Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:4.6.1.1 (adenylate cyclase)
19,190 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The present investigation was designed to study the direct role of PRL on testicular Leydig cell steroidogenesis, using the MA-10 murine Leydig tumor cell line as a model system. We have previously reported on the presence of specific PRL binding sites in those cells, and we now demonstrate the functionality of those sites and the biological responses induced by the binding of PRL. When cultured MA-10 cells were exposed for 24 h to increasing concentrations of PRL, washed, and then subjected to a 3-h human CG (hCG) stimulation test, a clear dose-dependent biphasic effect of PRL on the steroidogenic response was observed, even though PRL had no effect on MA-10 cell proliferation: at low PRL concentrations (0.1-10 ng/ml), hCG-induced steroidogenesis was stimulated (maximal stimulation by 1 ng/ml PRL being 200-250% of control); at higher concentrations, hCG-induced steroidogenesis was inhibited (60% inhibition was achieved by 1000 ng/ml PRL). When steroidogenesis was induced with various concentrations of cholera toxin, instead of hCG, no effect of the prior exposure to increasing concentrations of PRL was observed, indicating that PRL acts either at the level of the LH/hCG receptor or at some stage proximal to adenylate cyclase. Indeed, further study revealed that 24 or 72 h exposure of MA-10 cells to PRL caused a dose-dependent reduction in hCG binding. Thus, the maximal inhibition of 62% after 72 h with 500 ng/ml PRL, may explain, at least in part, the inhibitory effects of high PRL concentrations on hCG-induced progesterone secretion. Evidence demonstrating possible involvement of a pertussis toxin-(PT-)sensitive G protein in the signal transduction mechanism of PRL receptors is also presented: 1. GTP caused a dose-dependent reduction in affinity (Ka) of PRL binding by its receptors (from Ka = 1.66 +/- 0.2 x 10(9) M(-1) for control MA-10 cell membranes to Ka 3.03 +/- 0.6 x 10(8) M(-1) for membranes incubated with 8 mM GTP). 2. Prior exposure of MA-10 cells to PRL (10 pg/ml) caused a significant reduction in the ability of a 44-kDa membrane protein to undergo PT-induced [32P]ADP-ribosylation. These results demonstrate that MA-10 Leydig cells possess highly specific and biologically functional PRL receptors mediating direct and dose-dependent biphasic effects of PRL on hCG-induced progesterone secretion. These cells thus offer a suitable model to study the mechanism(s) of PRL action and signal transduction of its receptor on a physiologically relevant differentiated function.
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PMID:Prolactin and MA-10 Leydig cell steroidogenesis: biphasic effects of prolactin and signal transduction. 894 Mar 78

The testis is divided, both functionally and anatomically, into two specialised compartments, i.e. the vascular interstitial tissue and avascular seminiferous tubules. Leydig cell steroidogenesis is mainly controlled by luteinizing hormone (LH) secreted from the anterior pituitary. It is well known that LH is able to generate second messengers by adenylate cyclase and the cycle of inositolphospholipids. Also Ca2+ ions are taken into account as an important modulators of Leydig cell steroidogenesis. The aim of this study was to show the effect of macrophage-conditioned medium on the basal and LH stimulated testosterone secretion by mouse Leydig cells in vitro. A source of Leydig cells was the testis of mature, Swiss strain mice. Leydig cells were isolated and cultured for 48 hrs. Testosterone levels were measured radioimmunologically. Intracellular free Ca2+ ions were analysed by computer automatic system 'MAGICAL'. Results indicated that not only LH but also macrophage-conditioned medium were able to modulate testosterone secretion by Leydig cells and increase intracellular calcium concentration. Therefore, it seems possible that the action of proteins secreted from macrophages is mediated through Ca2+ ions which are involved in another signal transduction pathway leading to stimulation of testosterone biosynthesis by Leydig cells in vitro.
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PMID:[Immuno-endocrine interactions in the testis: possible involvement of Ca2+ in signal transduction pathway in Leydig cells]. 969 59

