scholarly journals Signalling through the leukotriene B4 receptor involves both αi and α16, but not αq or α11 G-protein subunits

1998 ◽  
Vol 335 (1) ◽  
pp. 15-18 ◽  
Author(s):  
Rémi GAUDREAU ◽  
Christian Le GOUILL ◽  
Salim MÉTAOUI ◽  
Stéphane LEMIRE ◽  
Jana STANKOVÀ ◽  
...  
Keyword(s):  
Phospholipase C ◽  
G Protein ◽  
Inositol Phosphates ◽  
Leukotriene B4 ◽  
The Other ◽  
Protein Subunits ◽  
Other Hand ◽  
Leukotriene B4 Receptor

COS-7 cells transfected with the leukotriene (LT) B4 receptor (BLTR) cDNA were unable to produce LTB4-induced inositol phosphates (IPs) in spite of the presence of endogenous Gαi, Gαq and Gα11 proteins. Co-transfection of BLTR with Gα16, however, resulted in high levels of IP production, which were 17-, 10- and 6-fold higher than with co-transfected Gα11, Gαq and Gα14, respectively. Co-transfection of BLTR with phospholipase C (PLC) β2, on the other hand, resulted in efficient IP production and co-transfection of BLTR with both Gα16 and PLCβ2 resulted in a greater than additive response.

Dexamethasone increases G alpha q-11 expression and hormone-stimulated phospholipase C activity in UMR-106-01 cells.

1997 ◽  
Vol 273 (3) ◽  
pp. E528 ◽  
Author(s):  
J Mitchell ◽  
A Bansal
Keyword(s):  
Adenylyl Cyclase ◽  
Phospholipase C ◽  
G Protein ◽  
G Protein Coupled Receptors ◽  
Beta Subunits ◽  
Adenylyl Cyclase Activity ◽  
Protein Subunits ◽  
G Alpha Q ◽  
Umr 106 ◽  
G Protein Coupled

Glucocorticoids regulate responsiveness of many cells to hormones that bind to G protein-coupled receptors. We examined the effect of glucocorticoids on parathyroid hormone (PTH) activation of two G protein-activated signal transduction pathways, phospholipase C (PLC) and adenylyl cyclase, in osteosarcoma UMR-106-01 cells. Dexamethasone (100 nM) increased PTH-stimulated and NaF-stimulated PLC activity by > 100% over 4 days (223 +/- 8 and 293 +/- 8.2% of control after 4 days for PTH and NaF-stimulated activity, respectively). The increase in PTH-stimulated adenylyl cyclase response in the same cells was more modest (162 +/- 5.4 and 171 +/- 6.8% of control after 4 days for PTH and NaF-stimulated activity, respectively). PTH activation of PLC was blocked by antiserums to G alpha q-11 and activation of adenylyl cyclase by G alpha s antiserums. Quantification of these G protein subunits in control and dexamethasone-treated cells showed a 78% increase in G alpha q-11 (from 18.1 +/- 1.2 to 32.2 +/- 1.5 pmol/mg), whereas G alpha s was increased only 34% (from 6.2 +/- 0.5 to 8.2 +/- 0.3 pmol/mg) and G beta-subunits were increased 40% (from 54 +/- 2.3 to 75.2 +/- 3.8 pmol/mg). These results suggest that glucocorticoids are more potent regulators of PLC activity than adenylyl cyclase activity in UMR cells, and this is mediated, at least in part, by differential increases in G alpha q-11 proteins.

scholarly journals Role of Dynamic Interactions in Effective Signal Transfer for Gβ Stimulation of Phospholipase C-β2

2002 ◽  
Vol 277 (51) ◽  
pp. 49707-49715 ◽  
Author(s):  
Elizabeth Buck ◽  
Peter Schatz ◽  
Suzanne Scarlata ◽  
Ravi Iyengar
Keyword(s):  
Phospholipase C ◽  
G Protein ◽  
Binding Affinity ◽  
Protein Interactions ◽  
Signal Transfer ◽  
Dynamic Interactions ◽  
Protein Subunits ◽  
Functional Studies ◽  
Effective Signal ◽  
Stimulation Of

