Characterization of CAF4 and CAF16 Reveals a Functional Connection between the CCR4-NOT Complex and a Subset of SRB Proteins of the RNA Polymerase II Holoenzyme

Abstract

The CCR4-NOT transcriptional regulatory complex affects transcription both positively and negatively and consists of the following two complexes: a core 1 ؋ 10 6 dalton (1 MDa) complex consisting of CCR4, CAF1, and the five NOT proteins and a larger, less defined 1.9-MDa complex. We report here the identification of two new factors that associate with the CCR4-NOT proteins as follows: CAF4, a WD40-containing protein, and CAF16, a putative ABC ATPase. Whereas neither CAF4 nor CAF16 was part of the core CCR4-NOT complex, both CAF16 and CAF4 appeared to be present in the 1.9-MDa complex. CAF4 also displayed physical interactions with multiple CCR4-NOT components and with DBF2, a likely component of the 1.9-MDa complex. In addition, both CAF4 and CAF16 were found to interact in a CCR4-dependent manner with SRB9, a component of the SRB complex that is part of the yeast RNA polymerase II holoenzyme. The three related SRB proteins, SRB9, SRB10, and SRB11, were found to interact with and to coimmunoprecipitate DBF2, CAF4, CCR4, NOT2, and NOT1. Defects in SRB9 and SRB10 also affected processes at the ADH2 locus known to be controlled by components of the CCR4-NOT complex; an srb9 mutation was shown to reduce ADH2 derepression and either an srb9 or srb10 allele suppressed spt10-enhanced expression of ADH2. In addition, srb9 and srb10 alleles increased ADR1 c -dependent ADH2 expression; not4 and not5 deletions are the only other known defects that elicit this phenotype. These results suggest a close physical and functional association between components of the CCR4-NOT complexes and the SRB9, -10, and -11 components of the holoenzyme.

A Complex of the Srb8, -9, -10, and -11 Transcriptional Regulatory Proteins from Yeast

“…Moreover, Srb8, -9, -10, and -11 can be found in association with the CCR4-NOT complex, which is involved in general negative regulation of transcription (34). Similarly, only the smaller human Mediator complex, lacking hSrb8, -9, -10, and -11, supports positive transcription control in vitro, whereas the larger human complex that includes these four proteins is inactive (17,35).…”

A genome-wide RNAi screen identifies genes regulating the formation of P bodies in C. elegans and their functions in NMD and RNAi

“…Previous studies have demonstrated an extensive link between mRNA degradation and other biological processes. For example, the CCR4/NOT deadenylase complex represses the activity of RNA Polymerase II (Liu et al, 2001). The RNA polymerase subunit Rbp4 is required for deadenylation and decay of a subset of transcripts that encodes proteins involved in protein synthesis (Lotan et al, 2005).…”

A Genomic Screen in Yeast Reveals Novel Aspects of Nonstop mRNA Metabolism

“…These genes have been implicated in both transcriptional activation and repression of a variety of genes, suggesting that they might increase his3-nonstop expression by increasing the strength of the HIS3 promoter (Kelleher et al 1990;Flanagan et al 1991;Thompson et al 1993). On the other hand, Ssn2p and Ssn3p interact with the CCR4/NOT complex, which is the major cytoplasmic deadenylation complex, suggesting that they might have a direct role in mRNA decay (Liu et al 2001;Tucker et al 2001;Huh et al 2003).…”