RNAi-dependent and RNAi-independent mechanisms contribute to the silencing of RIPed sequences in Neurospora crassa.
RNA interference (RNAi) can silence genes at the transcriptional level by targeting locus-specific Lys9H3 methylation or at the post-transcriptional level by targeting mRNA degradation. Here we have cloned and sequenced genomic regions methylated in Lys9H3 in Neurospora crassa to test the requirements for components of the RNAi pathway in this modification. We find that 90% of clones map to repeated sequences and relics of transposons that have undergone repeat-induced point mutations (RIP). We find siRNAs derived from transposon relics indicating that the RNAi machinery targets these regions. This is confirmed by the fact that the presence of these siRNAs depends on components of the RNAi pathway such as the RdRP (QDE-1), the putative RecQ helicase (QDE-3) and the two Dicer enzymes. We show that Lys9H3 methylation of RIP sequences is not affected in mutants of the RNAi pathway indicating that the RNAi machinery is not involved in transcriptional gene silencing in Neurospora. We find that RIP regions are transcribed and that the transcript level increases in the mutants of the RNAi pathway. These data suggest that the biological function of the Neurospora RNAi machinery is to control transposon relics and repeated sequences by targeting degradation of transcripts derived from these regions.
Chicas
Agustin
A
Istituto Pasteur e Fondazione Cenci Bolognetti, Dipartimento di Biotecnologie Cellulari ed Ematologia, Sezione di Genetica Molecolare, Università di Roma La Sapienza, Viale Regina Elena, 324, 00161 Roma, Italy.
Cogoni
Carlo
C
Macino
Giuseppe
G
D004251:DNA Transposable Elements / Q000235:genetics*; D004271:DNA, Fungal / Q000737:chemistry; D015966:Gene Expression Regulation, Fungal; D020868:Gene Silencing; D016681:Genome, Fungal; D006657:Histones / Q000737:chemistry / Q000378:metabolism; D008239:Lysine / Q000378:metabolism; D008745:Methylation; D009492:Neurospora crassa / Q000235:genetics* / Q000378:metabolism; D017354:Point Mutation; D034622:RNA Interference; D012331:RNA, Fungal / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D012091:Repetitive Sequences, Nucleic Acid
D004251:DNA Transposable Elements; D004271:DNA, Fungal; D006657:Histones; D012331:RNA, Fungal; D034741:RNA, Small Interfering; D008239:Lysine