scholarly journals Phospholipase C-δ1 and oxytocin receptor signalling: evidence of its role as an effector

1998 ◽  
Vol 331 (1) ◽  
pp. 283-289 ◽  
Author(s):  
Eu-Suk PARK ◽  
Jong Hyun WON ◽  
Kee Jung HAN ◽  
Pann-Ghill SUH ◽  
Sung Ho RYU ◽  
...  
Keyword(s):  
Phospholipase C ◽  
Oxytocin Receptor ◽  
Signalling Pathway ◽  
Human Myometrium ◽  
Signal Molecules ◽  
Receptor Signalling ◽  
Tightly Coupled ◽  
Oxytocin Antagonist ◽  
Stimulation Of

Although the oxytocin receptor modulates intracellular Ca2+ ion levels in myometrium, the identities of signal molecules have not been clearly clarified. Our previous studies on oxytocin receptor signalling demonstrated that 80 kDa Ghα is a signal mediator [Baek, Kwon, Lee, Kim, Muralidhar and Im (1996) Biochem. J. 315, 739–744]. To elucidate the effector in the oxytocin receptor signalling pathway, we evaluated the oxytocin-mediated activation of phospholipase C (PLC) by using solubilized membranes from human myometrium and a three-component preparation containing the oxytocin receptor–Ghα–PLC-δ1 complex. PLC-δ1 activity in the three-component preparation, as well as PLC activity in solubilized membranes, was increased by oxytocin in the presence of Ca2+ and activated Ghα (GTP-bound Ghα). Furthermore the stimulated PLC-δ1 activity resulting from activation of Ghα via the oxytocin receptor was significantly attenuated by the selective oxytocin antagonist desGly-NH2d(CH2)5[Tyr(Me)2,Thr4]ornithine vasotocin or GDP. Consistent with these observations, co-immunoprecipitation and co-immunoadsorption of PLC-δ1 in the three-component preparation by anti-Gh7α antibody resulted in the PLC-δ1 being tightly coupled to activated Ghα on stimulation of the oxytocin receptor. These results indicate that PLC-δ1 is the effector for Ghα-mediated oxytocin receptor signalling.

Oxytocin receptor couples to the 80 kDa Ghα family protein in human myometrium

1996 ◽  
Vol 315 (3) ◽  
pp. 739-744 ◽  
Author(s):  
Kwang Jin BAEK ◽  
Nyoun Soo KWON ◽  
Hee Sung LEE ◽  
Myoung Soo KIM ◽  
Pallaki MURALIDHAR ◽  
...  
Keyword(s):  
G Protein ◽  
Ternary Complex ◽  
Regulatory Protein ◽  
Oxytocin Receptor ◽  
Human Myometrium ◽  
Gtp Binding ◽  
Family Protein ◽  
Tightly Coupled ◽  
Complex Preparation ◽  
Oxytocin Antagonist

One of the primary functions of the oxytocin receptor is to modulate intracellular calcium levels in myometrium. The oxytocin receptor has been purified and cloned. Although it has been suggested that oxytocin receptor couples with a GTP-binding regulatory protein (G-protein), the identity of this G-protein remains unclear. To elucidate the mechanism of oxytocin receptor signalling, we used the oxytocin–receptor–G-protein ternary complex preparation from human myometrium, and evaluated oxytocin-mediated activation of [35S]guanosine 5´-[γ-thio]triphosphate ([35S]GTP[S]) binding and [α-32P]GTP photoaffinity labelling to a G-protein. Binding of [35S]GTP[S] and the intensity of the [α-32P]GTP photoaffinity labelled protein resulting from activation of the oxytocin receptor were significantly attenuated by the selective oxytocin antagonist, desGlyNH2d(CH2)5[Tyr(Me)2,Thr4]OVT. Furthermore, the molecular mass of the specific GTP-binding protein was ~80 kDa; homologous with the Ghα family, the new class of GTP-binding proteins first identified in rat liver that couples to the α1B-adrenoceptor. Consistent with these observations, in co-immunoprecipitation and co-immunoadsorption of the oxytocin receptor in the ternary complex preparation by anti-Gh7α antibody, the Ghα family protein tightly coupled to the oxytocin receptor. These findings demonstrate that oxytocin receptor couples with ~80 kDa Ghα in signal mediation.

