Mechanism of phospholipase D activation induced by extracellular ATP in osteoblast-like cells

1995 ◽  
Vol 145 (1) ◽  
pp. 81-86 ◽  
Author(s):  
A Suzuki ◽  
J Shinoda ◽  
Y Oiso ◽  
O Kozawa
Keyword(s):  
Phospholipase D ◽  
Inositol Phosphates ◽  
Binding Protein ◽  
Extracellular Atp ◽  
Prostaglandin E ◽  
Dependent Manner ◽  
Gtp Binding Protein ◽  
Proliferative Effect ◽  
Gtp Binding ◽  
Pkc Activation

Abstract We have previously reported that extracellular ATP stimulates Ca2+ influx from extracellular space, resulting in the production of prostaglandin E2 which mediates, at least in part, its proliferative effect on osteoblast-like MC3T3-E1 cells, and that the activation of protein kinase C (PKC) stimulates phospholipase D in these cells. In the present study, we examined the effect of extracellular ATP on phosphatidylcholine-hydrolysing phospholipase D activity in MC3T3-E1 cells. ATP stimulated the formation of both choline and inositol phosphates dose-dependently in the range between 0·1 and 1 mm. The formation of choline by a combination of ATP and NaF, an activator of GTP-binding protein, was synergistic, whereas that of inositol phosphates was not. A combination of ATP and 12-O-tetradecanoylphorbol-13-acetate, a PKC activating phorbol ester, additively stimulated the formation of choline. Staurosporine, an inhibitor of PKC, had little effect on ATP-stimulated formation of choline. Choline formation was significantly reduced by chelating extracellular Ca2+ with EGTA, while being inhibited by W-7, an antagonist of calmodulin. These results suggest that extracellular ATP stimulates phospholipase D in a Ca2+/calmodulin-dependent manner in osteoblast-like cells, and that neither PKC activation nor GTP-binding protein is involved in this mechanism. Journal of Endocrinology (1995) 145, 81–86

Rabphilin-3A, a putative target protein for smg p25A/rab3A p25 small GTP-binding protein related to synaptotagmin

1993 ◽  
Vol 13 (4) ◽  
pp. 2061-2068
Author(s):  
H Shirataki ◽  
K Kaibuchi ◽  
T Sakoda ◽  
S Kishida ◽  
T Yamaguchi ◽  
...  
Keyword(s):  
Binding Protein ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Bovine Brain ◽  
Target Protein ◽  
Dependent Manner ◽  
Gtp Binding Protein ◽  
Reading Frame ◽  
Gtp Binding ◽  
Putative Target

In a previous study (H. Shirataki, K. Kaibuchi, T. Yamaguchi, K. Wada, H. Horiuchi, and Y. Takai, J. Biol. Chem. 267:10946-10949, 1992), we highly purified from bovine brain crude membranes the putative target protein for smg p25A/rab3A p25, a ras p21-related small GTP-binding protein implicated in neurotransmitter release. In this study, we have isolated and sequenced the cDNA of this protein from a bovine brain cDNA library. The cDNA had an open reading frame encoding a protein of 704 amino acids with a calculated M(r) of 77,976. We tentatively refer to this protein as rabphilin-3A. Structural analysis of rabphilin-3A revealed the existence of two copies of an internal repeat that were homologous to the C2 domain of protein kinase C as described for synaptotagmin, which is known to be localized in the membrane of the synaptic vesicle and to bind to membrane phospholipid in a Ca(2+)-dependent manner. The isolated cDNA was expressed in COS7 cells, and the encoded protein was recognized with an anti-rabphilin-3A polyclonal antibody and was identical in size with rabphilin-3A purified from bovine brain by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Moreover, both rabphilin-3A purified from bovine brain and recombinant rabphilin-3A made a complex with the GTP gamma S-bound form of rab3A p25 but not with the GDP-bound form of rab3A p25. Immunoblot and Northern (RNA) blot analyses showed that rabphilin-3A was highly expressed in bovine and rat brains. These results indicate that rabphilin-3A is a novel protein that has C2 domains and selectively interacts with the GTP-bound form of rab3A p25.

