Trypsin stimulation of aldosterone and 18-hydroxycorticosterone production by rat adrenal zona glomerulosa tissue is mediated by activation of protein kinase C.
When rat adrenal whole capsules, containing the zona glomerulosa, were incubated, addition of the protein kinase C inhibitors TMB-8 (10 mumol/l), W7, H7, polymyxin-B and sphingosine (all 1 mumol/l) was found to inhibit the steroidogenic response to trypsin. Aldosterone and 18-hydroxycorticosterone were strongly, and corticosterone moderately, affected, while the production of 18-hydroxydeoxy-corticosterone was neither stimulated by trypsin nor inhibited by the protein kinase C inhibitors. Addition of neomycin, which prevents substrate interaction with phospholipase C, also inhibited the response to trypsin, while addition of phospholipase C itself stimulated aldosterone, 18-hydroxycorticosterone and corticosterone production with the same tissue sensitivity as trypsin. Addition of phospholipase A2 had no effect. Direct assay of protein kinase C activity showed that trypsin stimulation effected the translocation of Ca2+/phospholipid-activated protein kinase C from the cytosolic to the membrane fraction. When glomerulosa tissue was incubated with [32P]ATP, and cytosolic proteins were subjected to isoelectric focusing on polyacrylamide gels, autoradiography showed that incorporation of 32P into several protein components was increased by trypsin stimulation. It was concluded that trypsin exerts its stimulatory effects on steroidogenesis by activating protein kinase C; not, however, by generating the Ca2+/phospholipid-independent fragment, but possibly by enhancing the activity of phospholipase C.
Department of Biochemistry, Medical College of St Bartholomew's Hospital, London.
G P
GP
Vinson
S M
SM
Laird
J P
JP
Hinson
N
N
Mallick
S
S
Marsigliante
R
R
Teja
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D015069:18-Hydroxycorticosterone / Q000378:metabolism*; D000450:Aldosterone / Q000096:biosynthesis*; D000818:Animals; D003345:Corticosterone / Q000031:analogs & derivatives* / Q000096:biosynthesis; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D005707:Gallic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D006801:Humans; D007525:Isoelectric Focusing; D007546:Isoquinolines / Q000494:pharmacology; D008297:Male; D010879:Piperazines / Q000494:pharmacology; D011112:Polymyxin B / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D013449:Sulfonamides / Q000494:pharmacology; D014357:Trypsin / Q000494:pharmacology*; D010738:Type C Phospholipases / Q000494:pharmacology; D015384:Zona Glomerulosa / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*
D007546:Isoquinolines; D010879:Piperazines; D013449:Sulfonamides; D000450:Aldosterone; D015069:18-Hydroxycorticosterone; C006014:8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate; D005707:Gallic Acid; C017967:W 7; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D011493:Protein Kinase C; D010738:Type C Phospholipases; D014357:Trypsin; D011112:Polymyxin B; D003345:Corticosterone