scholarly journals Influence of oxytocin receptor single nucleotide sequence variants on contractility of human myometrium: an in vitro functional study

2019 ◽  
Vol 20 (1) ◽  
Author(s):  
F. Füeg ◽  
S. Santos ◽  
C. Haslinger ◽  
B. Stoiber ◽  
L. Schäffer ◽  
...  
Keyword(s):  
Mode Of Delivery ◽  
Oxytocin Receptor ◽  
Functional Study ◽  
Human Myometrium ◽  
Sequence Variants ◽  
Organ Bath ◽  
Single Nucleotide ◽  
Allele Distribution ◽  
Stimulation Of

Abstract Background Oxytocin receptor (OXTR) gene variants have been shown to affect the prevalence of preterm birth, mode of delivery and oxytocin (OXT) requirements for labor induction and augmentation. We hypothesized that this might be associated with different myometrium responses to oxytocin. Our aim was to investigate the influence of a selection of eight OXTR gene single nucleotide variants on oxytocin-induced stimulation of human myometrium contractility in vitro. Methods Human myometrium biopsies were collected during elective cesarean sections at term, if patients had given informed consent. Myometrial strips were submerged under tension in an organ bath and allowed to contract; the remaining material was stored at − 80 °C for further determination of relevant genetics and mRNA level. The area under the curve (AUC) of all contractions taking place in the absence of OXT and of those occurring upon OXT addition (for 30 min each) was measured. OXT stimulation, defined as the ratio between AUC measurements after OXT addition and those in the absence of OXT was calculated for each strip. TaqMan™ Assays were used to detect the allele distribution of the eight OXTR variants and to determine the relative amounts of OXTR-mRNA in the samples. For each variant, oxytocin stimulation of contractility was compared between samples homozygous for the reference allele (reference group) and samples with at least one variant allele (variant group) by linear regression. Results Sixty samples were included in the present study. For rs1042778, rs11706648, rs4686301, rs53576, rs237895, and rs237902, OXT stimulation was similar in the reference and in the variant groups. However, the values of OXT stimulation differed significantly between the reference and the variant groups for rs4686302 (3.1 vs. 4.1 times; p = 0.022) and rs237888 (3.2 vs. 5.5 times; p = 0.001). No significant differences between the levels of OXTR-mRNA in the various reference and corresponding variant groups were detected. Conclusions Patients with variant alleles of rs237888 and/or rs4686302 may be more sensitive to oxytocin stimulation, explaining why these sequence variants have been associated with lower cesarean section prevalence and premature birth, respectively.

scholarly journals Transcriptional Inhibition of Oxytocin Receptor Expression in Human Myometrial Cells by Melatonin Involves Protein Kinase C Signaling

2007 ◽  
Vol 92 (10) ◽  
pp. 4015-4019 ◽  
Author(s):  
James Sharkey ◽  
James Olcese
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phospholipase C ◽  
Gene Transcription ◽  
Late Pregnancy ◽  
Oxytocin Receptor ◽  
Receptor Expression ◽  
Human Myometrium ◽  
Myometrial Cells ◽  
Kinase C

