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Inhibition of PI 3‐kinase and RAS blocks IGF‐I and insulin‐induced uncoupling protein 1 gene expression in brown adipocytes
Author(s) -
Teruel Teresa,
Valverde Angela M.,
Navarro Paloma,
Benito Manuel,
Lorenzo Margarita
Publication year - 1998
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/(sici)1097-4652(199807)176:1<99::aid-jcp12>3.0.co;2-j
Subject(s) - transactivation , biology , insulin , protein kinase a , thermogenin , medicine , endocrinology , chimeric gene , kinase , chemistry , gene expression , microbiology and biotechnology , brown adipose tissue , adipose tissue , biochemistry , gene
Fetal brown adipocytes expressed uncoupling protein 1 (UCP1) mRNA, this expression being blunted throughout culture for 24 h in a serum‐free medium. At physiological doses, either insulin‐like growth factor I (IGF‐I) or insulin turned out to be as potent as dibutyryl cAMP (dbcAMP) in increasing UCP1 gene transcription rate (1 h) and also UCP1 mRNA accumulation (3 h), their maximal effect (15‐fold increase) reached upon treatment for 24 h. Upon treatment with either IGF‐I or insulin for 48 h, a 7‐fold increase in the UCP1 protein content relative to levels in the control cells was found, this induction being abolished in the presence of cycloheximide. Moreover, either IGF‐I or insulin transactivates the UCP1‐chloramphenicol acetyl transferase (CAT) fusion gene after transient transfection of primary brown adipocytes, these effects being tissue‐specific. Transient transfection of dominant‐negative form of phosphatidylinositol (PI) 3‐kinase completely blocked the transactivation of the fusion gene UCP1‐CAT induced by either IGF‐I or insulin, although inhibition of p70 S6kinase with rapamycin does not preclude transactivation of the UCP1 promoter by insulin. Furthermore, transient transfection of dominant‐negative form of p21‐ras or treatment of cells with a mitogen‐activated protein kinase kinase (MEK‐1) inhibitor (PD098059) completely abolished insulin‐induced UCP1‐CAT transactivation. Cotransfectionwith dominant‐negative p85 or with dominant‐negative Ras also produced down‐regulation of the insulin or IGF‐I‐induced 12‐O‐tetradecanoylphorbol‐13‐acetate response element (TRE)‐CAT (five AP‐1, activating protein‐1, binding sites arranged in tandem) transactivation. In addition, insulin induced AP‐1 DNA binding activity, this effect being totally prevented in the presence of MEK‐1 inhibitor. These results strongly suggest that either IGF‐I or insulin induced thermogenic‐differentiation through AP‐1 activity in a PI 3‐kinase and Ras/MAPK dependent manner in brown adipocytes. J. Cell. Physiol. 176:99–109, 1998. © 1998 Wiley‐Liss, Inc.