Vasopressin activates phospholipase D through pertussis toxin-insensitive GTP-binding protein in aortic smooth muscle cells: function of Ca2+/calmodulin

1995 ◽  
Vol 73 (3-4) ◽  
pp. 191-199 ◽  
Author(s):  
Masaichi Miwa ◽  
Atsushi Suzuki ◽  
Yasuko Watanabe ◽  
Junji Shinoda ◽  
Yutaka Oiso ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Phospholipase D ◽  
Pertussis Toxin ◽  
Binding Protein ◽  
Muscle Cells ◽  
Gtp Binding Protein ◽  
Aortic Smooth Muscle Cells ◽  
Aortic Smooth Muscle ◽  
Gtp Binding

In the present study, we examined the effect of vasopressin (AVP) on phosphatidylcholine-hydrolyzing phospholipase D activity in primary cultured rat aortic smooth muscle cells. AVP stimulation of choline formation was dose dependent. The time-course was quite different from those of inositol phosphates. The effect of AVP on the formation of inositol phosphates (EC50 was 3 nM) was more potent than that on the formation of choline (EC50 was 30 nM). 12-O-Tetradecanoylphorbol-13-acetate (TPA), an activator of protein kinase C (PKC), stimulated the formation of choline. However, 4α-phorbol 12,13-didecanoate, which is inactive for PKC, had little effect. Staurosporine, an inhibitor of protein kinases, which inhibited the TPA-induced formation of choline, had little effect on the AVP-induced formation of choline. Neither calphostin C, a highly specific PKC inhibitor, nor PKC down-regulation with TPA affected AVP-induced formation of choline. A combination of AVP and TPA additively stimulated the formation of choline. The depletion of extracellular Ca2+ by (ethylenebis(oxyethylenenitrilo))tetraacetic acid significantly reduced the AVP-induced formation of choline. W-7, an antagonist of calmodulin, inhibited the AVP-induced formation of choline in a dose-dependent manner. NaF, an activator for GTP-binding protein (G-protein), stimulated the formation of choline. However, the formation of choline by a combination of AVP and NaF was not additive. Pertussis toxin had little effect on the AVP-induced formation of choline. These results strongly suggest that AVP stimulates phospholipase D in a Ca2+/calmodulin-dependent manner in aortic smooth muscle cells, that a pertussis-toxin-insensitive G-protein is involved in the AVP-induced phospholipase D activation, and furthermore, that PKC is not essential for the activation.Key words: vasopressin, phospholipase D, protein kinase C, calmodulin, GTP-binding protein, aortic smooth muscle cells.

The C-Terminus of the Prostaglandin-E-Receptor EP3 Subtype is Essential for Activation of GTP-Binding Protein

1994 ◽  
Vol 224 (1) ◽  
pp. 161-166 ◽  
Author(s):  
Atsushi Irie ◽  
Yukihiko Sugimoto ◽  
Tsunehisa Namba ◽  
Tomiko Asano ◽  
Atsushi Ichikawa ◽  
...  
Keyword(s):  
Binding Protein ◽  
Prostaglandin E ◽  
Gtp Binding Protein ◽  
Gtp Binding ◽  
C Terminus

scholarly journals Increased Activity of Small GTP-binding Protein-dependent Phospholipase D during Differentiation in Human Promyelocytic Leukemic HL60 Cells

1997 ◽  
Vol 272 (3) ◽  
pp. 1990-1996 ◽  
Author(s):  
Kenji Ohguchi ◽  
Shigeru Nakashima ◽  
Zhiming Tan ◽  
Yoshiko Banno ◽  
Shuji Dohi ◽  
...  
Keyword(s):  
Phospholipase D ◽  
Binding Protein ◽  
Gtp Binding Protein ◽  
Hl60 Cells ◽  
Gtp Binding ◽  
Increased Activity

scholarly journals A human homologue of the yeast GST1 gene codes for a GTP-binding protein and is expressed in a proliferation-dependent manner in mammalian cells.

1989 ◽  
Vol 8 (12) ◽  
pp. 3807-3814 ◽  
Author(s):  
S. Hoshino ◽  
H. Miyazawa ◽  
T. Enomoto ◽  
F. Hanaoka ◽  
Y. Kikuchi ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Binding Protein ◽  
Dependent Manner ◽  
Human Homologue ◽  
Gtp Binding Protein ◽  
Gtp Binding
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