Abstract Context: Our laboratory recently characterized the expression of the melatonin receptors in the human myometrium and showed that the expression of these receptors is suppressed during late pregnancy. Objective: In an effort to understand better the significance of melatonin in the human myometrium, we explored the mechanisms through which this hormone influences the expression of the oxytocin receptor in vitro. Design: The stable melatonin analog iodomelatonin was presented to cultured telomerase-immortalized myometrial cells of the human telomerase reverse transcriptase line under physiological doses and durations. Pharmacological inhibitors of melatonin binding, gene transcription, phospholipase C, and protein kinase C signaling were used to define the mechanism of melatonin action. Results: Our results reveal that melatonin significantly inhibits oxytocin receptor mRNA expression primarily via the melatonin 2 receptor. The melatonin-dependent decrease in oxytocin receptor transcripts involves suppression of gene transcription rather than enhanced rates of transcript degradation. Melatonin effects were abolished by pretreating the cells with the phospholipase C inhibitor U73122 or the protein kinase C inhibitor C1. Conclusions: Melatonin, like oxytocin, can negatively regulate oxytocin receptor transcription in human myometrial cells via modulation of protein kinase C signaling. This is consistent with the hypothesis that the reduced melatonin receptor expression during late pregnancy, which occurs at a time when oxytocin receptors are up-regulated, may be physiologically important for the subsequent timing of labor.

scholarly journals The Effect of an Oxytocin Receptor Antagonist (Retosiban, GSK221149A) on the Response of Human Myometrial Explants to Prolonged Mechanical Stretch

Endocrinology ◽  
2015 ◽  
Vol 156 (10) ◽  
pp. 3511-3516 ◽  
Author(s):  
Alexandros A. Moraitis ◽  
Yolande Cordeaux ◽  
D. Stephen Charnock-Jones ◽  
Gordon C. S. Smith
Keyword(s):  
Preterm Birth ◽  
Receptor Antagonist ◽  
Multiple Pregnancy ◽  
Mechanical Stretch ◽  
Inhibitory Effect ◽  
Oxytocin Receptor ◽  
Human Myometrium ◽  
Spontaneous Preterm Birth ◽  
Myometrial Contractility ◽  
Stimulation Of

Multiple pregnancy is a major cause of spontaneous preterm birth, which is related to uterine overdistention. The objective of this study was to determine whether an oxytocin receptor antagonist, retosiban (GSK221149A), inhibited the procontractile effect of stretch on human myometrium. Myometrial biopsies were obtained at term planned cesarean delivery (n = 12). Each biopsy specimen was dissected into 8 strips that were exposed in pairs to low or high stretch (0.6 or 2.4 g) in the presence of retosiban (1 μM) or vehicle (dimethylsulfoxide) for 24 hours. Subsequently, we analyzed the contractile responses to KCl and oxytocin in the absence of retosiban. We found that incubation under high stretch in vehicle alone increased the response of myometrial explants to both KCl (P = .007) and oxytocin (P = .01). However, there was no statistically significant effect of stretch when explants were incubated with retosiban (P = .3 and .2, respectively). Incubation with retosiban in low stretch had no statistically significant effect on the response to either KCl or oxytocin (P = .8 and >.9, respectively). Incubation with retosiban in high stretch resulted in a statistically significant reduction (median fold change, interquartile range, P) in the response to both KCl (0.74, 0.60–1.03, P = .046) and oxytocin (0.71, 0.53–0.91, P = .008). The greater the effect of stretch on explants from a given patient, the greater was the inhibitory effect of retosiban (r = −0.65, P = .02 for KCl and r= −0.73, P = .007 for oxytocin). These results suggest that retosiban prevented stretch-induced stimulation of human myometrial contractility. Retosiban treatment is a potential approach for preventing preterm birth in multiple pregnancy.

Stimulation of Ca(2+)-dependent exocytosis of the sperm acrosome by cAMP acting downstream of phospholipase A2

Reproduction ◽  
2000 ◽  
pp. 57-68 ◽  
Author(s):  
J Garde ◽  
ER Roldan
Keyword(s):  
Arachidonic Acid ◽  
Phospholipase A2 ◽  
Phospholipase C ◽  
Phosphodiesterase Inhibitors ◽  
Dibutyryl Camp ◽  
Camp Phosphodiesterase ◽  
Ram Sperm ◽  
Camp Formation ◽  
Stimulation Of

