The protein kinase C system acts through the early growth response protein 1 to increase LHbeta gene expression in synergy with steroidogenic factor-1.
Expression of the LHbeta gene has been shown to be modulated by both the orphan nuclear receptor, steroidogenic factor-1 (SF-1), and the early growth response protein 1, Egr-1. It is also well known that LHbeta mRNA levels are increased after hormonal activation of the protein kinase C (PKC) signaling system, for example by GnRH; however, the mechanisms by which the PKC system exerts this effect has not been fully characterized. By transient transfection of the GH3 cell line, we demonstrate that activation of the PKC system with the phorbol ester, phorbol 12-myristate 13-acetate (PMA), increases activity of region -207/+5 of the rat LHbeta gene promoter (approximately 2-fold) and markedly augments SF-1-induced stimulation (95-fold in the presence of both factors vs. 13-fold for SF-1 alone). Mutation of the two previously identified Egr-1 sites not only prevents Egr-1 effects on the LHbeta gene promoter, but also eliminates the synergistic response to PMA and SF-1 together, findings that were confirmed in a longer construct spanning region -797/+5. In the gonadotrope-derived cell line, alphaT3-1, these mutations eliminate the GnRH responsiveness of the -207/+5 LHbeta promoter construct. We next show that PMA treatment (GH3 and alphaT3-1 cells) or GnRH treatment (alphaT3-1 cells) induces expression of Egr-1, as detected by Egr-1 interaction with Egr-1 DNA-binding sites in the rat LHbeta gene promoter sequence. Furthermore, we demonstrate that PMA increases steady-state Egr-1 mRNA levels via increased Egr-1 transcription. We conclude that PMA-induced stimulation of LHbeta gene expression is achieved, at least in part, by induction of Egr-1 expression.
Halvorson
L M
LM
Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, USA. lisa.halvorson@es.nemc.org
Kaiser
U B
UB
Chin
W W
WW
D000818:Animals; D001665:Binding Sites; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D051766:Early Growth Response Protein 1; D004789:Enzyme Activation; D051717:Fushi Tarazu Transcription Factors; D005786:Gene Expression Regulation / Q000187:drug effects; D007987:Gonadotropin-Releasing Hormone / Q000378:metabolism / Q000494:pharmacology; D018398:Homeodomain Proteins; D017874:Immediate-Early Proteins; D007986:Luteinizing Hormone / Q000187:drug effects / Q000235:genetics*; D051379:Mice; D009154:Mutation; D011401:Promoter Regions, Genetic; D011493:Protein Kinase C / Q000378:metabolism*; D012333:RNA, Messenger / Q000187:drug effects / Q000378:metabolism; D051381:Rats; D018160:Receptors, Cytoplasmic and Nuclear; D054339:Steroidogenic Factor 1; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014157:Transcription Factors / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*
D004268:DNA-Binding Proteins; D051766:Early Growth Response Protein 1; C486109:Egr1 protein, mouse; C486110:Egr1 protein, rat; D051717:Fushi Tarazu Transcription Factors; D018398:Homeodomain Proteins; D017874:Immediate-Early Proteins; D012333:RNA, Messenger; D018160:Receptors, Cytoplasmic and Nuclear; D054339:Steroidogenic Factor 1; D014157:Transcription Factors; C515058:steroidogenic factor 1, mouse; C515060:steroidogenic factor 1, rat; D007987:Gonadotropin-Releasing Hormone; D007986:Luteinizing Hormone; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate