White collar 1‐induced photolyase expression contributes to UV ‐tolerance of Ustilago maydis

Abstract

Ustilago maydis is a phytopathogenic fungus causing corn smut disease. It also is known for its extreme tolerance to UV‐ and ionizing radiation. It has not been elucidated whether light‐sensing proteins, and in particular photolyases play a role in its UV‐tolerance. Based on homology analysis, U. maydis has 10 genes encoding putative light‐responsive proteins. Four amongst these belong to the cryptochrome/photolyase family (CPF) and one represents a white collar 1 ortholog (wco1). Deletion mutants in the predicted cyclobutane pyrimidine dimer CPD‐ and (6–4)‐photolyase were impaired in photoreactivation. In line with this, in vitro studies with recombinant CPF proteins demonstrated binding of the catalytic FAD cofactor, its photoreduction to fully reduced FADH − and repair activity for cyclobutane pyrimidine dimers (CPDs) or (6–4)‐photoproducts, respectively. We also investigated the role of Wco1. Strikingly, transcriptional profiling showed 61 genes differentially expressed upon blue light exposure of wild‐type, but only eight genes in the Δwco1 mutant. These results demonstrate that Wco1 is a functional blue light photoreceptor in U. maydis regulating expression of several genes including both photolyases. Finally, we show that the Δwco1 mutant is less tolerant against UV‐B due to its incapability to induce photolyase expression.

Two Photolyases Repair Distinct DNA Lesions and Reactivate UVB-Inactivated Conidia of an Insect Mycopathogen under Visible Light

“…S1B). Notably, CryD is absent in the genomes of C. militaris, M. acridum, and M. robertsii but has two homologs in U. maydis, as reported previously (26).…”

“…This speculation was verified by the levels of CPDs and 6-4PPs quantified from the UVB-impaired DNAs of both mutants and their photorepair activities against the two cytotoxic products. In B. bassiana, the main roles of Phr1 and Phr2 in the decomposition of CPDs and 6-4PPs are well shown by the high DNA repair activity of Phr1 in the Δphr2 mutant and Phr2 in the Δphr1 mutant, respectively, and are partially consistent with the reported roles of their homologs in B. cinerea (21), M. robertsii (27), T. atroviride (23)(24)(25), and U. maydis (26). In the present study, intriguingly, both 6-4PP and CPD levels were significantly higher in the UVB-irradiated cells of the Δphr1 mutant and Δphr2 mutant than of the control strains, respectively.…”

“…The 6-4PP photolyase Cry1, but not cry-DASH, was also reported to play a significant role in photorepair of UVC-induced DNA damage in T. atroviride (25). Four CPF members exist in Ustilago maydis, a phytopathogenic fungus highly resistant to UV and ionizing irradiation, and the fungal DNA damage repair is dependent on two photolyases (Phr1 and Phr2) rather than two cry-DASHs (Cry1 and Cry2) (26). In Botrytis cinerea, the CPD photolyase BcCRY1 has been characterized as the major enzyme of light-driven DNA repair, while the cry-DASH BcCRY2 serves as a repressor of conidiation under white and black/near-UV lights but is dispensable for photoreactivation (21).…”

Fungal sensing of host environment

“…Fungal light sensing receptors belong to the family of phytochromes (Blumenstein et al, 2005), cryptochromes or DNA photolyases (Berrocal-Tito et al, 2007;Bluhm and Dunkle, 2008). The light-sensing protein white collar 1 ortholog was recently shown to contribute to UV tolerance of U. maydis (Brych et al, 2016). In brief, fungi have sensing mechanisms to respond to physical cues such as light, gravity and surfaces.…”

Rhodobacter sphaeroides CryB is a bacterial cryptochrome with (6–4) photolyase activity

“…For in vitro repair assays of CryB and variants, substrate generation was done as described by Brych et al [42]. One substrate molecule contained 1 (6-4)PP and 4.6 CPDs on average.…”