Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.6.3.14 (ATP synthase)
 7,042 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The regulation of cholesterol side-chain cleavage enzyme (P450scc) by glucocorticoids was investigated in mouse Leydig cell cultures. We recently demonstrated that P450scc is constitutively synthesized in Leydig cells and that the rate of P450scc synthesis is increased by chronic treatment of the cultures with 8-bromo-cAMP. We now report that glucocorticoids, specifically, decrease the constitutive and cAMP-induced synthesis of P450scc protein as well as the accumulation of P450scc mRNA. The treatment of cultures with as little as 10 nM dexamethasone resulted in a 50-60% decrease in the rate of synthesis of P450scc protein and mRNA content. The glucocorticoid-mediated decrease in P450scc synthesis was prevented when cultures were treated with the antiglucocorticoid RU-486. RU-486 alone had no effect on the rate of protein synthesis. The effect was specific for glucocorticoids; corticosterone (100 nM) or cortisol (100 nM) brought about a similar decrease as dexamethasone. Treatment of cultures with the progesterone agonist R5020 (100 nM), testosterone (2 microM), or estradiol (50 nM) had no effect on the rate of specific protein synthesis. The synthesis of iron sulfur protein reductase (ISP-reductase) and F1-ATPase were not affected by dexamethasone, indicating that the effect was specific for P450scc. The amount of P450scc mRNA was decreased 61% by dexamethasone and increased 144% by treatment with 8-bromo-cAMP. These data together with our previous finding on the negative regulation of P450(17 alpha) protein synthesis by testosterone suggest that the steroidogenic P450 enzymes in Leydig cells are negatively regulated by steroid hormones acting via their cognate receptors.
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PMID:Glucocorticoid-mediated repression of P450scc mRNA and de novo synthesis in cultured Leydig cells. 253 67

To investigate the basis for the pattern of ovarian steroid production during the bovine estrous cycle, the tissue concentrations of major steroidogenic enzymes, 17 alpha-hydroxylase cytochrome P-450 and cholesterol side-chain cleavage cytochrome P-450 (cytochrome P-450scc), and their respective electron donors, NADPH-cytochrome P-450 reductase and adrenodoxin, were estimated and compared with those of the nonsteroidogenic enzymes, cytochrome c oxidase and F1-ATPase. The levels of these enzymes were estimated in medium sized (9-11 mm) and large (14-18 mm) follicles after removal of follicular fluid by centrifugation, and corpora lutea from the early, early-mid late-mid, and late stages of the luteal phase (n = 5 per group). The specific contents of all enzymes and electron donors were determined by immunoblot analysis, except for cytochrome c oxidase, which was quantified by determination of specific activity. The specific (per microgram of tissue homogenate protein) and total (per follicle or corpus luteum) tissue contents of 17 alpha-hydroxylase cytochrome P-450 increased 0.5- and 5-fold respectively from medium sized to large follicles, but then decreased to undetectable levels in corpora lutea in the early luteal phase, and remained undetectable throughout the luteal phase. In contrast, the specific content of NADPH-cytochrome P-450 reductase was similar between follicles and corpora lutea. The specific contents of cytochrome P-450scc, adrenodoxin and cytochrome c oxidase in follicles were similar to those of corpora lutea of the early luteal phase. However, by the early-mid luteal phase the specific contents of luteal cytochrome P-450scc (490 +/- 46 vs. 5709 +/- 982 cpm/micrograms protein) and adrenodoxin (44 +/- 15 vs. 705 +/- 229 cpm/micrograms protein) were increased, by 12- and 15-fold, respectively (P less than 0.05). In contrast, cytochrome c oxidase activity (29.1 +/- 10.1 vs. 108.6 +/- 20.7 nmol/mg tissue protein X min) and the specific content of F1-ATPase increased only 3- to 4-fold reflective of an increase in numbers of mitochondria. The levels of these enzymes remained elevated until the late luteal phase when they declined markedly. It is concluded that the induction of synthesis of P-450scc and adrenodoxin after ovulation is specific and does not merely reflect biogenesis of mitochondria during luteinization. Moreover the changes in the types of steroids produced by the ovarian compartments throughout the estrous cycle are a reflection of changes in the tissue content of steroidogenic enzymes.
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PMID:Cytochromes P-450scc, P-450(17)alpha, adrenodoxin, and reduced nicotinamide adenine dinucleotide phosphate-cytochrome P-450 reductase in bovine follicles and corpora lutea. Changes in specific contents during the ovarian cycle. 300 11

