Chd1 and yFACT Act in Opposition in Regulating Transcription

Chromatin structure limits the accessibility of DNA sequences in eukaryotic chromosomes. Accessibility is enhanced through three major processes in vivo. First, posttranslational histone modifications either change the properties of the chromatin structure or create binding sites for other transcription factors (50). These posttranslational modifications include phosphorylation of serine residues and acetylation, methylation, and ubiquitylation of lysine residues (11). Several transcription factors that recognize specific histone modifications for their recruitment have been described. For example, the bromodomain-containing proteins recognize acetylation of histone proteins (61), and chromodomain-containing proteins are involved in recognizing methylation marks on histone proteins (6, 11). Second, ATP-dependent chromatin-remodeling factors promote accessibility by repositioning nucleosomes (9, 21, 55). These factors utilize the energy from ATP hydrolysis to establish or disrupt repressive chromatin structures (21). The third way by which the DNA sequence is made available is through ATP-independent chromatin-reorganizing factors that change the properties of nucleosomes in a localized manner (17). For example, the yFACT complex changes the properties of nucleosomes without requiring ATP hydrolysis (39, 41). The reorganization by yFACT has been shown to alter DNA accessibility both in vivo and in vitro (5, 14, 17, 30, 44, 45). The FACT complex (facilitates chromatin transcription) was first identified as a factor that promoted RNA polymerase II (Pol II) transcription in vitro using assembled chromatin as a template (33). The mammalian FACT complex is composed of two subunits, p140 and SSRP1. The homologs of p140 and SSRP1 in yeast are Spt16 and Pob3, respectively (34). The Spt16 and Pob3 proteins are always present in a heterodimer to form the SP complex in yeast (57). Although the N-terminal DNA binding domain of SSRP1 is absent in Pob3, Nhp6, a high-mobility group protein, is thought to serve as the DNA binding activity of the SP complex to form the yFACT complex (7, 18). Genetic and biochemical evidence suggests that yFACT is involved in regulating both transcription and DNA replication (1a, 3, 5, 18, 19, 27, 30, 34, 42, 56, 57). While the association of yFACT with elongation factors (27, 45) and with transcribed regions of genes (30, 42) supports an elongation role, studies also suggest that the FACT complex has a role in transcription initiation (5, 44). We have shown earlier that yFACT has a role in regulating TATA binding protein (TBP) binding during the transcriptional initiation step (5). The evidence for this included synthetic lethality between certain mutations of TBP and TFIIA and defective alleles of SPT16, reduced binding of TBP at some promoters in spt16 mutants, and enhanced binding of TBP to a TATA box within nucleosomal DNA in presence of TFIIA and yFACT. The yeast chromodomain protein (Chd1) is a member of the Snf2-like subfamily of nucleic acid-stimulated ATPases (21) and has ATP-dependent chromatin-remodeling activity in vitro (29, 40). Chd1 and other CHD proteins have two chromodomains near the N terminus, a centrally located Snf2-related helicase/ATPase domain, and a Myb-related DNA binding domain near the C terminus (59). Chd1 is thought to promote formation of inhibitory chromatin, as extracts derived from cells lacking Chd1 are unable to produce the same level of DNase I resistance at specific loci that results from similar preparations derived from normal cells (40). Genetic interactions have been reported between mutations of CHD1 and mutations in transcription elongation factors such as Spt5, Isw1, and Isw2 (45, 54). Chd1 also physically interacts with several transcription elongation factors, such as members of the Paf1 complex, the Spt4-Spt5 complex, and components of yFACT (24, 27, 45, 54). Recently Chd1 has been shown to physically associate with the SAGA/SLIK complex in yeast and to bind histone tail peptides methylated at K4 (37). However, binding of yeast Chd1 to methylated H3-K4 has not been observed by others (32, 46). Relatively little is known about the functional role of Chd1 in vivo in regulating transcription, although it was recently reported that a chd1 mutation affects chromatin structure of the ADH2 gene and the kinetics of ADH2 activation (60). In this report, we show that part of the role of Chd1 is to oppose the function of the positive transcription factor yFACT. We present evidence which suggests that Chd1 negates yFACT's ability to enhance TBP binding at promoters. We show that in a strain with a yFACT defect, deletion of CHD1 results in increased TBP binding and increased Pol II binding at promoters. Finally, we find that deletion of CHD1 suppresses synthetic lethalities between spt16 mutations and TBP mutations as well as between spt16-11 and TFIIA mutations.