The Bur1-Bur2 and Paf1 complexes function during transcription elongation and affect histone modifications. Here we describe new roles for Bur1-Bur2 and the Paf1 complex. We find that histone H3 K36 tri-methylation requires specific components of the Paf1 complex and that K36 trimethylation is more strongly affected at the 5 0 ends of genes in paf1D and bur2D strains in parallel with increased acetylation of histones H3 and H4. Interestingly, the 5 0 increase in histone acetylation is independent of K36 methylation, and therefore is mechanistically distinct from the methylation-driven deacetylation that occurs at the 3 0 ends of genes. Finally, Bur1-Bur2 and the Paf1 complex have a second methylation-independent function, since bur2D set2D and paf1D set2D double mutants display enhanced histone acetylation at the 3 0 ends of genes and increased cryptic transcription initiation. These findings identify new functions for the Paf1 and Bur1-Bur2 complexes, provide evidence that histone modifications at the 5 0 and 3 0 ends of coding regions are regulated by distinct mechanisms, and reveal that the Bur1-Bur2 and Paf1 complexes repress cryptic transcription through a Set2-independent pathway.

In Saccharomyces cerevisiae, Maf1 is essential for mediating the repression of transcription by RNA polymerase (pol) III in response to diverse cellular conditions. These conditions activate distinct signaling pathways that converge at or above Maf1. Thus, Maf1-dependent repression is thought to involve a common set of downstream inhibitory effects on the pol III machinery. Here we provide support for this view and define two steps in Maf1-dependent transcriptional repression. We show that chlorpromazine (CPZ)-induced repression of pol III transcription is achieved by inhibiting de novo assembly of transcription factor (TF) IIIB onto DNA as well as the recruitment of pol III to preassembled TFIIIB⅐DNA complexes. Additionally Brf1 was identified as a target of repression in extracts of CPZ-treated cells. Maf1-Brf1 and Maf1-pol III interactions were implicated in the inhibition of TFIIIB⅐DNA complex assembly and polymerase recruitment by recombinant Maf1. Co-immunoprecipitation experiments confirmed these interactions in yeast extracts and demonstrated that Maf1 does not differentially sequester Brf1 or pol III under repressing conditions. The results suggest that Maf1 functions by a non-stoichiometric mechanism to repress pol III transcription.Transcription of the large rRNAs by RNA polymerase (pol) 1 I and of 5 S rRNA and tRNAs by pol III is tightly co-regulated under essentially all conditions (1-3). This coordinate regulation is biologically important as it is conserved in all eukaryotes where transcription by pols I and III has been examined. The principal evolutionary imperatives that are thought to underlie this conserved regulation are the common function of rRNAs and tRNA in protein synthesis and the high energetic cost of their synthesis, which accounts for about 80% of nuclear gene transcription in actively growing cells (1, 3). The levels of pol I and pol III transcription are critical determinants of cell growth rate, and the deregulation of this transcription is a hallmark of cell transformation and tumorigenesis (4, 5). In addition, for single cell eukaryotes whose biological niche exposes them to periods when nutrients are in short supply and/or harsh environmental conditions, the ability to rapidly shut off the synthesis of rRNAs and tRNA is thought to be of vital importance for achieving metabolic economy (6) and hence is likely to impact cell survival.In higher eukaryotes, p53 and Rb and its relatives p107 and p130 play an important role in controlling pol I and pol III transcription and in coordinating the production of new protein synthetic capacity with cell proliferation (2, 5). These repressors function by binding directly to components of the pol I and pol III transcription machinery and thereby prevent proteinprotein interactions required for transcription. Specific components of this machinery are also substrates for phosphorylation by various kinases including extracellular signal-regulated kinase, casein kinase II, and cyclin-dependent kinases, which can either activate or repress tran...

BUR1 and BUR2 encode the catalytic and regulatory subunits of a cyclin-dependent protein kinase complex that is essential for normal growth and has a general role in transcription elongation. To gain insight into its specific role in vivo, we identified mutations that reverse the severe growth defect of bur1⌬ cells. This selection identified mutations in SET2, which encodes a histone methylase that targets lysine 36 of histone H3 and, like BUR1, has a poorly characterized role during transcription elongation. This genetic relationship indicates that SET2 activity is required for the growth defect observed in bur1⌬ strains. This SET2-dependent growth inhibition occurs via methylation of histone H3 on lysine 36, since a methylation-defective allele of SET2 or a histone H3 K36R mutation also suppressed bur1⌬. We have explored the relationship between BUR1 and SET2 at the biochemical level and find that histone H3 is monomethylated, dimethylated, and trimethylated on lysine 36 in wild-type cells, but trimethylation is significantly reduced in bur1 and bur2 mutant strains. A similar methylation pattern is observed in RNA polymerase II C-terminal domain truncation mutants and in an spt16 mutant strain. Chromatin immunoprecipitation assays reveal that the transcription-dependent increase in trimethylated K36 over open reading frames is significantly reduced in bur2⌬ strains. These results establish links between a regulatory protein kinase and histone methylation and lead to a model in which the Bur1-Bur2 complex counteracts an inhibitory effect of Set2-dependent histone methylation.