Heterotrimeric G protein subunits regulate their effectors by protein-protein interactions. The regions involved in these direct interactions have either signal transfer or general binding functions (Buck, E., Li, J., Chen, Y., Weng, G., Scarlata, S., and Iyengar, R. (1999) Science 283, 1332–1335). Although key determinants of signal transfer regions for G protein subunits have been identified, the mechanisms of signal transfer are not fully understood. We have used a combinatorial peptide approach to analyze one Gβ region, Gβ86–105, involved in signal transfer to the effector phospholipase C (PLC)-β2 to gain a more mechanistic understanding of Gβ/PLC-β2 signaling. Binding and functional studies with the combinatorial peptides on interaction with and stimulation/inhibition of phospholipase Cβ2 indicate that binding affinity can be resolved from EC50for functional effects, such that peptides that have wild type binding affinities have 15- to 20-fold lower EC50values. Although more potent, these peptides display a much lower extent of maximal stimulation. These peptides synergize with Gβγ or peptides encoding the second Gβ42–54 signal transfer region in maximally stimulating phospholipase C-β2. Other combinatorial peptides from the Gβ86–105 region that bind to PLC-β2 by themselves submaximally stimulate and extensively inhibit Gβγ stimulation of PLC-β2. The intrinsic stimulation function can be attributed to Arg-96 and Ser-97, the synergy function to Trp-99, and the binding affinity to Thr-87, Val-90, Pro-94, Arg-96, Ser-97, and Val-100. These results indicate that, even within signal transfer regions, residues involved in binding can be resolved from those involved in signal transfer and that signal transfer is likely to be achieved through dynamic rather than steady-state interactions.

scholarly journals G-proteins are not directly involved in the CD3-antigen-mediated production of inositol phosphates in HPB-ALL T-leukaemia cells expressing phospholipase C isoforms γ1 and β3

1993 ◽  
Vol 289 (2) ◽  
pp. 387-394 ◽  
Author(s):  
M Biffen ◽  
M Shiroo ◽  
D R Alexander
Keyword(s):  
Phospholipase C ◽  
Tyrosine Phosphorylation ◽  
G Protein ◽  
G Proteins ◽  
Inositol Phosphates ◽  
Inositol Phosphate ◽  
Cross Linking ◽  
Protein Tyrosine Phosphorylation ◽  
Protein Tyrosine ◽  
Inositol Phosphate Production

The possible involvement of G-proteins in T cell antigen-receptor complex (TCR)-mediated inositol phosphate production was investigated in HPB-ALL T-cells, which were found to express the phospholipase C gamma 1 and beta 3 isoforms. Cross-linking the CD3 antigen on streptolysin-O-permeabilized cells stimulated a dose-dependent increase in inositol phosphate production, as did addition of guanosine 5′-[gamma-thio]triphosphate (GTP[S]) or vanadate, a phosphotyrosine phosphatase inhibitor. It was possible, therefore, that the CD3-antigen-mediated production of inositol phosphates was either via a G-protein-dependent mechanism or by stimulation of protein tyrosine phosphorylation. The CD3-induced inositol phosphate production was potentiated by addition of vanadate, but not by addition of GTP[S]. Guanosine 5′-[beta-thio]diphosphate (GDP[S]) inhibited the rise in inositol phosphates induced by GTP[S], vanadate or cross-linking the CD3 antigen. The increase in protein tyrosine phosphorylation stimulated by vanadate or the OKT3 monoclonal antibody was not observed in the presence of GDP[S], showing that in permeabilized HPB-ALL cells, GDP[S] inhibits the actions of tyrosine kinases as well as G-protein function. Addition of either ADP[S] or phenylarsine oxide inhibited CD3- and vanadate-mediated increases in both tyrosine phosphorylation and inositol phosphate production, but did not inhibit GTP[S]-stimulated inositol phosphate production. On the other hand, pretreatment of cells with phorbol 12,13-dibutyrate inhibited subsequent GTP[S]-stimulated inositol phosphate production but did not inhibit significantly inositol phosphate production stimulated by either OKT3 F(ab')2 fragments or vanadate. Our results are consistent with the CD3 antigen stimulating inositol phosphate production by increasing the level of protein tyrosine phosphorylation, but not by activating a G-protein.

Ligand-induced formation of the leukotriene B4 receptor-G protein complex of human polymorphonuclear leukocytes

1992 ◽  
Vol 48 (4) ◽  
pp. 367-372 ◽  
Author(s):  
Jeffrey W. Sherman ◽  
Michelle A. Mendelson ◽  
John M. Boggs ◽  
Catherine H. Koo ◽  
Edward J. Goetzl
Keyword(s):  
G Protein ◽  
Protein Complex ◽  
Polymorphonuclear Leukocytes ◽  
Leukotriene B4 ◽  
Human Polymorphonuclear Leukocytes ◽  
Leukotriene B4 Receptor

Detection of phospholipase C-β 2 activation by G-protein subunits

1998 ◽  
Author(s):  
Suzanne Scarlata ◽  
Loren Runnels ◽  
Mario Rebecchi
Keyword(s):  
Phospholipase C ◽  
G Protein ◽  
Protein Subunits

