Regulation of the human general transcription initiation factor TFIIF by phosphorylation.
The transcription initiation factor, TFIIF, is essential not only for the initiation of transcription but also for efficient elongation of mRNA synthesis by mammalian RNA polymerase II and is extensively phosphorylated in vivo. The possible regulation of TFIIF activity by protein phosphorylation was investigated by comparing the biochemical properties of alkaline phosphatase-treated HeLa TFIIF with those of native or bacterially expressed factor. Alkaline phosphatase treatment decreased the size of the large subunit (RAP74) of TFIIF to that of the recombinant protein but did not change the size of the small subunit (RAP30). Both the transcription initiation and elongation stimulating activities of the alkaline phosphatase-treated TFIIF decreased to 15-20% of the native form under conditions in which the amount of TFIIF was rate-limiting for transcription. Furthermore, phosphatase-treated TFIIF assembled the DBPolF complex and bound to RNA polymerase II less efficiently than the native protein. When hybrid TFIIFs were reconstituted using native or recombinant subunits, a native form of RAP74 stimulated both transcription and DBPolF complex formation activity regardless of whether native or recombinant RAP30 was used. We propose that TFIIF activity is regulated by protein phosphorylation, particularly of the RAP74 subunit. The functional role of RAP74 in assembling the preinitiation complex and modulating TFIIF activity is discussed.
Department of Molecular Genetics, Tokyo Medical and Dental University, Japan.
S
S
Kitajima
T
T
Chibazakura
M
M
Yonaha
Y
Y
Yasukochi
D000469:Alkaline Phosphatase / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D010766:Phosphorylation; D012319:RNA Polymerase II / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D019473:Transcription Factors, TFII
D014157:Transcription Factors; D019473:Transcription Factors, TFII; D012319:RNA Polymerase II; D000469:Alkaline Phosphatase; C056604:transcription factor TFIIF