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Human transcription elongation factor NELF: Identification of novel subunits and reconstitution of the functionally active complex
被引:177
作者:
Narita, T
Yamaguchi, Y
Yano, K
Sugimoto, S
Chanarat, S
Wada, T
Kim, DK
Hasegawa, J
Omori, M
Inukai, N
Endoh, M
Yamada, T
Handa, H
机构:
[1] Tokyo Inst Technol, Frontier Collaborat Res Ctr, Yokohama, Kanagawa 2268503, Japan
[2] Tokyo Inst Technol, Grad Sch Biosci & Biotechnol, Yokohama, Kanagawa 2268503, Japan
[3] Japan Sci & Technol Corp, PRESTO, Yokohama, Kanagawa 2268503, Japan
[4] Kyowa Hakko Kogyo Co Ltd, Tokyo Res Labs, Tokyo 1948533, Japan
关键词:
D O I:
10.1128/MCB.23.6.1863-1873.2003
中图分类号:
Q5 [生物化学];
Q7 [分子生物学];
学科分类号:
071010 ;
081704 ;
摘要:
The multisubunit transcription elongation factor NELF (for negative elongation factor) acts together with DRB (5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole) sensitivity-inducing factor (DSIF)/human Spt4-Spt5 to cause transcriptional pausing of RNA polymerase II (RNAPII). NELF activity is associated with five polypeptides, A to E. NELF-A has sequence similarity to hepatitis delta antigen (HDAg), the viral protein that binds to and activates RNAPII, whereas NELF-E is an RNA-binding protein whose RNA-binding activity is critical for NELF function. To understand the interactions of DSIF, NELF, and RNAPII at a molecular level, we identified the B, C, and D proteins of human NELF. NELF-B is identical to COBRA1, recently reported to associate with the product of breast cancer susceptibility gene BRCA1 NELF-C and NELF-D are highly related or identical to the protein called TH1, of unknown function. NELF-B and NELF-C or NELF-D are integral subunits that bring NELF-A and NELF-E together, and coexpression of these four proteins in insect cells resulted in the reconstitution of functionally active NELF. Detailed analyses using mutated recombinant complexes indicated that the small region of NELF-A with similarity to HDAg is critical for RNAPII binding and for transcriptional pausing. This study defines several important protein-protein interactions and opens the way for understanding the mechanism of DSIF- and NELF-induced transcriptional pausing.
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页码:1863 / 1873
页数:11
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