Gene-specific RNA polymerase II phosphorylation and the CTD code

Abstract

Dynamic phosphorylation of the RNA polymerase II CTD repeats (YS2PTS5PS7) is coupled to transcription and may act as a “code” that controls mRNA synthesis and processing. To examine the "code" in budding yeast, we mapped genome-wide CTD S2, 5 and 7 phosphorylations (PO4) and compared them with the CTD-associated termination factors, Nrd1 and Pcf11. CTD-PO4 dynamics are not scaled to the size of the gene. At 5’ ends, the onset of S2-PO4 is delayed by about 450 bases relative to S5-PO4, regardless of gene length. Phospho-CTD dynamics are gene-specific, with high S5/7-PO4 at the 5' end being characteristic of well-expressed genes with nucleosome-occupied promoters. Furthermore, the CTD kinases Kin28 and Ctk1 profoundly affect pol II distribution along genes in a highly gene-specific way. The "code" is therefore written differently on different genes, probably under the control of promoters. S7-PO4 is enriched on introns and at sites of Nrd1 accumulation suggesting that this modification may function in splicing and Nrd1 recruitment. Nrd1 and Pcf11 frequently co-localized, suggesting functional overlap between these terminators. Surprisingly, Pcf11 is also recruited to centromeres and pol III transcribed genes.

Serine-7 but not serine-5 phosphorylation primes RNA polymerase II CTD for P-TEFb recognition

“…Particularly, Ser7 phosphorylation is found early in transcription initiation and retained until transcription termination in all RNAPII-dependent genes. In line with these observations, the dynamics of CTD phosphorylations were found to be not scaled to the gene length but differ among genes with different promoter structures and expression levels 19 . High levels of Ser7 and Ser5 phosphorylations occur at the transcription start site, whereas Ser2 phosphorylation levels increase until they reach their peak about 600-1,000 nucleotides downstream of the start site 20 .…”

Mechanism of alternative splicing and its regulation

“…For example, ser5 phosphorylation is associated with transcription initiation through cyclin-dependent kinase 7 (CDK7) of the general transcription factor IIH (TFIIH), whereas ser2 phosphorylation is preferentially linked with CTD activity at the 3'-end of genes through CDK9 of the positive transcription elongation factor b (42). In addition, phosphorylation of ser7 has been found to facilitate elongation and splicing (43). Thus, phosphorylation is a mechanism that clearly demonstrates that functional coupling exists between transcription and alternative splicing.…”

RNA Polymerase II transcription elongation and Pol II CTD Ser2 phosphorylation

“…In particular, Ser7 phosphorylation can be found throughout gene bodies in yeast. 12,[21][22][23] However, despite its presence on protein-coding genes, Ser7-P is particularly important for proper transcription of snRNA genes 16 and may or may not be required for transcription or processing of mRNA genes. 16,17 Thus, because Ser2 phosphorylation has the tightest association with transcription elongation regulation, we focus our discussion on the kinases that carry out this modification.…”