Abstract
In the fission yeast Schizosaccharomyces pombe, the RNA-Induced Transcriptional Silencing (RITS) complex has been proposed to target the chromosome via siRNA-dependent base-pairing interactions to initiate heterochromatin formation. Here we show that tethering of the RITS subunit, Tas3, to the RNA transcript of the normally active ura4+ gene silences ura4+ expression. This silencing depends on a functional RNAi pathway, requires the heterochromatin proteins, Swi6/HP1, Clr4/Suv39h, and Sir2, and is accompanied by the generation of ura4+ siRNAs, histone H3-lysine 9 methylation, and Swi6 binding. Furthermore, the ability of the newly generated ura4+ siRNAs to silence a second ura4+ allele in trans is strongly inhibited by the conserved siRNA nuclease, Eri1. Surprisingly, silencing of tethered ura4+, or ura4+ inserted within centromeric heterochromatin, or some of the endogenous centromeric repeat promoters, is not associated with changes in RNA polymerase II occupancy. These findings support a model in which targeting of nascent transcripts by RITS mediates chromatin modifications and suggest that cotranscriptional processing events play a primary role in the silencing mechanism.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Carrier Proteins / genetics
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Carrier Proteins / metabolism
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Cell Cycle Proteins / genetics
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Cell Cycle Proteins / metabolism
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Chromosomal Proteins, Non-Histone / genetics
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Chromosomal Proteins, Non-Histone / metabolism
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DNA Methylation
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Exoribonucleases / genetics
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Exoribonucleases / metabolism
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Gene Expression Regulation, Fungal / genetics
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Heterochromatin / genetics*
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Histone-Lysine N-Methyltransferase
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Histones / metabolism
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Methyltransferases / genetics
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Methyltransferases / metabolism
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RNA Interference / physiology*
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RNA Precursors / genetics*
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RNA, Small Interfering / genetics*
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RNA-Binding Proteins / genetics
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RNA-Induced Silencing Complex / genetics*
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Regulatory Elements, Transcriptional / genetics
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Schizosaccharomyces / genetics*
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Schizosaccharomyces pombe Proteins / genetics
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Schizosaccharomyces pombe Proteins / metabolism
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Sirtuins / genetics
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Sirtuins / metabolism
Substances
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Carrier Proteins
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Cell Cycle Proteins
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Chromosomal Proteins, Non-Histone
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Heterochromatin
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Histones
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RNA Precursors
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RNA, Small Interfering
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RNA-Binding Proteins
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RNA-Induced Silencing Complex
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Schizosaccharomyces pombe Proteins
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Swi6 protein, S pombe
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Tas3 protein, S pombe
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ura4 protein, S pombe
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Methyltransferases
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Histone-Lysine N-Methyltransferase
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clr4 protein, S pombe
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Eri1 protein, S pombe
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Exoribonucleases
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Sirtuins