The interaction of TRbeta1-N terminus with steroid receptor coactivator-1 (SRC-1) serves a full transcriptional activation function of SRC-1.
Steroid receptor coactivator-1 (SRC-1) plays a crucial role in nuclear receptor-mediated transcription including thyroid hormone receptor (TR)-dependent gene expression. Interaction of the TR-ligand binding domain and SRC-1 through LXXLL motifs is required for this action. However, potential interactions between the TRbeta1-N terminus (N) and SRC-1 have not been explored and thus are examined in this manuscript. Far-Western studies showed that protein construct containing TRbeta1-N + DNA binding domain (DBD) bound to nuclear receptor binding domain (NBD)-1 (amino acid residue, aa 595-780) of SRC-1 without ligand. Mammalian two-hybrid studies showed that NBD-1, as well as SRC-1 (aa 595-1440), bound to TRbeta1-N+DBD in the absence of ligand in CV-1 cells. However, NBD-2 (aa 1237-1440) did not bind to this protein. Glutathione-S-transferase pull-down studies showed that TRbeta1-N (aa 1-105) bound to the broad region of SRC-1-C terminus. Expression vectors encoding a series of truncations and/or point mutations of TRbeta1 were used in transient transfection-based reporter assays in CV-1 cells. N-terminal truncated TRbeta1 (DeltaN-TRbeta1) showed lower activity than that of wild-type in both artificial F2-thyroid hormone response element and native malic enzyme response element. These results suggest that there is the interaction between N terminus of TRbeta1 and SRC-1, which may serve a full activation of SRC-1, together with activation function-2 on TRbeta1-mediated transcription.
Iwasaki
Toshiharu
T
Department of Integrative Physiology, Gunma University Graduate School of Medicine, 3-39-22 Showa-machi, Maebashi 371-8511, Gunma, Japan. tiwasaki@med.gunma-u.ac.jp
Takeshita
Akira
A
Miyazaki
Wataru
W
Chin
William W
WW
Koibuchi
Noriyuki
N
D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D002522:Chlorocebus aethiops; D018076:DNA, Complementary / Q000378:metabolism; D005822:Genetic Vectors; D005982:Glutathione Transferase / Q000378:metabolism; D051548:Histone Acetyltransferases; D008024:Ligands; D056920:Nuclear Receptor Coactivator 1; D010957:Plasmids / Q000378:metabolism; D017354:Point Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011988:Receptors, Thyroid Hormone / Q000378:metabolism; D020218:Response Elements; D014157:Transcription Factors / Q000378:metabolism / Q000502:physiology*; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection
D018076:DNA, Complementary; D008024:Ligands; D011988:Receptors, Thyroid Hormone; D014157:Transcription Factors; D051548:Histone Acetyltransferases; D056920:Nuclear Receptor Coactivator 1; D005982:Glutathione Transferase