The cytosol as site of phosphorylation of the cyclic AMP-dependent protein kinase in adrenal steroidogenesis.
The mitochondria, the microsomes and the cytosol have been described as possible sites of cAMP-dependent phosphorylation. However, there has been no direct demonstration of a cAMP-dependent kinase associated with the activation of the side-chain cleavage of cholesterol. We have investigated the site of action of the cAMP-dependent kinase using a sensitive cell-free assay. Cytosol derived from cells stimulated with ACTH or cAMP was capable of increasing progesterone synthesis in isolated mitochondria when combined with the microsomal fraction. Cytosol derived from cyclase or kinase of negative mutant cells did not. Cyclic AMP and cAMP-dependent protein kinase stimulated in vitro a cytosol derived from unstimulated adrenal cells. This cytosol was capable of stimulating progesterone synthesis in isolated mitochondria. Inhibitor of cAMP-dependent protein kinase abolished the effect of the cAMP. ACTH stimulation of cytosol factors is a rapid process observable with a half maximal stimulation at about 3 pM ACTH. The effect was also abolished by inhibitor of arachidonic acid release. The function of cytosolic phosphorylation is still unclear. The effect of inhibitors of arachidonic acid release, and the necessity for the microsomal compartment in order to stimulate mitochondrial steroidogenesis, suggest that the factor in the cytosol may play a role in arachidonic acid release.
Dada
L A
LA
Department of Biochemistry, School of Medicine, University of Buenos Aires, Argentina.
Paz
C
C
Mele
P
P
Solano
A R
AR
Cornejo Maciel
F
F
Podesta
E J
EJ
D000305:Adrenal Cortex Hormones / Q000096:biosynthesis*; D000324:Adrenocorticotropic Hormone / Q000494:pharmacology; D000818:Animals; D002460:Cell Line; D003600:Cytosol / Q000201:enzymology / Q000378:metabolism*; D008297:Male; D008928:Mitochondria / Q000378:metabolism; D010766:Phosphorylation; D011374:Progesterone / Q000096:biosynthesis; D011494:Protein Kinases / Q000378:metabolism*; D051381:Rats; D015383:Zona Fasciculata / Q000201:enzymology / Q000378:metabolism*
D000305:Adrenal Cortex Hormones; D011374:Progesterone; D000324:Adrenocorticotropic Hormone; D011494:Protein Kinases