Spermatozoa undergo exocytosis in response to agonists that induce Ca2+ influx and, in turn, activation of phosphoinositidase C, phospholipase C, phospholipase A2, and cAMP formation. Since the role of cAMP downstream of Ca2+ influx is unknown, this study investigated whether cAMP modulates phospholipase C or phospholipase A2 using a ram sperm model stimulated with A23187 and Ca2+. Exposure to dibutyryl-cAMP, phosphodiesterase inhibitors or forskolin resulted in enhancement of exocytosis. However, the effect was not due to stimulation of phospholipase C or phospholipase A2: in spermatozoa prelabelled with [3H]palmitic acid or [14C]arachidonic acid, these reagents did not enhance [3H]diacylglycerol formation or [14C]arachidonic acid release. Spermatozoa were treated with the phospholipase A2 inhibitor aristolochic acid, and dibutyryl-cAMP to test whether cAMP acts downstream of phospholipase A2. Under these conditions, exocytosis did not occur in response to A23187 and Ca2+. However, inclusion of dibutyryl-cAMP and the phospholipase A2 metabolite lysophosphatidylcholine did result in exocytosis (at an extent similar to that seen when cells were treated with A23187/Ca2+ and without the inhibitor). Inclusion of lysophosphatidylcholine alone, without dibutyryl-cAMP, enhanced exocytosis to a lesser extent, demonstrating that cAMP requires a phospholipase A2 metabolite to stimulate the final stages of exocytosis. These results indicate that cAMP may act downstream of phospholipase A2, exerting a regulatory role in the exocytosis triggered by physiological agonists.

scholarly journals cAMP activates calcium signalling via phospholipase C to regulate cellulase production in the filamentous fungus Trichoderma reesei

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Yumeng Chen ◽  
Xingjia Fan ◽  
Xinqing Zhao ◽  
Yaling Shen ◽  
Xiangyang Xu ◽  
...  
Keyword(s):  
Signal Transduction ◽  
Phospholipase C ◽  
Trichoderma Reesei ◽  
Filamentous Fungus ◽  
Calcium Signalling ◽  
Signal Transduction Pathway ◽  
Cellulase Production ◽  
Signalling Pathway ◽  
Dependent Manner ◽  
Transduction Pathway

Abstract Background The filamentous fungus Trichoderma reesei is one of the best producers of cellulase and has been widely studied for the production of cellulosic ethanol and bio-based products. We previously reported that Mn2+ and N,N-dimethylformamide (DMF) can stimulate cellulase overexpression via Ca2+ bursts and calcium signalling in T. reesei under cellulase-inducing conditions. To further understand the regulatory networks involved in cellulase overexpression in T. reesei, we characterised the Mn2+/DMF-induced calcium signalling pathway involved in the stimulation of cellulase overexpression. Results We found that Mn2+/DMF stimulation significantly increased the intracellular levels of cAMP in an adenylate cyclase (ACY1)-dependent manner. Deletion of acy1 confirmed that cAMP is crucial for the Mn2+/DMF-stimulated cellulase overexpression in T. reesei. We further revealed that cAMP elevation induces a cytosolic Ca2+ burst, thereby initiating the Ca2+ signal transduction pathway in T. reesei, and that cAMP signalling causes the Ca2+ signalling pathway to regulate cellulase production in T. reesei. Furthermore, using a phospholipase C encoding gene plc-e deletion strain, we showed that the plc-e gene is vital for cellulase overexpression in response to stimulation by both Mn2+ and DMF, and that cAMP induces a Ca2+ burst through PLC-E. Conclusions The findings of this study reveal the presence of a signal transduction pathway in which Mn2+/DMF stimulation produces cAMP. Increase in the levels of cAMP activates the calcium signalling pathway via phospholipase C to regulate cellulase overexpression under cellulase-inducing conditions. These findings provide insights into the molecular mechanism of the cAMP–PLC–calcium signalling pathway underlying cellulase expression in T. reesei and highlight the potential applications of signal transduction in the regulation of gene expression in fungi.

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