scholarly journals Inhibition of RhoA Translocation and Calcium Sensitization by In Vivo ADP-Ribosylation with the Chimeric Toxin DC3B

1997 ◽  
Vol 8 (12) ◽  
pp. 2437-2447 ◽  
Author(s):  
Hideyoshi Fujihara ◽  
Lori A. Walker ◽  
Ming Cui Gong ◽  
Emmanuel Lemichez ◽  
Patrice Boquet ◽  
...  
Keyword(s):  
Membrane Fraction ◽  
Guanosine Triphosphate ◽  
Calcium Sensitization ◽  
Adp Ribosylation ◽  
Kinase C ◽  
Rabbit Portal Vein ◽  
Intact Rabbit ◽  
Sensitizing Effect ◽  
Tonic Phase

Pretreatment of intact rabbit portal vein smooth muscle with the chimeric toxin DC3B (10−6 M, 48 h; Aullo et al., 1993 ; Boquet et al. 1995 ) ADP-ribosylated endogenous RhoA, including cytosolic RhoA complexed with rhoGDI, and inhibited the tonic phase of phenylephrine-induced contraction and the Ca2+-sensitization of force by phenylephrine, endothelin and guanosine triphosphate (GTP)γS, but did not inhibit Ca2+-sensitization by phorbol dibutyrate. DC3B also inhibited GTPγS-induced translocation of cytosolic RhoA ( Gonget al., 1997a ) to the membrane fraction. In DC3B-treated muscles the small fraction of membrane-associated RhoA could be immunoprecipitated, even after exposure to GTPγS, which prevents immunoprecipitation of non-ADP–ribosylated RhoA. Dissociation of cytosolic RhoA–rhoGDI complexes with SDS restored the immunoprecipitability and ADP ribosylatability of RhoA, indicating that both the ADP-ribosylation site (Asn 41) and RhoA insert loop ( Weiet al., 1997 ) are masked by rhoGDI and that the long axes of the two proteins are in parallel in the heterodimer. We conclude that RhoA plays a significant role in G-protein-, but not protein kinase C-mediated, Ca2+ sensitization and that ADP ribosylation inhibits in vivo the Ca2+-sensitizing effect of RhoA by interfering with its binding to a membrane-associated effector.

scholarly journals Regulation of NAD+ glycohydrolase activity by NAD+-dependent auto-ADP-ribosylation

1996 ◽  
Vol 318 (3) ◽  
pp. 903-908 ◽  
Author(s):  
Myung-Kwan HAN ◽  
Ji-Young LEE ◽  
Yee-Sook CHO ◽  
Young M SONG ◽  
Nyeon-Hyoung AN ◽  
...  
Keyword(s):  
Enzyme Activity ◽  
Physiological Role ◽  
Enzymic Activity ◽  
Adp Ribosylation ◽  
Nad Glycohydrolase ◽  
Intact Rabbit ◽  
Hydrolysis Of

NAD+ glycohydrolase (NADase; EC 3.2.2.5) is an enzyme that catalyses hydrolysis of NAD+ to produce ADP-ribose and nicotinamide. Its physiological role and the regulation of its enzymic activity have not been fully elucidated. In the present study, the mechanism of self-inactivation of NADase by its substrate, NAD+, was investigated by using intact rabbit erythrocytes and purified NADase. Our results suggest that inactivation of NADase was due an auto-ADP-ribosylation reaction. ADP-ribosylated NADase of rabbit erythrocytes was deADP-ribosylated when incubated without NAD+, and thus enzyme activity was simultaneously restored. These findings suggest that reversible auto-ADP-ribosylation of NADase might regulate the enzyme's activity in vivo.

