The Mechanism of Small Interfering RNA Biogenesis in Neurospora Crassa
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RNA interference is a well-conserved post-transcriptional gene silencing mechanism that regulates various biological processes including development, genome defense, and heterochromatin formation. In filamentous fungus Neurospora crassa, quelling is an RNAi-related phenomenon that post-transcriptionally silences repetitive DNA and transposon. We previously identified a type of DNA damage-induced small RNA called qiRNAs that originate from ribosomal DNA. Ribosomal DNA cluster remains the only highly repetitive sequences in Neurospora genome. To understand how small RNAs are generated from repetitive DNA, we carried out a genetic screen to identify genes required for qiRNA biogenesis. Factors directly involved in homologous recombination (HR) and chromatin remodeling factors required for HR are essential for qiRNA production. HR is also required for quelling, and quelling is also the result of DNA damage, indicating that quelling and qiRNA production share a common mechanism. These results suggest that DNA damage triggered HR-based recombination allows the RNAi pathway to recognize repetitive DNA to produce small RNA. The involvement of chromatin remodeling factors indicates that siRNA biogenesis is regulated in the chromatin level. From our systematic knock out library screen, we identified a novel component, RTT109, which is required for both qiRNA and quelling-induced small RNA production. RTT109 is a fungal-specific histone acetyltransferase (HAT) for histone H3 on lysine 56 and its catalytic activity is required for its function in small RNA production. Furthermore, we show that RTT109 is required for homologous recombination and H3K56Ac is enriched around double strand break, which overlaps with RAD51 binding. Taken together, our results suggest that H3K56 acetylation is required for small RNA production through its role in homologous recombination.