Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.13 (protein kinase C)
 49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The role of arachidonic acid in the regulation of steroidogenesis in rat Leydig cells was studied. A dose- and time-dependent biphasic effect on maximal and submaximal LH- and dibutyryl-cAMP-stimulated testosterone production was found. The locus of the inhibition, which occurred during 3 h incubation, was prior to the side chain cleavage of cholesterol and after cAMP production. The same inhibitory effect was found with the protein kinase C (PKC) activators, phorbol-12-myristate, 13-acetate (PMA) and oleic acid, also with no change in LH-stimulated cAMP production. Arachidonic acid, PMA, and diolein, all stimulated PKC activity in a dose-dependent fashion in partially purified Leydig cell homogenates. When the cells were incubated for 5 h, arachidonic acid potentiated LH- and dibutyryl-cAMP-stimulated testosterone production. Similarly, incubation with PMA for 5 h, potentiated subsequent basal and dibutyryl-cAMP-stimulated testosterone production. PKC was down-regulated over 5 h (but not during 3 h) by pretreating Leydig cells with PMA or arachidonic acid in the presence of LH. Lipoxygenase and cyclooxygenase inhibitors did not alter the stimulatory effects of arachidonic acid. We conclude that the short-term inhibitory effect of arachidonic acid (and PMA) is via activation of PKC, but when protein kinase C (PKC) is down-regulated by these ligands, steroidogenesis is enhanced. These results suggest that steroidogenesis is normally under tonic inhibitory control by PKC.
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PMID:Direct effect of arachidonic acid on protein kinase C and LH-stimulated steroidogenesis in rat Leydig cells; evidence for tonic inhibitory control of steroidogenesis by protein kinase C. 131 Dec 29

Luteinizing hormone (LH) interacts with its plasma membrane receptor to stimulate steroidogenesis not only via cyclic AMP but also other pathways which include arachidonic acid and leukotrienes and regulation of chloride and calcium channels. The same stimulatory pathways may lead to desensitization and down-regulation of the LH receptor and steroidogenesis. The LH receptor exists in a dynamic state, being truncated, or internalized, degraded or recycled. Desensitization is controlled by protein kinase C (PKC) in the rat and by cyclic AMP dependent protein kinase and PKC in the mouse Leydig cells. Using an adapted anti-sense oligonucleotide strategy we have shown that the cytoplasmic C-terminal sequence of the LH receptor is essential for desensitization to occur. In contrast, these sequences of the LH receptor are not required for the stimulation of cyclic AMP and steroid production. We have also shown that the extracellular domain of the LH receptor is secreted from the Leydig cell and may act as a LH-binding protein.
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PMID:Control of steroidogenesis in Leydig cells. 139 Feb 94

The role of cyclic AMP and phorbol esters in luteinizing hormone (LH) receptor down-regulation in Leydig cells has been studied. Dibutyryl cyclic AMP (db-cAMP) (0.01, 0.1 and 1 mM), forskolin (80 microM) and cholera toxin (1.19 nM) caused a 30-50% loss of [125I]hCG binding sites and an inhibition of receptor-[125I]hCG complex internalization in mouse tumour Leydig (MA10, MLTC-1) cells during 2 h. In contrast, db-cAMP had no effect on the level of binding sites or internalization of the hormone receptor complex in rat testis Leydig cells or a rat tumour (R2C) Leydig cell. Phorbol 12-myristate 13-acetate (PMA) at concentrations from 10(-9) to 10(-5) M had no effect on hormone binding or hormone-receptor complex internalization in any of the Leydig cells. In contrast a 2 h preincubation of MLTC-1 cells with 10(-7) M PMA caused a loss of subsequent LH-stimulated cyclic AMP and pregnenolone production. These results indicate that LH receptor down-regulation is mediated by cyclic AMP dependent kinase, but not protein kinase C, in mouse Leydig cells. No down-regulation of rat Leydig cell LH receptor occurs with either kinase.
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PMID:Differences in LH receptor down-regulation between rat and mouse Leydig cells: effects of 3',5'-cyclic AMP and phorbol esters. 166 61

