TLS dependent and independent functions of DNA polymerase eta (Polη/Rad30) from Pathogenic Yeast Candida albicans

Abstract

Polη, a unique TLS DNA polymerase that promotes efficient bypass of UV-induced CPDs and cisplatin adducts, has not been explored in Candida species yet. Here, we show that CaPolη plays a vital role in protecting Candida albicans genome from diverse array of DNA damaging agents, not limited to UV and cisplatin. Polη deficient strain did not exhibit any hyphal development in the presence of UV and cisplatin while the wild type strain profusely developed DNA damage induced filamentation. The polarized growth induced by HU and MMS was found to be Polη independent. No common regulatory pathway of morphogenesis operates in C. albicans due to genomic stress, rather Polη branches away from RAD53 dependent pathway to be specific to UV/cisplatin. Interestingly, serum that does not inflict any DNA damage also induces hyphal growth in C. albicans, and requires a functionally active Polη. Importantly, deletion of RAD30 sensitized the strain to amphotericin B; but its presence resulted in azole drug tolerance only in DNA damaging conditions. We suggest that the roles of CaPolη in genome stability and genotoxins induced filamentation are due to its TLS activities; whereas its TLS independent functions play a vital role in serum induced morphogenesis and amphotericin B resistance.

Virulence and pathogenicity of aCandida albicansmutant with reduced filamentation

“…Thus, the involvement of TLS that maintains genome stability in extreme genomic stress would become essential for C. albicans survival, colonisation, and in the development of systemic candidiasis in the host. As reported earlier (Manohar et al, ), in this study as well, we found that serum or spider media constituents do not inflict any DNA damage and induce hyphal growth in C. albicans , but it requires a functionally active Polη. Additionally, we observed that Polη does not have a regulatory role in serum‐induced cell proliferation, but it does have an important role in the expression of certain virulent genes.…”

“…In our earlier study, we observed that the wild‐type strain of C. albicans switches from yeast cell to filamentous morphology on agar plates in response to genotoxic stress inflicted by HU, MMS, H 2 O 2 , UV, and cisplatin (Manohar et al, ). However, deletion of RAD30 drastically reduced such transition induced by UV/cisplatin but retained filamentation in the presence of HU, MMS, and H 2 O 2 .…”

“…However, significantly less percentage of macrophages interacted with treated wild‐type C. albicans than the untreated (Figure S2c). This partial inhibition could be due to hyphal transition in treated wild‐type C. albicans (Manohar et al, ), and macrophages may not have been able to interact with such large structures. This analysis suggests that macrophages exhibit differential affinity for different strains of C. albicans and vice versa, and the prior hyphal transition could inhibit interaction with macrophages to a certain extent.…”

“…(d) A polymerase chain reaction of genomic DNA isolated from six random C. albicans colonies from above to confirm the genotypes where absence or presence of RAD30 gene was verified. Due to deletion, a doublet truncated open reading frames are observed in rad30Δ but a single open reading frame band in WT (Manohar et al, ). A statistical significance of P < .05 was tested with unpaired Student's t test…”

“…As C. albicans exhibits considerable genotypic variation both in vitro and in clinical isolates due to genetic alterations (Ahmad et al, ; Ciudad, Hickman, Bellido, Berman, & Larriba, ; Rustchenko, ), DNA polymerases (Pol) could play a crucial role in fungal pathogenesis. Recently, we have characterized C. albicans Polη (Rad30), a Y‐family DNA polymerase, which promotes efficient bypass of ultraviolet (UV)‐induced cyclobutane pyrimidine dimers and cisplatin adducts (Acharya et al, ; Manohar, Peroumal, & Acharya, ; Satpati, Manohar, Acharya, & Dixit, ). In our earlier study, we showed that the roles of CaPolη in genome stability, genotoxins‐induced filamentation, and azole drug tolerance are due to its translesion DNA synthesis (TLS) activities, whereas its TLS‐independent functions play a pivotal role in serum‐induced morphogenesis and amphotericin B resistance.…”