Global Role for Polyadenylation-Assisted Nuclear RNA Degradation in Posttranscriptional Gene Silencing

Abstract

Fission yeast Cid14, a component of the TRAMP (Cid14/Trf4-Air1-Mtr4 polyadenylation) complex, polyadenylates nuclear RNA and stimulates degradation by the exosome for RNA quality control. Here, we analyze patterns of global gene expression in cells lacking the Cid14 or the Dis3/Rpr44 subunit of the nuclear exosome. We found that transcripts from many genes induced during meiosis, including key regulators, accumulated in the absence of Cid14 or Dis3. Moreover, our data suggest that additional substrates include transcripts involved in heterochromatin assembly. Mutant cells lacking Cid14 and/or Dis3 accumulate transcripts corresponding to naturally silenced repeat elements within heterochromatic domains, reflecting defects in centromeric gene silencing and derepression of subtelomeric gene expression. We also uncover roles for Cid14 and Dis3 in maintaining the genomic integrity of ribosomal DNA. Our data indicate that polyadenylation-assisted nuclear RNA turnover functions in eliminating a variety of RNA targets to control diverse processes, such as heterochromatic gene silencing, meiotic differentiation, and maintenance of genomic integrity.Polyadenylation is important for the maturation of mRNAs (26), while our recent work has also uncovered unexpected links between polyadenylation and chromosome replication and segregation (33,35,37). In addition to its function in the maturation of mRNAs, nuclear polyadenylation is important for diverse cellular activities, such as RNA interference (RNAi)-mediated heterochromatin assembly and quality control of noncoding RNAs (33,35,37). These functions involve Cid12 and Cid14, members of a widespread family of noncanonical poly(A) polymerases found in eukaryotes from yeasts to humans (30). Besides its function in checkpoint control, Cid12 is required for faithful chromosome segregation and RNAi-mediated heterochromatin assembly at centromeres (19,37). RNAi silencing is triggered by double-stranded RNA, which is processed by the RNase III-like RNase Dicer into small interfering RNA (siRNA) molecules of around 21 nucleotides. These siRNAs become incorporated into the RNAinduced transcriptional silencing (RITS) complex, directing the complex to homologous RNA targets (6). In worms, plants, and fungi, RNAi also requires RNA-directed RNA polymerases (RDRs), which are involved in the siRNA-and template-directed production of double-stranded RNA (1, 17, 29). Motamedi et al. (19) identified Cid12 in an RDR complex (RDRC) which also contains Rdp1 (the fission yeast RDR homolog) and Hrr1 (an RNA helicase). RDRC physically interacts with the RITS complex in a manner that requires Dicer and the histone methyltransferase Clr4. In cells lacking Cid12, RITS complexes are devoid of siRNA and fail to localize to centromeric DNA repeats to initiate heterochromatin assembly.In Saccharomyces cerevisiae, the Cid1-like proteins Trf4 and Trf5 share an essential function that involves the polyadenylation of nuclear RNAs as part of a pathway of exosomemediated RNA turnover (10,12,31,38). We have ide...

Negative Regulation of Meiotic Gene Expression by the Nuclear Poly(a)-binding Protein in Fission Yeast

“…This therefore suggests rapid co-transcriptional degradation of meiotic transcripts, similar to what has been seen for cryptic unstable transcripts that are also untraceable in normal growing cells (26,50). Also similar to the pathway of cryptic unstable transcript decay, recent studies suggest that the exosome complex contributes to the degradation of fission yeast meiotic transcripts (51,52). Consistently, we found several meiotic transcripts to be up-regulated in cells in which genes that encode exosome components were deleted or mutated (Fig.…”

Common themes in siRNA-mediated epigenetic silencing pathways

“…In parallel to RNAi, another RNA degradation process, requiring the protein Cid14, which belongs to an atypical class of polyA polymerases (like Cid12 a subunit of RDRC), and Dis3 the main exosome ribonuclease, has also been found implicated in transcriptional silencing of all major heterochromatic sites in fission yeast Murakami et al, 2007;Wang et al, 2008). In a cid14 deficient strain, heterochromatic silencing at centromeres is relieved and the level of centromeric siRNAs dramatically reduces.…”

Activities of human RRP6 and structure of the human RRP6 catalytic domain

“…In addition to its role in RNA degradation, the exosome is required for efficient processing and maturation of ribosomal RNA, stable small nucleolar RNA, and small nuclear RNA. Evidence also supports a role for the exosome in controlling the levels of histone mRNAs (Chen et al 2001;Canavan and Bond 2007;Reis and Campbell 2007), cryptic unstable transcripts (CUTs), and antisense transcripts through an association with the TRAMP complex in processes that contribute to post-transcriptional gene silencing and heterochromatin formation (LaCava et al 2005;Vanacova et al 2005;Wyers et al 2005;Davis and Ares 2006;Camblong et al 2007;Houseley and Tollervey 2008;Wang et al 2008a).…”