Some environmental chemicals exhibit estrogenic or antiandrogenic activity. Some of these, such as bisphenol A (bis A) and octylphenols, are used in large amounts in many applications. We have analyzed the effects of bis A and octylphenols on steroidogenesis in Leydig cells by measuring the LH receptor-mediated cAMP and progesterone (P) production in cultured mouse Leydig tumor cells (mLTC-1 cells). After preincubation of mLTC-1 cells for 48 h in the presence of bis A or one of the octylphenols in micromolar concentration, the hCG-stimulated cAMP and P formation in these cells was inhibited. Bis A or octylphenols could neither inhibit cAMP nor P formation stimulated by forskolin (Fk) or cholera toxin (CT) nor steroidogenesis stimulated by 8-Br-cAMP. The preincubation of mLTC-1 cells with estradiol or diethylstilbesterol (DES) at the concentration of 10(-8) mol/liter had no inhibitory effect on cAMP formation stimulated by hCG or Fk but P production was inhibited. Similarly, both estrogens inhibited P production stimulated by 8-Br-cAMP. Bis A or octylphenols had no effect on 125I-hCG binding to Leydig cell LH-receptors. Thus, these environmental chemicals appear to inhibit cAMP formation and steroidogenesis in mLTC-1 Leydig tumor cells by preventing the coupling between LH receptor and the adenylate cyclase. Since, estradiol did not inhibit hCG-stimulated cAMP production, the effects of bis A and octylphenols may not be estrogen related. This emphasizes the complexity of endocrine disruption: chemicals show multiple endocrine activities that may disturb several organs in distinct ways.
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PMID:Inhibition of hCG-stimulated steroidogenesis in cultured mouse Leydig tumor cells by bisphenol A and octylphenols. 1037

The involvement of adenylate cyclase-cyclic adenosine monophosphate (AC-cAMP) in gonadotropin-stimulated testicular steroidogenesis is well known. Little is known about the role of guanylate cyclase-cyclic guanosine monophosphate (GC-cGMP) or early chloride conductance stimulated by gonadotropins in steroidogenesis. Human chorionic gonadotropin (hCG) 1 IU/L caused significant androgen secretion without a discernible effect on cAMP production. Despite negligible intracellular cAMP, the protein kinase A inhibitor H89 blocked basal and hCG-stimulated steroidogenesis. The GC inhibitors methylene blue (MB) and LY83583 decreased androgen secretion, but hCG did not stimulate cGMP production and there was not a steroidogenic response to exogenous cGMP. A chloride-channel inhibitor, diphenylamine-2-carboxylate (DPC), at concentrations up to 0.6 mmol/L stimulated basal steroid secretion and hCG 10 IU/L stimulated cAMP production, but higher concentrations had an inhibitory effect. Substitution of chloride by gluconate enhanced basal steroid secretion, but nitrate completely abolished the effect of 1 IU/L hCG on androgen secretion, which could be partially overcome by increasing the gonadotropin concentration. In conclusion, chloride, perhaps by activating AC-cAMP, mediates the steroidogenic action of gonadotropins in mouse Leydig tumor cells (MLTC-1). Inorganic nitrate probably inhibited steroidogenesis via conversion to nitric oxide (NO) without involving the GC-cGMP pathway. Nevertheless, the results obtained with GC inhibitors suggest a role for the GC-cGMP pathway in Leydig cell steroidogenesis.
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PMID:Role of chloride and inhibitory action of inorganic nitrate on gonadotropin-stimulated steroidogenesis in mouse Leydig tumor cells. 1038 Nov 42

Secreted peptide hormones and components of the steroidogenic machinery are molecules that are expressed usually in high amounts and in a time- and cell-specific fashion within the cells that give rise to the bovine corpus luteum. They thus serve as useful markers for the events occurring within the nuclei of these cells that result in differentiation and the expression of the specific luteal phenotype. We have studied the bovine genes of three such luteal products: oxytocin, the new relaxin-like factor (RLF), and the steroidogenic acute regulatory protein (StAR). The oxytocin gene is expressed in the granulosal cells of the preovulatory follicle and in the large luteal cells of the immediately resulting early corpus luteum. The RLF gene is a major thecal cell product in antral and atretic follicles. It is also transcribed in luteal cells, but only in the mid- to late ovarian cycle and in pregnancy, following a temporal pattern of expression very similar to that of relaxin in pigs. The StAR gene appears to be upregulated only in the mid- to late ovarian cycle, several days after the increase in steroidogenic enzymes associated with luteinization and progesterone production. All three genes make use of the transcription factor SF-1 (Ad4BP) and, although they all respond to LH activation of adenylate cyclase, none utilize CRE-linked systems. Specific transcriptional activation must involve other factors to encode the information for the widely diverse temporal and cellular patterns of gene expression for these three genes.
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PMID:Luteal peptides and their genes as important markers of ovarian differentiation. 1069 56

