Pertussis Toxin-sensitive Activation of Phospholipase C by the C5a and fMet-Leu-Phe Receptors*
- From the ‡ Department of Pharmacology and Physiology and Biochemistry, University of Rochester, Rochester, New York 14534 and
- § Division of Biology, California Institute of Technology, Pasadena, California 91125
- ¶ To whom correspondence should be addressed. Tel.: 716-275-2029; Fax: 716-244-9283.
Abstract
Signal transduction pathways that mediate C5a and fMet-Leu-Phe (fMLP)-induced pertussis toxin (PTx)-sensitive activation of phospholipase C (PLC) have been investigated using a cotransfection assay system in COS-7 cells. The abilities of the receptors for C5a and fMLP to activate PLC β2 and PLC β3 through the Gβγ subunits of endogenous Gi proteins in COS-7 cells were tested because both PLC β2 and PLC β3 were shown to be activated by the βγ subunits of G proteins in in vitro reconstitution assays. Neither of the receptors can activate endogenous PLC β3 or recombinant PLC β3 in transfected COS-7 cells. However, both receptors can clearly activate PLC β2 in a PTx-sensitive manner, suggesting that the receptors may interact with endogenous PTx-sensitive G proteins and activate PLC β2 probably through the Gβγ subunits. These findings were further corroborated by the results that PLC β3 could only be slightly activated by Gβ1γ1 or Gβ1γ5 in the cotransfection assay, whereas the Gβγ subunits strongly activated PLC β2 under the same conditions. PLC β3 can be activated by Gαq, Gα11, and Gα16 in the cotransfection assay. In addition, the Gγ2 and Gγ3 mutants with substitution of the C-terminal Cys residue by a Ser residue, which can inhibit wild type Gβγ-mediated activation of PLC β2, were able to inhibit C5a or fMLP-mediated activation of PLC β2. These Gγ mutants, however, showed little effect on m1-muscarinic receptor-mediated PLC activation, which is mediated by the Gq class of G proteins. These results all confirm that the Gβγ subunits are involved in PLC β2 activation by the two chemoattractant receptors and suggest that in COS-7 cells activation of PLC β3 by Gβγ may not be the primary pathway for the receptors.
Footnotes
-
↵* This work was supported by National Institutes of Health grants (to D. W., and M. I. S.) and by an American Cancer Society Institutional Grant IRG-18 (to H. J.). The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
-
↵1 The abbreviations used are:
- G protein
-
GTP-binding protein
- C5a
-
complement factor 5a
- fMLP
-
formyl-Met-Leu-Phe
- GγCS
-
Gγ mutants with substitution of C-terminal Cys residues by Ser residues
- PLC
-
phospholipase C
- PTx
-
pertussis toxin
- IP
-
inositol phosphate
- IL-8
-
interleukin-8.
-
- Received February 15, 1996.
- © 1996 by The American Society for Biochemistry and Molecular Biology, Inc.
