Seven Helix cAMP Receptors Stimulate Ca Entry in the Absence of Functional G Proteins in Dictyostelium

doi:10.1074/jbc.270.11.5926

Seven Helix cAMP Receptors Stimulate Ca Entry in the Absence of Functional G Proteins in Dictyostelium(*)

From the Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205-2185

^§§ To whom correspondence should be addressed:
the Dept. of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, MD 21205-2185.
Tel.: 410-955-4699; Fax: 410-955-5759.

↵^¶ Current address: Leukosite, Inc., 215 First St., Cambridge, MA 02142.

Abstract

Surface cAMP receptors (cARs) in Dictyostelium transmit a variety of signals across the plasma membrane. The best characterized cAR, cAR1, couples to the heterotrimeric guanine nucleotide-binding protein (G protein) α-subunit Gα2 to mediate activation of adenylyl and guanylyl cyclases and cell aggregation. cAR1 also elicits other cAMP-dependent responses including receptor phosphorylation, loss of ligand binding (LLB), and Ca influx through a Gα2-independent pathway that may not involve G proteins. Here, we have expressed cAR1 and a related receptor, cAR3, in a gβ strain (Lilly, P., Wu. L., Welker, D. L., and Devreotes, P. N.(1993) Genes & Dev. 7, 986-995), which lacks G protein activity. Both cell lines failed to aggregate, a process requiring the Gα2 and Gβ-subunits. In contrast, cAR1 phosphorylation in cAR1/gβ cells showed a time course and cAMP dose dependence indistinguishable from those of cAR1/Gβ controls. cAMP-induced LLB was also normal in the cAR1/gβ cells. Finally, cAR1/gβ cells and cAR3/gβ cells showed a Ca response with kinetics, agonist dependence, ion specificity, and sensitivity to depolarization agents that were like those of Gβ controls, although they accumulated fewer Ca ions per cAMP receptor than the control strains. Together, these results suggest that the Gβ-subunit is not required for the activation or attenuation of cAR1 phosphorylation, LLB, or Ca influx. It may, however, serve to amplify the Ca response, possibly by modulating other intracellular Ca signal transduction pathways.

Footnotes

↵^§ Recipient of a Centennial Fellowship from the Medical Research Council of Canada.
↵** Supported by a National Institutes of Health Medical Scientist Training Program grant at The Johns Hopkins School of Medicine.
↵* This study was supported in part by National Institutes of Health Grant GM28007 (to P. N. D.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

cAR

cAMP receptor

CCCP

carbonylcyanide-m-chlorophenylhydrazone

IP₃

inositol 1,4,5-trisphosphate

LLB

loss of ligand binding.
↵² J. M. Louis and A. Kimmel, personal communication.
↵³ L. Wu, H. Kuwayama, P. Van Dijkan, P. Van Haastert, and P. N. Devreotes, manuscript in preparation.
↵⁴ R. L. Johnson and P. N. Devreotes, unpublished observations.
↵⁵ Caterina, M. J., Hereld, D., and Devreotes, P. N.(1995) J. Biol. Chem.270, 4418-4423.
↵⁶ J. L. S. Milne, unpublished observations.
- Received September 12, 1994.
- Revision received December 20, 1994.