Different endocrine, steroidogenic cell types were examined for their content in protein kinase C (PKC) subtypes I, II and III. The expression of the PKC isoforms was assayed following high-performance liquid chromatography separation and characterization using a set of antipeptide antibodies specific for each enzyme isotype. Bovine and rat adrenocortical cells, as well as porcine Sertoli cells expressed only the type III PKC. By contrast, Leydig cell expressed both the isotypes I, II and II at similar levels. Taking into account the biological effect observed in these various cell types upon PKC activation, it may be suggested that the type III iso-PKC is involved in the steroidogenic activation pathways, whereas the expression of the types I, II, and III, only in Leydig cells, may contribute to a different array of cross-talk regulation pathways in these cells.
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PMID:Expression of protein kinase C isoforms in various steroidogenic cell types. 205 Feb 74

We have recently demonstrated the presence in the rat Leydig cells of a corticotropin releasing factor (CRF) receptor and an inhibitory action of the peptide on human chorionic gonadotropin (hCG)-induced cAMP generation and steroidogenesis. The inhibitory action of CRF was unaffected by pertussis toxin and was completely reversed by 8-bromo-cAMP (Ulisse, S., Fabbri, A., and Dufau, M. L. (1989) J. Biol. Chem. 264, 2156-2163). In this study, we have evaluated the participation of protein kinase C in CRF action in the Leydig cells and the level of the gonadotropin signal pathway affected by CRF. Binding of 125I-labeled ovine CRF to Leydig cell membranes was reduced by GTP and guanyl-5'-yl imidodiphosphate (Gpp(NH)p), in a dose-dependent manner. Phorbol 12-myristate 13-acetate, like CRF, caused time-dependent inhibition of hCG-induced cAMP generation and steroidogenesis. This inhibitory action was reversed by 8-bromo-cAMP. Both CRF and 12-O-tetradecanoylphorbol-13-acetate did not affect 125I-hCG binding. No additive effects of CRF and the phorbol ester were observed in these studies. CRF caused a rapid translocation of protein kinase C in Leydig cells. Preincubation of cells with protein kinase C inhibitors or TPA-induced depletion of protein kinase C prevented the inhibitory actions of CRF and TPA. CRF and TPA were able to inhibit the stimulation of cAMP and testosterone production by cholera toxin and forskolin. Adenylate cyclase stimulation by Gpp(NH)p, luteinizing hormone + Gpp(NH)p, and NaF in crude membranes or by forskolin and manganese in solubilized membranes, prepared from CRF- and TPA-treated cells, was also markedly inhibited. We conclude that CRF receptors interact with a pertussis toxin-insensitive G protein (possibly Gp) in the Leydig cell and that the inhibitory action of CRF on Leydig cell function is exerted mainly on the catalytic subunit of adenylate cyclase through a direct or indirect action of protein kinase C.
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PMID:A novel mechanism of action of corticotropin releasing factor in rat Leydig cells. 215 73

Serum-free primary cultures of neonatal (1-day-old) porcine Leydig cells were used to study the effects of phorbol-12-myristate-13-acetate and 1,2-dioctanoylglycerol on testosterone and pregnenolone production. Phorbol-12-myristate-13-acetate alone from 0.001-10 mumol/l stimulated testosterone and pregnenolone production, whereas 1,2-dioctanoylglycerol alone had no effect on steroid production, relative to control. Phorbol-12-myristate-13-acetate and 1,2-dioctanoylglycerol each inhibited pLH-stimulated testosterone and pregnenolone production. To further clarify the influence of these protein kinase C activators on steroidogenesis, cultured Leydig cells were treated with either phorbol-12-myristate-13-acetate or 1,2-dioctanoylglycerol plus forskolin (an adenylate cyclase activator). Both phorbol-12-myristate-13-acetate and 1,2-dioctanoylglycerol inhibited forskolin-stimulated testosterone production. Phorbol-12-myristate-13-acetate had no effect on forskolin-stimulated pregnenolone production and only the highest concentration of 1,2-dioctanoylglycerol (100 mumol/l) inhibited forskolin-stimulated production of pregnenolone. These data demonstrate that porcine Leydig cell steroidogenesis can be modulated by interactions of the protein kinase C and protein kinase A second messenger systems.
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PMID:In vitro modulation of porcine Leydig cell steroidogenesis by phorbol-12-myristate-13-acetate and 1,2-dioctanoylglycerol. 230 1

