The Paf1 Complex Is Essential for Histone Monoubiquitination by the Rad6-Bre1 Complex, Which Signals for Histone Methylation by COMPASS and Dot1p

Abstract

Monoubiquitination of histone H2B, catalyzed byRad6-Bre1, is required for methylation of histone H3 on lysines 4 and 79, catalyzed by the Set1-containing complex COMPASS and Dot1p, respectively. The Paf1 protein complex, which associates with RNA polymerase II, is known to be required for these histone H3 methylation events. During the early elongation stage of transcription, the Paf1 complex is required for association of COMPASS with RNA polymerase II, but the role the Paf1 complex plays at the promoter has not been clear. We present evidence that the Paf1 complex is required for monoubiquitination of histone H2B at promoters. Strains deleted for several components of the Paf1 complex are defective in monoubiquitination of histone H2B, which results in the loss of methylation of lysines 4 and 79 of histone H3. We also show that Paf1 complex is required for the interaction of Rad6 and COMPASS with RNA polymerase II. Finally, we show that the Paf1 complex is required for Rad6-Bre1 catalytic activity but not for the recruitment of Rad6-Bre1 to promoters. Thus, in addition to its role during the elongation phase of transcription, the Paf1 complex appears to activate the function but not the placement of the Rad6-Bre1 ubiquitinprotein ligase at the promoters of active genes.The DNA of eukaryotic organisms assembles around histone proteins in nucleosomes to form highly organized structures known as chromatin (1). Alterations in chromatin structure play a major role in regulating gene expression, and for this reason much attention has been focused recently on the covalent modifications of histone proteins and their outcomes in transcriptional elongation (1-4). Essential to this process are the N-terminal tails of histone proteins. Because they protrude from the globular body of the nucleosome and are available for interactions with other proteins, the tails are the site of many covalent modifications that alter nucleosome structure. A myriad of modifications, such as acetylation, phosphorylation, ubiquitination, and methylation, decorate each histone tail (2, 5) The combinatorial effects of such modifications can produce an array of different responses involved in transcriptional activation and repression (1-6).One histone modification of major consequence is the methylation of histone H3 at lysines 4 and 79, catalyzed by the Set1-containing complex COMPASS 1 and Dot1p, respectively (7)(8)(9)(10)(11)(12)(13)(14). It has been shown that methylation of both lysine residues impacts the expression of genes within the rDNA loci and telomeric regions of DNA in Saccharomyces cerevisiae (7-8, 15, 16). It has been demonstrated that some of the components of the Paf1 complex, a complex that associates with the initiating and elongating RNA polymerase II, is also required for histone H3 methylation on lysines 4 and 79 (17, 18). Accordingly, previous studies have demonstrated a role for the Paf1 complex in transcriptional elongation and initiation (19 -22). A further requirement for the methylation of both lysines 4 and 79 of hi...

Molecular implementation and physiological roles for histone H3 lysine 4 (H3K4) methylation

“…The methyltransferase function of MLLs in human are non-redundant as deletion of either MLLs result in embryonic lethality. Previous studies demonstrated that COMPASS interacts with the elongating form of Pol II via its interaction with the Paf1 complex [32,43]. Therefore, (A) in the absence of H2B monoubiquitination, COMPASS can still interact with the transcribing Pol II via the Paf1 complex and monomethylate H3K4.…”

“…We now know that monoubiquitination of H2B is required for H3K4 di-and trimethylation in yeast and that this crosstalk pathway is highly conserved from yeast to human [6,40,41]. Although the past several years brought about a watershed of information regarding the factors/proteins required for proper H2B monoubiquitination [6,[40][41][42][43][44][45][46][47][48], the molecular mechanism for this histone crosstalk was poorly understood. Recent studies in yeast S. cerevisiae demonstrated that monoubiquitination of histone H2B regulates COMPASS' compositional assembly, and therefore, proper H3K4 methylation [49].…”

The human PAF complex coordinates transcription with events downstream of RNA synthesis

“…Therefore, yPAF was postulated to be involved in both RNA polymerase IImediated transcription initiation and elongation. Recently, the yPAF complex has been shown to be required for Rad6-mediated histone H2B-K123 monoubiquitination (Ng et al 2003b;Wood et al 2003), Set1-mediated histone H3-K4 methylation (Krogan et al 2003;Ng et al 2003a), and Dot1-mediated histone H3-K79 methylation (Krogan et al 2003), placing PAF at the center of cotranscriptional histone modifications (for review, see Hampsey and Reinberg 2003). PAF subunits are also required for maintaining proper poly(A) tail length in yeast, suggesting a post-transcriptional role for PAF complex in mRNA processing and maturation (Mueller et al 2004).…”

The eleven-nineteen-leukemia protein ENL connects nuclear MLL fusion partners with chromatin

“…After thorough washing, the retained proteins were liberated by heating in SDS sample buffer and analysed by immunoblotting with an anti-H3 antibody (Feng et al, 2002), was found in a mammalian twohybrid assay to bind to ENL (Guoliang Xi, Shanghai, personal communication). Histone H3 K79 methylation is an activating epigenetic mark that is imposed during transcriptional elongation (Krogan et al, 2003;Wood et al, 2003). Also ELL, the first identified MLL partner with a known function works as elongation factor (Shilatifard et al, 1996).…”