Previous results from our group have indicated that arachidonic acid decrease cAMP production through a modification of heterotrimeric G proteins. In the present study, we have characterized the high affinity GTPase activity present in Leydig cell membranes and its regulation by fatty acids. The high-affinity GTPase activity, measured as [gamma32P] GTP hydrolysis rate, was both time and protein concentration dependent. Arachidonic acid elicited a dose-dependent inhibition of enzyme activity with an IC50 = 26.7+/-1.1 microM. The existence of only two double bonds in linoleic acid is reflected by a decrease in its inhibitory activity (IC50 = 34+/-2.3 microM). Saturated fatty acids showed no effect at this level. The kinetic analysis as interpreted by Lineweaver-Burk plots, indicated that 50 microM arachidonic acid had no effect on the apparent affinity for GTP, but resulted in a 40% decreases in the maximal velocity of the reaction. Arachidonic acid modulation of GTPase activity was not attenuated by blocking eicosanoid metabolism with inhibitors of 5'-lipoxygenase, cyclooxygenase, or epoxygenase P-450. The addition of arachidonic acid to pertussis toxin-treated membranes had no effect on the enzyme activity, indicating that arachidonic acid does not modify the GTPase activity present in Galphas protein. However, ADP-ribosylation with cholera toxin followed by arachidonic acid treatment led to a further 40% inhibition when compared with cholera toxin treatment alone. These results allowed us to postulate that arachidonic acid inhibits the GTPase activity of Gi protein family. To further analyze the mechanism of arachidonic acid inhibition of GTPase activity, the effect of arachidonic acid on the [35S]GTPgammaS binding was studied. No effect of this fatty acid on GTP binding was found. Combining our previous results with those found here, we can conclude that arachidonic acid maintains Gi proteins in their active state, which in turn inhibit adenylate cyclase and results in decrease cAMP levels.
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PMID:Modulation of guanosine triphosphatase activity of G proteins by arachidonic acid in rat Leydig cell membranes. 1069 85

Aging in Brown Norway rats is associated with reduced Leydig cell T production. To address the mechanism by which aging Leydig cells become steroidogenically hypofunctional, Leydig cells from young and old rat testes were isolated and cultured long-term with LH. Leydig cells isolated from young rats that had received LH-suppressive T implants served as positive controls. The ability of young control Leydig cells to produce T at high levels was sustained over a 3-d culture period. T production by cells from young LH-suppressed rats increased over this period, almost to control levels. In contrast, culture of the steroidogenically hypofunctional old Leydig cells with LH failed to increase their T production, suggesting that LH stimulation, by itself, is unable to reverse the steroidogenic deficits of old Leydig cells. Reduced numbers of LH binding sites characterized Leydig cells from old rats and LH-suppressed young rats. However, whereas Leydig cells from young LH-suppressed rats produced cAMP at the high levels of young control cells, the old cells produced far less cAMP, suggesting that old Leydig cells have defects in the LH-cAMP signaling cascade. When stimulated with forskolin, old cells produced the same amount of cAMP as young control and young LH-suppressed cells, suggesting that adenylate cyclase is maintained in the old cells. Taken together, these results suggest that inefficient signal transduction may explain the reduced steroidogenesis that characterizes old Leydig cells.
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PMID:Age-related decreases in Leydig cell testosterone production are not restored by exposure to LH in vitro. 1195 44

