title: Decoupling the downstream effects of germline nuclear RNAi reveals that H3K9me3 is dispensable for heritable RNAi and the maintenance of endogenous siRNA-mediated transcriptional silencing inCaenorhabditis elegans.
abstract: Germline nuclear RNAi inC. elegansis a transgenerational gene-silencing pathway that leads to H3K9 trimethylation (H3K9me3) and transcriptional silencing at the target genes. H3K9me3 induced by either exogenous double-stranded RNA (dsRNA) or endogenous siRNA (endo-siRNA) is highly specific to the target loci and transgenerationally heritable. Despite these features, the role of H3K9me3 in siRNA-mediated transcriptional silencing and inheritance of the silencing state at native target genes is unclear. In this study, we took combined genetic and whole-genome approaches to address this question.
  
  
  
  Here we demonstrate that siRNA-mediated H3K9me3 requires combined activities of three H3K9 histone methyltransferases: MET-2, SET-25, and SET-32.set-32single,met-2 set-25double, andmet-2 set-25;set-32triple mutant adult animals all exhibit prominent reductions in H3K9me3 throughout the genome, withmet-2 set-25;set-32mutant worms losing all detectable H3K9me3 signals. Surprisingly, loss of high-magnitude H3K9me3 at the native nuclear RNAi targets has no effect on the transcriptional silencing state. In addition, the exogenous dsRNA-induced transcriptional silencing and heritable RNAi atoma-1, a well-established nuclear RNAi reporter gene, are completely resistant to the loss of H3K9me3.
  
  
  
  Nuclear RNAi-mediated H3K9me3 inC. elegansrequires multiple histone methyltransferases, including MET-2, SET-25, and SET-32. H3K9me3 is not essential for dsRNA-induced heritable RNAi or the maintenance of endo-siRNA-mediated transcriptional silencing inC. elegans. We propose that siRNA-mediated transcriptional silencing inC. eleganscan be maintained by an H3K9me3-independent mechanism.
authors.affiliation: Department of Molecular Biology and Biochemistry, Rutgers the State University of New Jersey, Piscataway, NJ 08854 USA.
authors.forename: Natallia
authors.initials: N
authors.lastname: Kalinava
authors.affiliation: Department of Molecular Biology and Biochemistry, Rutgers the State University of New Jersey, Piscataway, NJ 08854 USA.
authors.forename: Julie Zhouli
authors.initials: JZ
authors.lastname: Ni
authors.affiliation: Department of Molecular Biology and Biochemistry, Rutgers the State University of New Jersey, Piscataway, NJ 08854 USA.
authors.forename: Kimberly
authors.initials: K
authors.lastname: Peterman
authors.affiliation: Department of Molecular Biology and Biochemistry, Rutgers the State University of New Jersey, Piscataway, NJ 08854 USA.
authors.forename: Esteban
authors.initials: E
authors.lastname: Chen
authors.affiliation: Department of Molecular Biology and Biochemistry, Rutgers the State University of New Jersey, Piscataway, NJ 08854 USA.|Nelson Labs A125, 604 Allison Road, Piscataway, NJ 08854 USA.
authors.forename: Sam Guoping
authors.initials: SG
authors.lastname: Gu
mesh_terms: D000818:Animals; D017173:Caenorhabditis elegans / Q000235:genetics* / Q000378:metabolism; D029742:Caenorhabditis elegans Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D002352:Carrier Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D047369:Chromatin Immunoprecipitation; D016678:Genome; D042822:Genomic Instability; D005854:Germ Cells / Q000378:metabolism; D059014:High-Throughput Nucleotide Sequencing; D000076983:Histone Methyltransferases; D011495:Histone-Lysine N-Methyltransferase / Q000235:genetics / Q000378:metabolism; D006657:Histones / Q000235:genetics / Q000378:metabolism*; D008745:Methylation; D008856:Microscopy, Fluorescence; D016296:Mutagenesis; D034622:RNA Interference; D012330:RNA, Double-Stranded / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction; D017423:Sequence Analysis, RNA; D014158:Transcription, Genetic
chemical_list: D029742:Caenorhabditis elegans Proteins; D002352:Carrier Proteins; D006657:Histones; C440205:OMA-1 protein, C elegans; D012330:RNA, Double-Stranded; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D000076983:Histone Methyltransferases; D011495:Histone-Lysine N-Methyltransferase