An enriched rat Leydig cell preparation was preincubated with [14C]arachidonic acid. Stimulation of the cells with arginine vasopressin (AVP) (1 microM) for 2 min caused a significant increase in labelled phosphatidic acid and a significant fall in radioactivity in phosphatidylinositol and phosphatidylinositol 4-monophosphate + phosphatidylinositol 4,5-bisphosphate. Preincubation with dibutyryl cyclic AMP had no effect on the AVP-induced phospholipid turnover. Leydig cells were preincubated with myo-[2-3]inositol for 22 h and then with 10 mM LiCl for 10 min. Exposure to AVP (1 microM) induced a rise in labelled inositol phosphates. The response was inhibited when the cells were preincubated with the phorbol ester, 4 beta-phorbol 12-myristate 13-acetate (0.16 microM) for 10 min. These results provide evidence for an AVP-induced phospholipase C stimulation in rat Leydig cells and suggest a protein kinase C-dependent feedback inhibition of the stimulation. Other agonists that might have a regulatory function in the testis were tested for possible effects on phosphoinositide metabolism. Of prostaglandin E2 (10 microns,) angiotensin II (0.1 microM), and bradykinin (0.9 microM), only the latter induced a significant increase in the labelled inositol phosphates. This suggests that Leydig cells possess a bradykinin receptor which can activate phospholipase C.
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PMID:Arginine vasopressin stimulates phosphoinositide turnover in an enriched rat Leydig cell preparation. 253 41

Luteinizing hormone is the major regulator of Leydig cell differentiation and steroidogenic function. A number of hormones produced by the Leydig cell (e.g. estrogen, angiotensin, CRF, vasopressin) and the tubular compartment (inhibin, TGF beta), can influence both acute and long-term actions of LH. Conversely, hormones produced in the Leydig cells modulate tubular function (e.g. androgen, beta-endorphin, oxytocin). The LH stimulatory event can be negatively influenced by the action of angiotensin II through the guanyl nucleotide inhibitory unit of adenylate cyclase. We have recently discovered an action of corticotrophin releasing hormone through specific high-affinity low-capacity receptors in the Leydig cells which involves a pertussis toxin insensitive guanyl nucleotide regulatory unit with interaction between signalling pathways and resulting inhibition of LH induced cAMP generation and consequently of steroidogenesis. In contrast to other tissues the CRF receptor in the Leydig cells did not couple to Gs. CRF action is exerted through direct or indirect action of protein kinase C, at the level of the catalytic subunit of adenylate cyclase. Physiological increases in endogenous LH cause positive regulation of membrane receptors and steroidogenesis, while major elevations in circulating gonadotropin can induce down-regulation of LH receptors and desensitization of steroid responses in the adult cell. Gonadotropin-induced desensitization in adult rat tests include an estrogen mediated steroidogenic lesion of the microsomal enzymes 17 alpha-hydroxylase/17,20-desmolase. For further understanding of the regulation of this key enzyme of the androgen pathway the rat P450(17) alpha cDNA was cloned and sequenced. This cDNA expressed in COS-1 cells 17 alpha-hydroxylase/17,20-desmolase activities. From the deduced amino acid sequence, two transmembrane regions were identified, a signal peptide for insertion in the ER, and a 2nd transmembrane region separated from the first by 122 amino acids. The carboxy terminal non-transmembrane region possesses 4 hydrophobic clefts, of which cleft II would contain the putative steroid binding site for both hydroxylase and lyase activities. The rat cDNA was employed to evaluate the hormonal regulation of mRNA levels in adult and fetal Leydig cells. Low dose hCG treatment caused an early increase in mRNA levels followed by a return to control values at later times, while with higher desensitizing doses the initial increase in mRNA was followed by a marked reduction in mRNA at 24 h and a small recovery at 48 h. Fetal rat Leydig cells treated with E2 showed a 70% decrease in P450 mRNA levels, and testosterone production closely followed the changes in mRNA.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:LH action in the Leydig cell: modulation by angiotensin II and corticotropin releasing hormone, and regulation of P450(17) alpha mRNA. 269 45

