title: Activation of MAPK cascades by GnRH: ERK and Jun N-terminal kinase are involved in basal and GnRH-stimulated activity of the glycoprotein hormone LHbeta-subunit promoter.
abstract: The role of ERK and Jun N-terminal kinase (JNK) in basal- and GnRH-stimulated LHbeta-promoter activity was examined in the gonadotroph cell line LbetaT-2. GnRH agonist (GnRH-A) stimulates the MAPK cascades ERK, JNK, and p38MAPK, with a peak at 7 min for ERK and at 60 min for JNK and p38MAPK. The rat glycoprotein hormone LHbeta-subunit promoter, linked to the chloramphenicol acetyl transferase (CAT) reporter gene, was used to follow its activation. Addition of GnRH-A (10 nM) to LbetaT-2 cells resulted in a 6-fold increase in LHbeta-CAT activity at 8 h, which was markedly reduced by a GnRH antagonist. The PKC activator 12-O-tetradecanoylphorbol-13-acetate (TPA), but not the Ca(2+) ionophore ionomycin, stimulated LHbeta-CAT activity. Addition of GnRH-A and TPA together did not produce an additive response. Down-regulation of PKC, but not removal of Ca(2+), abolished the GnRH-A and the TPA response. Cotransfection of the LHbeta-promoter and the constitutively active form of Raf-1 stimulated basal and GnRH-A-induced LHbeta-CAT activity. The dominant negative forms of the ERK cascade members Ras, Raf-1, and MAPK/ERK kinase (MEK) markedly reduced basal and GnRH-A-induced LHbeta-CAT activity, Similar results were obtained with the MEK inhibitor PD 098059. Cotransfection of the LHbeta-promoter and the constitutively active CDC42 stimulated basal and GnRH-A-induced LHbeta-CAT activity. The dominant negative forms of the JNK cascade members Rac, CDC42, and SEK markedly diminished basal and GnRH-A-induced LHbeta-CAT activity. Interestingly, the constitutively active form of c-Src stimulated the basal and the GnRH-A response, whereas the dominant negative form of c-Src, or the c-Src inhibitor PP1 diminished basal and the GnRH-A response. We conclude that ERK and JNK are involved in basal and GnRH-A stimulation of LHbeta-CAT activity. c-Src participates also in LHbeta-promoter activation by a mechanism which might be linked to ERK and JNK activation.
authors.affiliation: Department of Biochemistry, The George S. Wise Faculty of Life Sciences, Tel Aviv University, Ramat Aviv 69978, Israel.
authors.forename: Dagan
authors.initials: D
authors.lastname: Harris
authors.forename: David
authors.initials: D
authors.lastname: Bonfil
authors.forename: Dana
authors.initials: D
authors.lastname: Chuderland
authors.forename: Sarah
authors.initials: S
authors.lastname: Kraus
authors.forename: Rony
authors.initials: R
authors.lastname: Seger
authors.forename: Zvi
authors.initials: Z
authors.lastname: Naor
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000187:drug effects; D016391:Genes, src / Q000235:genetics; D007987:Gonadotropin-Releasing Hormone / Q000502:physiology*; D048031:JNK Mitogen-Activated Protein Kinases; D007986:Luteinizing Hormone / Q000096:biosynthesis* / Q000235:genetics*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism* / Q000502:physiology*; D011493:Protein Kinase C / Q000378:metabolism; D051381:Rats; D014162:Transfection
chemical_list: D007987:Gonadotropin-Releasing Hormone; D007986:Luteinizing Hormone; D011493:Protein Kinase C; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D002118:Calcium