Recent studies suggest that LSH and DDM1 are also critical for genome integrity, but their precise functions are not understood.
Similarly, the plant protein is required for development, and both proteins are important for regulating levels of DNA methylation, an important epigenetic mark. In mammals, the LSH enzyme is required for normal development, as well as oogenesis, spermatogenesis and T-lymphocyte proliferation. Mammalian LSH (Lymphoid-specific helicase also known as HELLS, PASG, and SMARCA6) and Arabidopsis DDM1 (Decreased DNA methylation 1) are the founding members of the LSH/DDM1 subfamily of ATP-dependent chromatin remodelers. It is therefore important for cells to regulate chromatin structure so that the genome can function properly. Chromatin packaging often inhibits enzymes that need to access the genetic information. Inside cells, eukaryotic DNA exists in a highly packaged structure called chromatin. Overall, our study provides important new information about the functions of the LSH/DDM1 family of enzymes. Together, our findings suggest that MUS-30, an LSH/DDM1 homolog, is required to prevent DNA damage arising from toxic base excision repair intermediates. We found that MUS-30-deficient cells are not defective for DSB repair, and we observed a negative genetic interaction between Δ mus-30 and Δ mei-3, the Neurospora RAD51 homolog required for homologous recombination.
DNA damage-sensitivity of Δ mus-30 is partially suppressed by deletion of methyl adenine glycosylase-1, a component of the base excision repair machinery (BER) however, the rate of BER is not affected in Δ mus-30 strains. Deletion of wdr76 rescued DNA damage-hypersensitivity of Δ mus-30 strains, demonstrating that the MUS-30-WDR76 interaction is functionally important. MUS-30 co-purifies with Neurospora WDR76, a homolog of yeast Changed Mutation Rate-1 and mammalian WD40 repeat domain 76. MUS-30 is a nuclear protein, consistent with its predicted role as a chromatin remodeling enzyme, and levels of MUS-30 are increased following DNA damage. We show that MUS-30-deficient cells have normal DNA methylation, but are hypersensitive to DNA damaging agents. We report that the Neurospora LSH/DDM1 enzyme is encoded by mutagen sensitive-30 ( mus-30), a locus identified in a genetic screen over 25 years ago. The filamentous fungus Neurospora crassa is a tractable system that encodes a single LSH/DDM1 homolog (NCU06306). LSH/DDM1 enzymes are required for DNA methylation in higher eukaryotes and have poorly defined roles in genome maintenance in yeast, plants, and animals.
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