TY - JOUR
T1 - Formation and fate of a complete 31-protein RNA polymerase II transcription preinitiation complex
AU - Murakami, Kenji
AU - Calero, Guillermo
AU - Brown, Christopher R.
AU - Liu, Xin
AU - Davis, Ralph E.
AU - Boeger, Hinrich
AU - Kornberg, Roger D.
PY - 2013/3/1
Y1 - 2013/3/1
N2 - Whereas individual RNA polymerase II (pol II)-general transcription factor (GTF) complexes are unstable, an assembly of pol II with six GTFs and promoter DNA could be isolated in abundant homogeneous form. The resulting complete pol II transcription preinitiation complex (PIC) contained equimolar amounts of all 31 protein components. An intermediate in assembly, consisting of four GTFs and promoter DNA, could be isolated and supplemented with the remaining components for formation of the PIC. Nuclease digestion and psoralen crosslinking mapped the PIC between positions -70 and -9, centered on the TATA box. Addition of ATP to the PIC resulted in quantitative conversion to an open complex, which retained all 31 proteins, contrary to expectation from previous studies. Addition of the remaining NTPs resulted in run-off transcription, with an efficiency that was promoter-dependent and was as great as 17.5% with the promoters tested.
AB - Whereas individual RNA polymerase II (pol II)-general transcription factor (GTF) complexes are unstable, an assembly of pol II with six GTFs and promoter DNA could be isolated in abundant homogeneous form. The resulting complete pol II transcription preinitiation complex (PIC) contained equimolar amounts of all 31 protein components. An intermediate in assembly, consisting of four GTFs and promoter DNA, could be isolated and supplemented with the remaining components for formation of the PIC. Nuclease digestion and psoralen crosslinking mapped the PIC between positions -70 and -9, centered on the TATA box. Addition of ATP to the PIC resulted in quantitative conversion to an open complex, which retained all 31 proteins, contrary to expectation from previous studies. Addition of the remaining NTPs resulted in run-off transcription, with an efficiency that was promoter-dependent and was as great as 17.5% with the promoters tested.
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U2 - 10.1074/jbc.M112.433623
DO - 10.1074/jbc.M112.433623
M3 - Article
C2 - 23303183
AN - SCOPUS:84874770600
SN - 0021-9258
VL - 288
SP - 6325
EP - 6332
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 9
ER -