Positive transcription elongation factor B phosphorylates hSPT5 and RNA polymerase II carboxyl-terminal domain independently of cyclin-dependent kinase-activating kinase.
The CDK9-cyclin T kinase complex, positive transcription elongation factor b (P-TEFb), stimulates the process of elongation of RNA polymerase (Pol) II during transcription of human immunodeficiency virus. P-TEFb associates with the human immunodeficiency virus Tat protein and with the transactivation response element to form a specific complex, thereby mediating efficient elongation. Here, we show that P-TEFb preferentially phosphorylates hSPT5 as compared with the carboxyl-terminal domain of RNA Pol II in vitro. Phosphorylation of hSPT5 by P-TEFb occurred on threonine and serine residues in its carboxyl-terminal repeat domains. In addition, we provide several lines of evidence that P-TEFb is a CDK-activating kinase (CAK)-independent kinase. For example, CDK9 was not phosphorylated by CAK, whereas CDK2-cyclin A kinase activity was dramatically enhanced by CAK. Therefore, it is likely that P-TEFb participates in regulation of elongation by RNA Pol II by phosphorylation of its substrates, hSPT5 and the CTD of RNA Pol II, in a CAK-independent manner.
Kim
J B
JB
Center for Cancer Research, Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA.
Sharp
P A
PA
D001483:Base Sequence; D002868:Chromosomal Proteins, Non-Histone; D017931:DNA Primers; D005656:Fungal Proteins / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism*; D010766:Phosphorylation; D042925:Positive Transcriptional Elongation Factor B; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D012319:RNA Polymerase II / Q000737:chemistry / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*; D035602:Transcriptional Elongation Factors
D002868:Chromosomal Proteins, Non-Histone; D017931:DNA Primers; D005656:Fungal Proteins; D009687:Nuclear Proteins; D014157:Transcription Factors; D035602:Transcriptional Elongation Factors; C069940:SPT5 transcriptional elongation factor; D042925:Positive Transcriptional Elongation Factor B; D017346:Protein Serine-Threonine Kinases; D012319:RNA Polymerase II