Abstract MP122: Effects of Cardiac Beta3-Adrenoceptors on the Inhibition of G Protein-coupled Receptor Kinase 2 via S-nitrosylation

2020 ◽  
Vol 127 (Suppl_1) ◽  
Author(s):  
Gizem Kayki Mutlu ◽  
Kimberly Ferrero ◽  
Rajika Roy ◽  
Anna Maria Lucchese ◽  
Erhe Gao ◽  
...  
Keyword(s):  
Cardiac Function ◽  
Infarct Size ◽  
G Protein ◽  
Contractile Function ◽  
Ischemia Reperfusion ◽  
The Other ◽  
Receptor Kinase ◽  
G Protein Coupled Receptor ◽  
Other Hand ◽  
G Protein Coupled

G Protein-Coupled Receptor Kinase 2 (GRK2) is a culprit in the loss of cardiac contractile function in heart failure due to β-Adrenoceptor (AR) desensitization after its upregulation. Indeed, its inhibition has been demonstrated to improve cardiac function and increased GRK2 in the heart leads to larger injury after an ischemic insult. Nitric oxide (NO) via S-nitrosothiol (SNO) at residue Cys340 is a reported endogenous inhibitor of GRK2 activity. ß3ARs, on the other hand, are known to be resistant to desensitization by GRK2 and they are upregulated in cardiac pathologies. Activation of ß3ARs can be cardioprotective via NO signalling. Thus, in the present study, we aimed to investigate the interaction between ß3ARs and GRK2 through NO signaling to determine if ß3AR cardioprotection can occur via NO-mediated GRK2 inhibition. We used wild type C57BL/6 mice (WT), global β3AR knockout (KO) mice and GRK2-C340S knockin mice, which harbor a point mutation that changes Cys340 with a serine, meaning all endogenous GRK2 globally cannot be inhibited via NO-mediated S-nitrosylation. We exposed WT, GRK2-C340s KI and β3AR KO mice to ischemia/reperfusion (I/R) injury (40 min ischemia followed by 24 hrs reperfusion). We found that WT mice had significantly diminished cardiac function evaluated by echocardiography and Millar Catheterization, and this was rescued by treating these mice with CL316,243 (a selective β3AR agonist) at the time of reperfusion. On the other hand, GRK2-C340S KI mice did worse after I/R injury compared to WT mice and CL316,243 did not rescue this dysfunction as it did in WT mice. As expected, β3AR KO mice had worsened cardiac function compared to WT mice and CL316,243 had no functional benefit. Infarct size measurements after I/R revealed that β3AR KO mice had larger infarcts than WT mice supporting β3ARs as being protective. Indeed, CL316,243 induced robust cardioprotection in WT mice, reducing infarct size. GRK2-C340S mice had larger infarcts than WT mice and CL316,243 failed to offer any cardioprotection. Thus, β3AR-mediated cardioprotection clearly involves inhibition of GRK2 as part of its therapeutic mechanism and this appears to involve NO-mediated S-nitrosylation.

scholarly journals Novel Computational Methodologies for Structural Modeling of Spacious Ligand Binding Sites of G-Protein-Coupled Receptors: Development and Application to Human Leukotriene B4 Receptor

2012 ◽  
Vol 2012 ◽  
pp. 1-11 ◽  
Author(s):  
Yoko Ishino ◽  
Takanori Harada
Keyword(s):  
G Protein ◽  
Time Scale ◽  
Thermal Fluctuations ◽  
Leukotriene B4 ◽  
G Protein Coupled Receptors ◽  
Dynamics Simulation ◽  
Ligand Docking ◽  
Leukotriene B4 Receptor ◽  
G Protein Coupled ◽  
Gpcr Protein

This paper describes a novel method to predict the activated structures of G-protein-coupled receptors (GPCRs) with high accuracy, while aiming for the use of the predicted 3D structures inin silicovirtual screening in the future. We propose a new method for modeling GPCR thermal fluctuations, where conformation changes of the proteins are modeled by combining fluctuations on multiple time scales. The core idea of the method is that a molecular dynamics simulation is used to calculate average 3D coordinates of all atoms of a GPCR protein against heat fluctuation on the picosecond or nanosecond time scale, and then evolutionary computation including receptor-ligand docking simulations functions to determine the rotation angle of each helix of a GPCR protein as a movement on a longer time scale. The method was validated using human leukotriene B4 receptor BLT1 as a sample GPCR. Our study demonstrated that the proposed method was able to derive the appropriate 3D structure of the active-state GPCR which docks with its agonists.

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