HEXIM1 forms a transcriptionally abortive complex with glucocorticoid receptor without involving 7SK RNA and positive transcription elongation factor b.
The HEXIM1 protein has been shown to form a protein-RNA complex composed of 7SK small nuclear RNA and positive transcription elongation factor b (P-TEFb), which is composed of cyclin-dependent kinase 9 (CDK9) and cyclin T1, and to inhibit the kinase activity of CDK9, thereby suppressing RNA polymerase II-dependent transcriptional elongation. Here, we biochemically demonstrate that HEXIM1 forms a distinct complex with glucocorticoid receptor (GR) without RNA, CDK9, or cyclin T1. HEXIM1, through its arginine-rich nuclear localization signal, directly associates with the ligand-binding domain of GR. Introduction of HEXIM1 short interfering RNA and adenovirus-mediated exogenous expression of HEXIM1 positively and negatively modulated glucocorticoid-responsive gene activation, respectively. In the nucleus, HEXIM1 was shown to localize in a distinct compartment from that of the p160 coactivator transcriptional intermediary factor 2. Overexpression of HEXIM1 decreased ligand-dependent association between GR and transcriptional intermediary factor 2. Antisense-mediated disruption of 7SK blunted the negative effect of HEXIM1 on arylhydrocarbon receptor-dependent transcription but not on GR-mediated one, indicating that a class of transcription factors are direct targets of HEXIM1. These results indicate that HEXIM1 has dual roles in transcriptional regulation: inhibition of transcriptional elongation dependent on 7SK RNA and positive transcription elongation factor b and interference with the sequence-specific transcription factor GR via a direct protein-protein interaction. Moreover, the fact that the central nuclear localization signal of HEXIM1 is essential for both of these actions may argue the crosstalk of these functions.
Shimizu
Noriaki
N
Division of Clinical Immunology and Department of Rheumatology and Allergy, Institute of Medical Science, University of Tokyo, 4-6-1, Shirokanedai, Minato-ku, Tokyo 108-8639, Japan.
Ouchida
Rika
R
Yoshikawa
Noritada
N
Hisada
Tetsuya
T
Watanabe
Hajime
H
Okamoto
Kensaku
K
Kusuhara
Masatoshi
M
Handa
Hiroshi
H
Morimoto
Chikao
C
Tanaka
Hirotoshi
H
D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D019084:Fluorescent Antibody Technique, Indirect; D005786:Gene Expression Regulation; D005982:Glutathione Transferase; D006367:HeLa Cells; D006801:Humans; D047468:Immunoprecipitation; D008156:Luciferases; D013058:Mass Spectrometry; D046912:Multiprotein Complexes / Q000378:metabolism*; D051958:Nuclear Receptor Coactivator 2; D020411:Oligonucleotide Array Sequence Analysis; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D034741:RNA, Small Interfering / Q000235:genetics; D016601:RNA-Binding Proteins / Q000378:metabolism*; D011965:Receptors, Glucocorticoid / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors / Q000378:metabolism; D015533:Transcriptional Activation; D014162:Transfection
C476123:HEXIM1 protein, human; D046912:Multiprotein Complexes; C498001:NCOA2 protein, human; D051958:Nuclear Receptor Coactivator 2; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins; D011965:Receptors, Glucocorticoid; D014157:Transcription Factors; D008156:Luciferases; D005982:Glutathione Transferase