TY - JOUR
T1 - Inhibition by toxin B of inositol phosphate formation induced by G protein-coupled and tyrosine kinase receptors in N1E-115 neuroblastoma cells
T2 - Involvement of Rho proteins
AU - Zhang, Chunyi
AU - Schmidt, Martina
AU - Von Eichel-Streiber, Christoph
AU - Jakobs, Karl H.
PY - 1996/10
Y1 - 1996/10
N2 - G protein-coupled receptors activate phospholipase C (PLC)-β isoforms by the α or βγ subunits of G proteins, whereas growth-factor receptors activate PLC-γ isoforms by phosphorylating tyrosine residues of the enzyme. As a common substrate for PLC enzymes, phosphatidylinositol 4,5-bisphosphate [Ptdlns(4,5)P2] may play a pivotal role in the regulation of cellular PLC activity. Because small-molecular-weight G proteins have been implicated in the synthesis of Ptdlns(4,5)P2, we studied the effect of Clostridium difficile toxin B, which glucosylates and thereby inactivates small G proteins of the Rho family, on receptor-stimulated PLC activity. We report here that in N1E-115 neuroblastoma cells, stimulation of inositol phosphate formation by the G protein-coupled receptor agonists bradykinin and lysophosphatidic acid and by the tyrosine kinase receptor agonist platelet- derived growth factor is largely attenuated by toxin B treatment. Furthermore, inositol phosphate production stimulated by the stable GTP analog guanosine 5'-O-(3-thio)-triphosphate in permeabilized N1E-115 cells was inhibited by C3 exoenzyme, which specifically inactivates Rho proteins. The inhibition by toxin B was apparently not caused by its effect on the cytoskeleton. In addition, the level of platelet-derived growth factor receptors, which was studied with immunoblotting, was unaffected by toxin B. Using exogenous Ptdlns(4,5)P2 as PLC substrate, it was found that the intrinsic enzymatic activity of PLC activated either by Ca2+ or by guanosine 5'-O(3-thio)triphosphate was not altered by toxin B. However, toxin B decreased strongly, by up to 80%, the cellular level of Ptdlns(4,5)P2 in a concentration-dependent manner, without changing those of phosphatidylinositol and phosphatidylinositol 4-phosphate. These results, together with the recent finding that Rho family proteins can regulate phosphatidyiinositol 4-phosphate 5-kinase activity, demonstrate that Rho proteins are presumably important regulators of Ptdlns(4,5)P2 synthesis and, thereby, play an integral role in the regulation of cellular signaling by PLC enzymes.
AB - G protein-coupled receptors activate phospholipase C (PLC)-β isoforms by the α or βγ subunits of G proteins, whereas growth-factor receptors activate PLC-γ isoforms by phosphorylating tyrosine residues of the enzyme. As a common substrate for PLC enzymes, phosphatidylinositol 4,5-bisphosphate [Ptdlns(4,5)P2] may play a pivotal role in the regulation of cellular PLC activity. Because small-molecular-weight G proteins have been implicated in the synthesis of Ptdlns(4,5)P2, we studied the effect of Clostridium difficile toxin B, which glucosylates and thereby inactivates small G proteins of the Rho family, on receptor-stimulated PLC activity. We report here that in N1E-115 neuroblastoma cells, stimulation of inositol phosphate formation by the G protein-coupled receptor agonists bradykinin and lysophosphatidic acid and by the tyrosine kinase receptor agonist platelet- derived growth factor is largely attenuated by toxin B treatment. Furthermore, inositol phosphate production stimulated by the stable GTP analog guanosine 5'-O-(3-thio)-triphosphate in permeabilized N1E-115 cells was inhibited by C3 exoenzyme, which specifically inactivates Rho proteins. The inhibition by toxin B was apparently not caused by its effect on the cytoskeleton. In addition, the level of platelet-derived growth factor receptors, which was studied with immunoblotting, was unaffected by toxin B. Using exogenous Ptdlns(4,5)P2 as PLC substrate, it was found that the intrinsic enzymatic activity of PLC activated either by Ca2+ or by guanosine 5'-O(3-thio)triphosphate was not altered by toxin B. However, toxin B decreased strongly, by up to 80%, the cellular level of Ptdlns(4,5)P2 in a concentration-dependent manner, without changing those of phosphatidylinositol and phosphatidylinositol 4-phosphate. These results, together with the recent finding that Rho family proteins can regulate phosphatidyiinositol 4-phosphate 5-kinase activity, demonstrate that Rho proteins are presumably important regulators of Ptdlns(4,5)P2 synthesis and, thereby, play an integral role in the regulation of cellular signaling by PLC enzymes.
UR - http://www.scopus.com/inward/record.url?scp=0029912023&partnerID=8YFLogxK
M3 - Article
C2 - 8863831
AN - SCOPUS:0029912023
SN - 0026-895X
VL - 50
SP - 864
EP - 869
JO - Molecular Pharmacology
JF - Molecular Pharmacology
IS - 4
ER -