Although several reports indicate effects of histamine (HA) on female reproductive functions, scant literature exists to suggest a physiological role of HA in the male gonad. In the present study, we report a dual concentration-dependent effect of HA on steroidogenesis in MA-10 murine Leydig cells and purified rat Leydig cells. Although 1 nM HA can stimulate steroid production and significantly increase the response to LH/hCG in these cells, 10 microM HA exerts an inhibitory effect. We also provide confirming evidence for the existence of functional HRH1 and HRH2 receptors in both experimental models. The use of HRH1 and HRH2 selective agonists and antagonists led us to suggest that HRH2 activation would be largely responsible for stimulation of steroidogenesis, while HRH1 activation is required for inhibition of steroid synthesis. Our results regarding signal transduction pathways associated with these receptors indicate the coupling of HRH2 to the adenylate cyclase system through direct interaction with a Gs protein. Moreover, we show HRH1 activation mediates increases in inositol phosphate production, possibly due to coupling of this receptor to Gq protein and phospholipase C activation. The data compiled in this report clearly indicate that HA can modulate Leydig cell steroidogenesis in the testis and suggest a possible new physiological site of action for HA. Given that many drugs binding to HRH1, HRH2, or both, are widely prescribed for the treatment of diverse HA-related pathologies, it seems necessary to increase the knowledge regarding histaminergic regulation of testicular functions, to avoid possible unexpected side effects of such substances in the testis.
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PMID:Dual role of histamine in modulation of Leydig cell steroidogenesis via HRH1 and HRH2 receptor subtypes. 1591 47

This study aimed to improve, using the zebrafish model, our understanding of the distinct roles of pituitary gonadotropins FSH and LH in regulating testis functions in teleost fish. We report, for the first time in a vertebrate species, that zebrafish Leydig cells as well as Sertoli cells express the mRNAs for both gonadotropin receptors (fshr and lhcgr). Although Leydig cell fshr expression has been reported in other piscine species and may be a common feature of teleost fish, Sertoli cell lhcgr expression has not been reported previously and might be related to the undifferentiated gonochoristic mode of gonadal sex differentiation in zebrafish. Both recombinant zebrafish (rzf) gonadotropins (i.e. rzfLH and rzfFSH) stimulated androgen release in vitro and in vivo, with rzfFSH being significantly more potent than rzfLH. Forskolin-induced adenylate cyclase activation mimicked, whereas the protein kinase A inhibitor H-89 significantly reduced, the gonadotropin-stimulated androgen release. Therefore, we conclude that both FSH receptor and LH/choriogonadotropin receptor signaling are predominantly mediated through the cAMP/protein kinase A pathway to promote steroid production. Despite this similarity, other downstream mechanisms seem to differ. For example, rzfFSH up-regulated the testicular mRNA levels of a number of steroidogenesis-related genes both in vitro and in vivo, whereas rzfLH or human chorionic gonadotropin did not. Although not fully understood at present, these differences could explain the capacity of FSH to support both steroidogenesis and spermatogenesis on a long-term basis, whereas LH-stimulated steroidogenesis might be a more acute process, possibly restricted to periods during which peak steroid levels are required.
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PMID:Studies in zebrafish reveal unusual cellular expression patterns of gonadotropin receptor messenger ribonucleic acids in the testis and unexpected functional differentiation of the gonadotropins. 2030 33

The relaxin-like factor (RLF) also named insulin-like 3 (INSL3) consists of two polypeptide chains linked by two interchain and one intrachain disulfide bond. RLF binds to its receptor (LGR8 also named RXFP2) through the B chain and initiates transmembrane communication by activating the adenylate cyclase through the N-terminal region of both chains. Cystine A11-B10 occupies a unique position on the molecular surface just outside the binding region and between the two signaling ports. We have synthesized an RLF analogue in which the disulfide A11-B10 was replaced by a peptide bond and found that cAMP production ceased while receptor binding was not affected. In contrast, replacing the disulfide A24-B22 by a peptide bond reduced potency proportional to the binding affinity and lowered efficacy to 65%, while replacing disulfide A10-A15 by a peptide bond reduced binding affinity to 32% and lowered potency to 7% but maintained 100% efficacy. The exceptional properties of the derivative bearing an A11-B10 isopeptide cross-link suggests that the disulfide has a special role in signal transduction. We propose that disulfide A11-B10 serves as an insulator between the two ports, whereas the amide functionality disturbs the signal transmission complex likely due to changes in polarity. The clear separation between receptor binding and signal activation sites within this small protein permits one to study how the relaxin-like factor initiates the signal on the receptor that induces intracellular cAMP production.
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PMID:Replacement of disulfides by amide bonds in the relaxin-like factor (RLF/INSL3) reveals a role for the A11-B10 link in transmembrane signaling. 2257 50


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