Genetic analysis of the Neurospora crassa RAD14 homolog mus-43 and the RAD10 homolog mus-44 reveals that they belong to the mus-38 pathway of two nucleotide excision repair systems

Abstract

Genetic analysis of the Neurospora crassa RAD14 homolog mus-43 and the RAD10 homolog mus-44 reveals that they belong to the mus-38 pathway of two nucleotide excision repair systemsMasahito Sato, Takaharu Niki, Takeru Tokou, Kouhei Suzuki, Makoto Fujimura and Akihiko Ichiishi * Sciences, Toyo University, Itakura, Oura-gun, Gunma 374-0193, Japan (Received 30 October 2007, accepted 19 November 2007 Saccharomyces cerevisiae Rad14 and Rad10 proteins are essential for nucleotide excision repair (NER). Rad14 is a UV-damaged DNA binding protein and Rad10 is a structure-specific endonuclease that functions in a complex with Rad1. In this study, we identified and characterized the RAD14 and RAD10 homolog genes in Neurospora crassa, which we named mus-43 and mus-44, respectively. Disruption of mus-43 and mus-44 conferred sensitivity to UV and 4-nitroquinoline 1-oxide, but not to methyl methanesulfonate, N-methyl-N'-nitro-N-nitrosoguanidine, camptothecin, hydroxyurea, or bleomycin. The mus-44 mutant was more sensitive to UV than the mus-43 mutant. Genetic analysis indicated that mus-43 and mus-44 are epistatic to mus-38 which is a homolog of the S. cerevisiae RAD1, but not to mus-18 which belongs to a second excision repair pathway. Immunological assays demonstrated that both mus-43 and mus-44 retained the ability to excise UV-induced cyclobutane pyrimidine dimers and 6-4 photoproducts, but that excision ability was completely abolished in the mus-43 mus-18 and mus-44 mus-18 double mutants. These double mutants exhibited extremely high sensitivity to UV. In mus-43 and mus-44 mutants, the UV-induced mutation frequency increased compared to that of the wild-type. The mus-44 mutants also exhibited a partial photoreactivation defect phenotype similar to mus-38. These results suggest that both mus-43 and mus-44 function in the mus-38 NER pathway, but not in the mus-18 excision repair pathway. Faculty of Life

A partial photoreactivation defect phenotype is not due to unrepaired ultraviolet-induced pyrimidine dimers in ultraviolet-sensitive mutants of <i>Neurospora crassa</i>

“…These results indicate that most of the CPD repair activity during photoreactivation for 60 min depends on the photolyase. These results, together with a previous report (Sato et al, 2008), suggested that some cause of functional loss of MUS-44 protein resulted in the PPD phenotype.…”

“…1B). These results were similar to mutants produced by repeat-induced point mutation (Sato et al, 2008). Furthermore, to evaluate the photoreactivation ability of these strains, a photoreactivation assay was performed in which conidia were irradiated with visible light for 60 min after irradiation with UV light of each dose.…”

“…The Δmus-44 strain shows the PPD phenotype The mus-43 and mus-44 genes in N. crassa are homologs of S. cerevisiae RAD14 and RAD10, respectively. Their gene products are considered to be involved in NER, even in N. crassa, as mutants of these genes are sensitive to UV and 4-nitroquinoline 1-oxide (4NQO) and exhibit an epistatic relationship with the RAD1 homolog mus-38 (Sato et al, 2008). In this study, deletion strains of these genes produced by the KO project of the Fungal Genetics Stock Center were used.…”

“…In many organisms, these types of damage are repaired by nucleotide excision repair (NER), and the repair mechanism is highly A partial photoreactivation defect phenotype is not due to unrepaired ultraviolet-induced pyrimidine dimers in ultraviolet-sensitive mutants of Neurospora crassa conserved from bacteria to higher eukaryotes. NERrelated genes have been isolated in Neurospora crassa, such as mus-38 (a homolog of Saccharomyces cerevisiae RAD1), mus-40 (RAD2), mus-43 (RAD14) and mus-44 (RAD10) (Hatakeyama et al, 1998;Sato et al, 2008;S. Hatakeyama et al, unpublished data).…”

Effect of DNA Repair Deficiencies on the Cytotoxicity of Drugs Used in Cancer Therapy - A Review

“…265], deficiencies of other proteins do not change its activity or even reduce it [68,165,188,227,262]. Deficiencies in the BER, MMR and DRR pathways seem to increase the cytotoxicity of hydroxyurea [158,235,266], and defects in NER, FA and TLS do not seem to have a marked effect in its cytotoxicity [267][268][269]. With regard to the other drugs compiled in Table 7, deficiencies in BER increase the cytotoxicity of bortezomib and tretitoin [270][271][272] and decrease that of L-asparaginase [273], deficiencies in NER increase the cytotoxicity of arsenic trioxide [274,275] but decrease that of thalidomide [276], and deficiencies in MMR do not modify the cytotoxicity of L-asparaginase and vorinostat [277,278].…”