A Posttranscriptional Role for the Yeast Paf1-RNA Polymerase II Complex Is Revealed by Identification of Primary Targets

Department of Biochemistry and Molecular Genetics and Molecular Biology Program, University of Colorado, Denver, USA.
Molecular Cell (Impact Factor: 14.02). 11/2005; 20(2):213-23. DOI: 10.1016/j.molcel.2005.08.023
Source: PubMed


The yeast Paf1 complex (Paf1C: Paf1, Cdc73, Ctr9, Rtf1, and Leo1) is associated with RNA Polymerase II (Pol II) at promoters and coding regions of transcriptionally active genes, but transcript abundance for only a small subset of genes is altered by loss of Paf1. By using conditional and null alleles of PAF1 and microarrays, we determined the identity of both primary and secondary targets of the Paf1C. Neither primary nor secondary Paf1C target promoters were responsive to loss of Paf1. Instead, Paf1 loss altered poly(A) site utilization of primary target genes SDA1 and MAK21, resulting in increased abundance of 3'-extended mRNAs. The 3'-extended MAK21 RNA is sensitive to nonsense-mediated decay (NMD), as revealed by its increased abundance in the absence of Upf1. Therefore, although the Paf1C is associated with Pol II at initiation and during elongation, these critical Paf1-dependent changes in transcript abundance are due to alterations in posttranscriptional processing.

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    • "Additionally, PAF1C targets Rad6-Bre1 to promoters, allowing H2BK123 monoubiquitylation, which in turn helps the methylation of H3K4 by Set1/COMPASS, a hallmark of active transcription (Wood et al. 2003). Finally, PAF1C is required for proper 3 ′ end processing and termination of mRNA and snoRNA (Penheiter et al. 2005). Although transcription–replication interference has been identified as a major source of genome instability in recent years, few studies have examined the role of the Mec1/ATR checkpoint kinase in such conflicts. "
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    ABSTRACT: Little is known about how cells ensure DNA replication in the face of RNA polymerase II (RNAPII)-mediated transcription, especially under conditions of replicative stress. Here we present genetic and proteomic analyses from budding yeast that uncover links between the DNA replication checkpoint sensor Mec1-Ddc2 (ATR-ATRIP), the chromatin remodeling complex INO80C (INO80 complex), and the transcription complex PAF1C (PAF1 complex). We found that a subset of chromatin-bound RNAPII is degraded in a manner dependent on Mec1, INO80, and PAF1 complexes in cells exposed to hydroxyurea (HU). On HU, Mec1 triggers the efficient removal of PAF1C and RNAPII from transcribed genes near early firing origins. Failure to evict RNAPII correlates inversely with recovery from replication stress: paf1Δ cells, like ino80 and mec1 mutants, fail to restart forks efficiently after stalling. Our data reveal unexpected synergies between INO80C, Mec1, and PAF1C in the maintenance of genome integrity and suggest a mechanism of RNAPII degradation that reduces transcription-replication fork collision.
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    • "It was suggested that Cdc73 might regulate mRNA processing by facilitating the recruitment of 3′ factors to transcribed loci. Other evidence of PAF involvement in 3′ processing comes from yeast, where PAF was shown to affect poly (A) tail length (Mueller et al., 2004) and poly (A) site selection (Penheiter et al., 2005). In addition, another study (Nagaike et al., 2011) found a role of PAF in mediating stimulation of 3′ processing by transcriptional activators, confirming its potential role in bridging polyadenylation to transcription (Fig. 3B). "
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    • "ticular , Paf1 interacts both genetically and physically with Spt16 ( Squazzo et al . , 2002 ) . Consistent with this , Spt16 recruitment is reduced in the absence of Paf1 ( Pruneski et al . , 2011 ) . Additionally , Paf1 is involved in regulating transcription termination and mRNA 3 ' processing ( Mueller et al . , 2004 ; Nagaike et al . , 2011 ; Penheiter et al . , 2005 ; Sheldon et al . , 2005 ) ."
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    ABSTRACT: Cmr1 (Changed mutation rate) is a histone interacting, WD40 domain containing protein, with a previously identified role in the DNA damage repair pathway. Recent interest in Cmr1 has suggested roles in additional cellular processes. The current study was carried out to determine whether Cmr1 is involved in Pol II mediated transcription. Results presented herein provide the first evidence that Cmr1 is recruited to the coding regions of Pol II transcribed genes. It is recruited to the 5’ open reading frame in a manner dependent on Ser5 phosphorylation of the Pol II carboxyl terminal domain by the kinase Kin28. Strong evidence is provided that a primary function of Cmr1 is to coordinate recruitment and retention of the multifunctional elongation factor Paf1. Cmr1 was observed to occupy the coding region of the inducible genes ARG1 and HIS4 at significant levels, but not that of the constitutively expressed genes ADH1 and PMA1, suggesting that it regulates a subset of the multiple functions identified for Paf1. Additionally, it is important for efficient recruitment of the histone chaperone complex FACT and of the mRNA processing factor Rna14, both of which are consistent with previously identified roles for Paf1.
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