), Spider (1% peptone, 1% mannitol, 0.2% K 2 HPO 4 ), and Lee's medium (Goyard et al, 2008;Feng et al, 2013). 5-Fluoroorotic acid (5-FOA) was purchased from Nuotai Chemicals Inc., Shanghai, China.…”

Cadmium-induced activation of high osmolarity glycerol pathway through its Sln1 branch is dependent on the MAP kinase kinase kinase Ssk2, but not its paralog Ssk22, in budding yeast

“…To investigate the phosphorylation state of Hog1, we introduced the plasmid YCp33-HOG1-GFP (YCp; URA3) (Jiang et al, 2004) into the wild-type BY4743 and its isogenic gene deletion mutants. Protein extraction and Western blot analysis was carried out as described (Jiang et al, 2004;Feng et al, 2013). Total proteins of HOG1-GFP were detected by a monoclonal antiGFP antibody purchased from Abcam Inc. (Hong Kong, China), and phosphorylated proteins of HOG1-GFP were detected by the phospho-p38 MAP kinase (Thr180/Tyr182) antibody purchased from Cell signaling (Product number: 9211).…”