The C-terminal domain of pol II and a DRB-sensitive kinase are required for 3' processing of U2 snRNA

Abstract

The human snRNA genes transcribed by RNA polymerase II (e.g. U1 and U2) have a characteristic TATA-less promoter containing an essential proximal sequence element. Formation of the 3¢ end of these non-polyadenylated RNAs requires a specialized 3¢ box element whose function is promoter speci®c. Here we show that truncation of the C-terminal domain (CTD) of RNA polymerase II and treatment of cells with CTD kinase inhibitors, including DRB (5,6-dichloro-1-b-D-ribofuranosylbenzimidazole), causes a dramatic reduction in proper 3¢ end formation of U2 transcripts. Activation of 3¢ box recognition by the phosphorylated CTD would be consistent with the role of phospho-CTD in mRNA processing. CTD kinase inhibitors, however, have little effect on initiation or elongation of transcription of the U2 genes, whereas elongation of transcription of the b-actin gene is severely affected. This result highlights differences in transcription of snRNA and mRNA genes.

Transcription termination by nuclear RNA polymerases

“…CTD truncation and CTD kinase inhibitors affect 39-box recognition of U2 but not transcription termination (Medlin et al 2003;Jacobs et al 2004). Medlin et al (2005) showed that P-TEFb and phosphorylation of both Ser2 and Ser5 are required for cotranscriptional recognition of the 39-box.…”

“…While transcription of U1 genes terminates close to the 39-box (Cuello et al 1999), U2 transcription terminates ;1 kb from its mature 39-end (Medlin et al 2003). An unidentified termination factor or complex appears to bind the U1 precursor RNA immediately downstream from the 39-box at a G track, just upstream of a T stretch (Cuello et al 1999).…”

Formation of the 3′ end of histone mRNA: Getting closer to the end

“…Pre-snRNAs are processed by a 12-subunit complex called Integrator that contains a putative endonuclease RC-68/Int11 and directly associates with the CTD (Baillat et al, 2005). Inhibition of Cdk9 activity does not affect generation of pre-snRNAs although it abolishes their 3' end processing (Jacobs et al, 2004;Medlin et al, 2005;Medlin et al, 2003), demonstrating that the CTD phosphorylated on serine 2 is selectively required for 3' end processing of pre-snRNAs but is dispensable for transcription elongation. In agreement with these results, addition of the phosphorylated CTD stimulates in vitro 3' end processing of snRNA precursors .…”

Biogenesis of small nuclear RNPs

“…Stable pre-snRNA intermediates carrying only a short (5-10 nt) 3′ overhang are generated by a transcription-linked processing event that requires a cis-acting signal sequence (3′ box) and phosphorylation of the Cterminal domain of the largest subunit of Pol II (Medlin et al, 2003). The presnRNA is assembled into a large export complex that includes the 7mG-cap-binding complex (CBC), the phosphorylated adaptor for RNA export (PHAX) and the CRM1/RanGTP complex.…”