Novel function of the transactivation domain of a pituitary-specific transcription factor, Pit-1.
Pit-1 stimulates the expression of growth hormone, prolactin, and thyrotropin beta subunit genes. Consequently, abnormality of the Pit-1 gene results in combined pituitary hormone deficiency (CPHD). In this study, we analyzed the function of Pit-1 with a mutation (proline to leucine at codon 24) in the transactivation domain, P24L, which has a normal POU domain important for binding to DNA, because this mutation had been reported in a patient with CPHD. We found that codon 24 proline in the transactivation domain as well as the POU domain of Pit-1 was crucial to recruit coactivator CREB-binding protein (CBP) in the cultured cells. P24L completely lost the responsiveness to cAMP to stimulate the expression of the Pit-1-targeted genes. Furthermore, CBP and Pit-1, but not P24L, markedly enhanced the expression of the Pit-1-targeted gene to cAMP, and adenovirus E1a that binds to CBP and abrogates its function blocked the induction by cAMP of Pit-1-stimulated gene transcription in the pituitary-derived GH3 cells. These results suggest that CBP and proline at codon 24 in the transactivation domain of Pit-1 are important for the cAMP-induced activation of Pit-1-targeted genes. However, P24L maintained basal transcriptional activity, suggesting that CBP is unlikely to be an essential coactivator for Pit-1.
Division of Endocrinology/Metabolism, Neurology and Hematology/Oncology, Department of Clinical Molecular Medicine, Kobe University Graduate School of Medicine, Kobe University School of Medicine, 7-10-2, Tomogaoka, Suma-ku, Kobe 654-0142, Japan.
Masahiko
M
Kishimoto
Yasuhiko
Y
Okimura
Kazuhiro
K
Yagita
Genzo
G
Iguchi
Mariko
M
Fumoto
Keiji
K
Iida
Hidesuke
H
Kaji
Hitoshi
H
Okamura
Kazuo
K
Chihara
D000818:Animals; D019556:COS Cells; D050882:CREB-Binding Protein; D002675:Child, Preschool; D000242:Cyclic AMP / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D005260:Female; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D013006:Growth Hormone / Q000172:deficiency / Q000235:genetics / Q000378:metabolism; D006801:Humans; D046911:Macromolecular Substances; D008297:Male; D009154:Mutation; D009687:Nuclear Proteins / Q000378:metabolism; D010902:Pituitary Gland / Q000378:metabolism*; D011388:Prolactin / Q000172:deficiency / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015290:Second Messenger Systems / Q000502:physiology; D013972:Thyrotropin / Q000172:deficiency; D015534:Trans-Activators / Q000378:metabolism; D050817:Transcription Factor Pit-1; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D014158:Transcription, Genetic; D020798:Two-Hybrid System Techniques
D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D009687:Nuclear Proteins; C494187:POU1F1 protein, human; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; D050817:Transcription Factor Pit-1; D014157:Transcription Factors; D011388:Prolactin; D013972:Thyrotropin; D013006:Growth Hormone; D000242:Cyclic AMP; D050882:CREB-Binding Protein; C494363:CREBBP protein, human