Chronic treatment by the calcium channel blocker ± PN 200-110 in the rat counteracts the stimulations of pituitary weight, prolactin release and pituitary C-kinase activity induced by a chronic estradiol treatment, in vivo

1990 ◽  
Vol 122 (3) ◽  
pp. 403-408
Author(s):  
Ph. Touraine ◽  
P. Birman ◽  
F. Bai-Grenier ◽  
C. Dubray ◽  
F. Peillon ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Calcium Channel ◽  
Calcium Channel Blocker ◽  
Channel Blocker ◽  
Plasma Prolactin ◽  
Estradiol Treatment ◽  
Kinase C ◽  
Protein Kinase C Activity

Abstract In order to investigate whether a calcium channel blocker could modulate the protein kinase C activity in normal and estradiol pretreated rat pituitary, female Wistar rats were treated or not (controls) with ± PN 200-110 (3 mg · kg−1 · day−1, sc) for 8 days or with estradiol cervical implants for 8 or 15 days, alone or in combination with PN 200-110 the last 8 days. Estradiol treatment induced a significant increase in plasma prolactin levels and pituitary weight. PN 200-110 administered to normal rats did not modify these parameters, whereas it reduced the effects of the 15 days estradiol treatment on prolactin levels (53.1 ± 4.9 vs 95.0 ±9.1 μg/l, p<0.0001) and pituitary weight (19.9 ± 0.4 vs 23.0 ± 0.6 mg, p <0.001), to values statistically comparable to those measured after 8 days of estradiol treatment. PN 200-110 alone did not induce any change in protein kinase C activity as compared with controls. In contrast, PN 200-110 treatment significantly counteracted the large increase in soluble activity and the decrease in the particulate one induced by estradiol between day 8 and day 15. We conclude that PN 200-110 opposed the stimulatory effects of chronic in vivo estradiol treatment on plasma prolactin levels and pituitary weight and that this regulation was related to a concomitant modulation of the protein kinase C activity.

scholarly journals Anti-Malignant Ascites Effect of Total Diterpenoids from Euphorbiae ebracteolatae Radix Is Attributable to Alterations of Aquaporins via Inhibiting PKC Activity in the Kidney

Molecules ◽  
2021 ◽  
Vol 26 (4) ◽  
pp. 942
Author(s):  
Yuanbin Zhang ◽  
Dongfang Liu ◽  
Fan Xue ◽  
Hongli Yu ◽  
Hao Wu ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Antitumor Activity ◽  
Membrane Fraction ◽  
Collecting Duct ◽  
Human Kidney ◽  
Underlying Mechanism ◽  
Malignant Ascites ◽  
Kinase C ◽  
Collecting Duct Cells ◽  
Medullary Collecting Duct

This study evaluated the anti-ascites effect of total diterpenoids extracted from Euphorbiae ebracteolatae Radix (TDEE) on malignant ascitic mice and elucidated its underlying mechanism. TDEE was extracted by dichloromethane and subjected to column chromatography. The purity of six diterpenoids isolated from TDEE was determined to be 77.18% by HPLC. TDEE (3 and 0.6 g raw herbs/kg, p.o.) reduced ascites and increased urine output. Meanwhile, analysis of tumor cell viability, cycle and apoptosis indicated that TDEE had no antitumor activity. In addition, the expression levels of aquaporins (AQPs) and the membrane translocation levels of protein kinase C (PKC) α and PKCβ in kidney and cells were measured. TDEE reduced the levels of AQP1–4, and inhibited PKCβ expression in membrane fraction. Four main diterpenoids, except compound 2, reduced AQP1 level in human kidney-2 cells. Compounds 4 and 5 inhibited AQP2–4 expression in murine inner medullary collecting duct cells. The diterpenoid-induced inhibition of AQP1–4 expression was blocked by phorbol-12-myristate-13-acetate (PMA; agonist of PKC). The diterpenoids from TDEE are the main anti-ascites components. The anti-ascites effect of diterpenoids may be associated with alterations in AQPs in the kidneys to promote diuresis. The inhibition of AQP1–4 expression by TDEE is related to the inhibition of PKCβ activation.

scholarly journals ADP-ribosylation of integrin α7 modulates the binding of integrin α7β1 to laminin