The effects of FSH and (Bu)2cAMP on synthesis of the components of the cholesterol side-chain cleavage (SCC) enzyme complex, namely SCC cytochrome P-450 (P-450scc), the iron-sulfur protein adrenodoxin (ISP), and NADPH:ISP reductase (Red), were investigated in granulosa cells obtained from ovaries of immature estrogen-primed rats cultured for up to 72 h in defined medium in the presence or absence of FSH and (Bu)2cAMP. The cells were lysed, and proteins were subjected to polyacrylamide gel electrophoresis, followed by immunoblotting using antibodies specific to bovine adrenocortical P-450scc, ISP, and Red. A time-dependent increase was observed in the specific contents of these three components of SCC, but not of the reference mitochondrial protein, F1-ATPase, upon treatment with FSH or (Bu)2cAMP. The increase in the content of these three enzymes was accompanied by a rise in progesterone and 20 alpha-hydroxyprogesterone production. The synthesis of P-450scc, ISP, and Red increased 3- to 4-fold with time upon FSH or (Bu)2cAMP treatment respectively, as evidenced by pulse labeling of the cell proteins with [35S]methionine, followed by immunoprecipitation. Immunoprecipitation of P-450scc and ISP from an in vitro translation system programmed by RNA isolated from cultured cells revealed that treatment with FSH or (Bu)2cAMP resulted in an increase in the levels of translatable mRNA specific for these proteins, and that the initial products of translation were precursor forms of cytochrome P-450scc and ISP, similar to those observed in bovine adrenal and granulosa cells. It is concluded that in cultured rat ovarian granulosa cells, FSH induces the synthesis of cytochrome P-450scc, ISP, and Red by increasing the content of translatable mRNA coding for the precursor forms of these enzymes and that this action is mediated by cAMP. Furthermore, the effects of FSH and (Bu)2cAMP provide an explanation for the action of these compounds to stimulate progestin synthesis in cultured ovarian cells.
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PMID:Synthesis of the cholesterol side-chain cleavage enzymes in cultured rat ovarian granulosa cells: induction by follicle-stimulating hormone and dibutyryl adenosine 3',5'-monophosphate. 301 93

The effect of epidermal growth factor (EGF) on the synthesis of the components of the cholesterol side-chain cleavage enzyme complex (SCC) was studied in rat ovarian granulosa cells. The cells were cultured for 48 h in the presence or absence of EGF (15 ng/ml) and/or FSH (50 ng/ml) after which proteins were radiolabeled by incubation with [35S]methionine followed by immunoprecipitation of newly synthesized P-450scc or adrenodoxin (ISP) with polyclonal antibodies directed against the corresponding proteins from bovine adrenal cortex. In addition the action of EGF on the level of translatable RNA for P-450scc was evaluated using a cell-free translation system programmed with RNA isolated from treated and untreated cells, followed by immunoisolation of newly synthesized proteins. Immunoisolated proteins were separated by polyacrylamide-gel electrophoresis, visualized by fluorography and quantified by densitometry. EGF stimulated progesterone formation by the cells 3-fold and potentiated the FSH-induced stimulation of progesterone formation, but had no effect on cAMP accumulation. EGF also stimulated the synthesis of P-450scc and ISP, and enhanced the FSH-induced synthesis of P-450scc and ISP in a concentration-dependent fashion with a maximal stimulation attained at concentrations ranging from 1.0 to 100 ng/ml. No appreciable changes in the induction pattern were observed when EGF and dibutyryl cyclic AMP (Bt2cAMP) were added together, as compared to when Bt2cAMP was added alone. Neither treatment affected the synthesis of the constitutive mitochondrial enzyme, F1-ATPase. Immunoisolation of P-450scc from the proteins synthesized in a rabbit reticulocyte in vitro translation system programmed with RNA isolated from EGF- and/or FSH-treated cells, revealed that EGF enhanced the FSH-stimulated synthesis of the precursor form of P-450scc.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Effects of epidermal growth factor on the synthesis of the cholesterol side-chain cleavage enzyme complex in rat ovarian granulosa cells in primary culture. 349 31

To clarify whether ATP synthase and side chain cleavage enzymes (SCC) can coexist in a mitochondrion of the adrenal cortical cell, and whether the functional heterogeneity of mitochondria can occur in an adrenal cortical cell, the immunostainability of these two enzymes in mitochondria was compared by electron microscope using pre- and post-embedding methods. In the pre-embedding method, the inner membrane of most mitochondria in the adrenal cortical cell was positively stained for SCC, 11 beta-hydroxylase, and ATP synthase, but among these immunopositive mitochondria we often observed negatively immunoreacted ones in the same adrenal cortical cell. In the positively stained mitochondria, immunoreaction products were evenly localized along the inner mitochondrial membrane. We therefore think that these differences in the immunostainability are caused by technical artifacts, namely the inadequate penetration of the antibody. In the post-embedding immunocytochemistry, gold particles showing the presence of the enzymes were observed on all mitochondria. Two different antibodies, anti-ATP synthase antibody and anti-cytochrome P-450scc antibody, which were labelled with gold particles of varying diameter (5 nm, 10 nm each) could be observed on all the mitochondria of the bovine adrenal cortical cells. No heterogeneities as to the stainability for both ATP synthase and SCC were detected in mitochondria of the rat and bovine adrenal cortical cells. These results indicate that in the rat and bovine adrenal cortical cells, both ATP synthase and SCC are evenly and simultaneously present on the inner membrane of all mitochondria. Hence, mitochondria in the adrenal cortical cell seem to be homogeneous in their steroid and ATP synthesizing abilities.
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PMID:Electron microscopic immunocytochemistry on the colocalization of ATP synthase and cytochrome P-450 of side chain cleavage enzymes in mitochondria of rat and bovine adrenal cortical cells. 837 64