Gonadotropin-releasing hormone (GnRH) has specific receptor sites in rat Leydig cells and has direct effects on their steroidogenesis. The purpose of the present study was to examine whether activation of the calcium- and phospholipid-dependent protein kinase C (PK-C) is involved in GnRH effects on rat Leydig cells, as has been shown in pituitary gonadotrophs. Testosterone production of Percoll-purified Leydig cells was similarly stimulated (about 50-100%) by a GnRH agonist (buserelin, maximum effect at concentration of 10(-9) mol/l and above) and a tumor promoting phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA, maximum effect at 10(-8) mol/l), which is known to activate PK-C. In contrast, a GnRH antagonist (10(-5) mol/l) and an inactive phorbol ester, 4 alpha-phorbol-12,13-didecanoate (10(-6) mol/l), were without effect on testosterone. None of these substances had clear effects on cAMP production. The maximum steroidogenic effects of GnRH agonist and TPA were the same whether used separately or together, suggesting that they share a common mechanism of action. TPA translocated the cytosolic proportion of Leydig cell PK-C activity to a membrane-associated form almost instantaneously, within 0.5-1 min. A similar translocation, though less complete, was observed in the presence of buserelin in 1-4 min. Inclusion of a 100-fold excess of a potent GnRH antagonist completely prevented the translocation of PK-C. These results provide evidence that GnRH agonist activates PK-C also in the testis tissue, and this may be the mechanism whereby it affects Leydig cell endocrine function.
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PMID:Gonadotropin-releasing hormone agonist activates protein kinase C in rat Leydig cells. 283 41

The distribution and role of the calcium-activated, phospholipid-dependent protein kinase C (PK-C) was studied in rat testis. When testis tissue was homogenized in the presence of 2 mmol/l EDTA and EGTA, the majority (greater than 70%) of the PK-C activity was soluble, the rest was released from the particulate fraction by solubilization with 0.3% Triton X-100. Without chelating agents the soluble PK-C activity was undetectable, and only partially recovered from solubilized membranes. Preincubation of the tissue with the tumor-promoting phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA, 10(-7) mol/l) translocated PK-C to the membranes, and the majority of this activity was recovered by solubilization. Mobility of testicular soluble PK-C activity in HPLC-DEAE cellulose chromatography was similar to that of the brain enzyme. This single step purified testicular PK-C activity 140-fold. The specific activity and subcellular distribution of PK-C was similar in whole testis tissue and separated seminiferous tubules (160-210 pmol 32P X mg protein-1 X min-1 in the soluble and particulate fractions), but 2- to 3-fold higher in purified Leydig cells. However, the majority of total testicular PK-C activity appeared to be of tubular origin. Unilateral cryptorchidism for 1 week reduced PK-C of the abdominal testis by 50%, and the activity of dissected seminiferous tubules varied according to the epithelial wave. Both findings suggest that the bulk of the activity resides in the seminiferous epithelium. Involvement of PK-C in Leydig cell function was demonstrated using the TPA, which at 10(-7) mol/l inhibited basal cAMP production by 50% (P less than 0.01) but increased that of testosterone by 2- to 3-fold (P less than 0.01). On the other hand, when incubated with hCG, TPA inhibited both cAMP and testosterone production; the ED50s of hCG stimulation increased 4- to 10-fold with both parameters. It is concluded that PK-C activity is present in both the seminiferous tubules and Leydig cells, and is involved in the regulation of these testicular compartments. Its total activity and subcellular distribution are at variance according to the functional state and endocrine milieu of the testis.
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PMID:Distribution and activation of protein kinase C in the rat testis tissue. 288 17


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