The activities of several mRNA processing factors are coupled to transcription through binding to RNA polymerase II (Pol II). The largest subunit of Pol II contains a repetitive carboxy-terminal domain (CTD) that becomes highly phosphorylated during transcription. mRNA-capping enzyme binds only to phosphorylated CTD, whereas other processing factors may bind to both phosphorylated and unphosphorylated forms. Capping occurs soon after transcription initiation and before other processing events, raising the question of whether capping components remain associated with the transcription complex after they have modified the 5 end of the mRNA. Chromatin immunoprecipitation in Saccharomyces cerevisiae shows that capping enzyme cross-links to promoters but not coding regions. In contrast, the mRNA cap methyltransferase and the Hrp1/CFIB polyadenylation factor cross-link to both promoter and coding regions. Remarkably, the phosphorylation pattern of the CTD changes during transcription. Ser 5 phosphorylation is detected primarily at promoter regions dependent on TFIIH. In contrast, Ser 2 phosphorylation is seen only in coding regions. These results suggest a dynamic association of mRNA processing factors with differently modified forms of the polymerase throughout the transcription cycle. Eukaryotic mRNAs undergo 5Ј end capping, splicing of introns, and polyadenylation. Targeting of capping enzyme and other RNA processing factors is through binding to the carboxy-terminal domain (CTD) of the RNA polymerase II (Pol II) largest subunit McCracken et al. 1997a,b;Yue et al. 1997;Hirose and Manley 1998;Pillutla et al. 1998;Hirose et al. 1999). Capping is the earliest modification, occurring when the transcript is 20 to 40 nucleotides long (Jove and Manley 1984;Rasmussen and Lis 1993). Phosphorylation of the CTD occurs soon after initiation and is necessary for capping enzyme recruitment. Other RNA-processing factors bind to both phosphorylated and unphosphorylated CTD and act much later during transcription. This raises the question of whether capping enzyme and other processing factors are simultaneously associated with RNA pol II throughout transcription or instead interact transiently at different stages. In vivo cross-linking is used here to show that capping enzyme is recruited to promoter regions dependent on TFIIH kinase activity, but does not remain associated with elongating polymerase. In contrast, the mRNA cap methyltransferase Abd1 and the polyadenylation factor CFIB/Hrp1 cross-link throughout transcribed regions. Surprisingly, Ser 5 phosphorylation of the CTD also localizes to promoters, suggesting dephosphorylation not long after escape into elongation phase. Ser 2 phosphorylation of the CTD shows a complementary pattern, with no cross-linking at the promoter and higher levels near the 3Ј end of the gene. These results suggest a dynamic association of RNA processing factors with differently modified forms of the polymerase throughout the transcription cycle. Results Experimental designTo determine the in vivo ...

The yeast histone deacetylase Rpd3 can be recruited to promoters to repress transcription initiation. Biochemical, genetic, and gene-expression analyses show that Rpd3 exists in two distinct complexes. The smaller complex, Rpd3C(S), shares Sin3 and Ume1 with Rpd3C(L) but contains the unique subunits Rco1 and Eaf3. Rpd3C(S) mutants exhibit phenotypes remarkably similar to those of Set2, a histone methyltransferase associated with elongating RNA polymerase II. Chromatin immunoprecipitation and biochemical experiments indicate that the chromodomain of Eaf3 recruits Rpd3C(S) to nucleosomes methylated by Set2 on histone H3 lysine 36, leading to deacetylation of transcribed regions. This pathway apparently acts to negatively regulate transcription because deleting the genes for Set2 or Rpd3C(S) bypasses the requirement for the positive elongation factor Bur1/Bur2.

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Summary The C-terminal domain of the RNA polymerase II largest subunit undergoes dynamic phosphorylation during transcription, and the different phosphorylation patterns that predominate at each stage of transcription recruit the appropriate set of mRNA processing and histone modifying factors. Recent papers help explain how the changes in CTD phosphorylation pattern are linked to the progression from initiation through elongation to termination.