Activation of cAMP-dependent protein kinase increases the protein level of steroidogenic factor-1.
The orphan nuclear receptor steroidogenic factor 1 (SF-1) is an essential regulator of endocrine organogenesis, sexual differentiation, and steroidogenesis. SF-1 is a transcriptional regulator of cAMP responsive genes, but the exact mechanisms by which cAMP-dependent PKA modulates SF-1 dependent transcription leading to increased steroidogenic output have not been determined. In this report the effects of PKA activation on SF-1 in living cells have been examined by the use of full-length SF-1 cDNA fused to the cDNA encoding green fluorescent protein (GFP). The GFP-SF-1 fusion protein localized to the nucleus of both steroidogenic Y1 cells and nonsteroidogenic COS-1 cells, and the functional properties of wild-type SF-1 were conserved. When the catalytic subunit of PKA was coexpressed with GFP-SF-1, we observed that the fluorescence emission was markedly elevated. These findings were confirmed by Western blot analysis, showing that stimulation of PKA increased SF-1 protein levels. The PKA- induced expression of SF-1 protein was not accompanied by an increase in SF-1 mRNA levels. However, pulse-chase studies showed a decrease in SF-1 degradation rate in response to activation of PKA, indicating that PKA elevates the level of SF-1 by increasing the stability of SF-1 protein.
Aesøy
Reidun
R
Department of Anatomy and Cell Biology, University of Bergen, Aarstadveien 19, N-5009 Bergen, Norway. reidun.asoy@pki.uib.no
Mellgren
Gunnar
G
Morohashi
Ken-Ichirou
K
Lund
Johan
J
D000818:Animals; D019556:COS Cells; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004355:Drug Stability; D004789:Enzyme Activation / Q000502:physiology; D051717:Fushi Tarazu Transcription Factors; D049452:Green Fluorescent Proteins; D018398:Homeodomain Proteins; D008164:Luminescent Proteins / Q000235:genetics; D051379:Mice; D009154:Mutation; D010766:Phosphorylation; D012333:RNA, Messenger / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D054339:Steroidogenic Factor 1; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D004268:DNA-Binding Proteins; D051717:Fushi Tarazu Transcription Factors; D018398:Homeodomain Proteins; D008164:Luminescent Proteins; D012333:RNA, Messenger; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D054339:Steroidogenic Factor 1; D014157:Transcription Factors; C515058:steroidogenic factor 1, mouse; D049452:Green Fluorescent Proteins; D017868:Cyclic AMP-Dependent Protein Kinases