2004 ◽  
Vol 385 (1) ◽  
pp. 309-317 ◽  
Author(s):  
Zhefeng ZHAO ◽  
Joanna GRUSZCZYNSKA-BIEGALA ◽  
Anna ZOLKIEWSKA
Keyword(s):  
C2c12 Myotubes ◽  
Post Translational Modification ◽  
Integrin Β1 ◽  
Adp Ribosylation ◽  
In Vitro Binding ◽  
Vitro Binding ◽  
C2c12 Skeletal Muscle Cells ◽  
Adp Ribosyltransferase

The extracellular domain of integrin α7 is ADP-ribosylated by an arginine-specific ecto-ADP-ribosyltransferase after adding exogenous NAD+ to intact C2C12 skeletal muscle cells. The effect of ADP-ribosylation on the structure or function of integrin α7β1 has not been explored. In the present study, we show that ADP-ribosylation of integrin α7 takes place exclusively in differentiated myotubes and that this post-translational modification modulates the affinity of α7β1 dimer for its ligand, laminin. ADP-ribosylation in the 37-kDa ‘stalk’ region of α7 that takes place at micromolar NAD+ concentrations increases the binding of the α7β1 dimer to laminin. Increased in vitro binding of integrin α7β1 to laminin after ADP-ribosylation of the 37-kDa fragment of α7 requires the presence of Mn2+ and it is not observed in the presence of Mg2+. In contrast, ADP-ribosylation of the 63-kDa N-terminal region comprising the ligand-binding site of α7 that occurs at approx. 100 μM NAD+ inhibits the binding of integrin α7β1 to laminin. Furthermore, incubation of C2C12 myotubes with NAD+ increases the expression of an epitope on integrin β1 subunit recognized by monoclonal antibody 9EG7. We discuss our results based on the current models of integrin activation. We also hypothesize that ADP-ribosylation may represent a mechanism of regulation of integrin α7β1 function in myofibres in vivo when the continuity of the membrane is compromised and NAD+ is available as a substrate for ecto-ADP-ribosylation.

Effect of phorbol myristate acetate on secretion of parathyroid hormone

1988 ◽  
Vol 254 (1) ◽  
pp. E63-E70 ◽  
Author(s):  
J. J. Morrissey
Keyword(s):  
Protein Kinase C ◽  
Parathyroid Hormone ◽  
Protein Kinase ◽  
Membrane Fraction ◽  
Hormone Secretion ◽  
Myristate Acetate ◽  
Kinase C ◽  
Low Calcium ◽  
Protein Kinase C Activity ◽  
High Calcium

The influence of phorbol myristate acetate (PMA), an activator of protein kinase c, on the secretion of parathyroid hormone from collagenase-dispersed bovine parathyroid cells was tested. The cells were incubated at low (0.5 mM) or high (2.0 mM) concentrations of calcium in the medium, and the hormone secreted into the medium was measured by a radioimmunoassay that recognizes both intact and C-terminal fragments of hormone. At low calcium, the secretory rate averaged 32 +/- 3.8 ng.h-1.(10(5) cells)-1. The addition of 1.6 microM PMA did not affect secretion. At high calcium there was a significant suppression of secretion by 38% to 19.8 +/- 3 ng.h-1.(10(5) cells)-1. The addition of 1.6 microM PMA significantly stimulated hormone secretion to 35.8 +/- 8 ng.h-1.(10(5) cells)-1, a rate indistinguishable from low calcium. This stimulatory effect of PMA at high calcium was seen at PMA concentrations as low as 1.6 nM, did not occur with a biologically inactive 4 alpha-isomer of phorbol ester, and was independent of changes in cellular adenosine 3',5'-cyclic monophosphate levels. Examination of 32P-labeled phosphoproteins by two-dimensional gel electrophoresis revealed acidic proteins of approximately 20,000 and 100,000 Da that were phosphorylated at low and high calcium + 1.6 microM PMA but not at high calcium alone. The protein kinase c activity associated with the membrane fraction of parathyroid cells significantly decreased 40% when the cells were incubated at high vs. low calcium. The data suggest that calcium may regulate parathyroid hormone secretion through changes in protein kinase c activity of the membrane fraction of the cell and protein phosphorylation.

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