A Novel G Protein α Subunit Containing Atypical Guanine Nucleotide-binding Domains Is Differentially Expressed in a Molluscan Nervous System

Jaco C Knol,Arno R Van Der Slik,Ellen R Van Kesteren,Rudi J Planta,Harm Van Heerikhuizen,Erno Vreugdenhil

doi:10.1074/jbc.270.32.18804

Abstract

We described the characterization of a novel G protein alpha subunit, G alpha a. cDNA encoding this subunit was cloned from the central nervous system of the mollusc Lymnaea stagnalis. The deduced protein contains all characteristic guanine nucleotide-binding domains of G alpha subunits but shares only a limited degree of overall sequence identity with known subtypes (approximately 30%). Moreover, two of the nucleotide-binding domains exhibit salient deviations from corresponding sequences in other G protein alpha subunits. The A domain, determining kinetic features of the GTPase cycle, contains a markedly unique amino acid sequence (ILIIGGPGAGK). In addition, the C domain is also clearly distinct (DVAGQRSL). The presence of a leucine in this motif, instead of glutamic acid, has important implications for hypotheses concerning the GTPase mechanism. In contrast to other G alpha subtypes, G alpha a has no appropriate N-terminal residues that could be acylated. It does contain the strictly conserved arginine residue that serves as a cholera toxin substrate in G alpha s and G alpha t but lacks a site for ADP-ribosylation by pertussis toxin. In situ hybridization experiments indicate that G alpha a-encoding mRNA is expressed in a limited subpopulation of neurons within the Lymnaea brain. These data suggest that G alpha a defines a separate class of G proteins with cell type-specific functions.

Highlights

Heterotrimeric G proteins (Ga{3y) form a family of molecular go-betweens that couple stimulus-triggered cell surface receptors to response-generating effectors within the cell [1,2,3]
We have cloned a diverse set of snail Go subunits, i.e. Gao' Gai' Gas' and Gaq, as well as a G{3 subunit [8,9,10]. All of these Lymnaea subunits appear to be remarkably similar to their mammalian counterparts (76-82% amino acid sequence identity). Taking into consideration such a striking resemblance and the fact that at least 16 Ga subtypes exist in mammals, it seemed reasonable to assume that Lymnaea expresses more than merely four G protein a subunits
Identification of a Novel G Protein 0' Subunit Expressed in Lymnaea-Degenerate oligonucleotides were based on well conserved GO' amino acid motifs within domains C and G of the nucleotide-binding pocket (Ref. 16; see "Materials and Methods")

Summary

Introduction

Heterotrimeric G proteins (Ga{3y) form a family of molecular go-betweens that couple stimulus-triggered cell surface receptors to response-generating effectors within the cell [1,2,3]. Specific G protein subtypes can interact with more than one receptor or effector subtype [1] Such a promiscuity might, allow the integration or distribution of extracellular signals in vivo [4]. One system offering such opportunities is the simple central nervous system of the pond snail, Lymnaea stagnalis, which we use to study the function of G protein-mediated signaling networks in neuronal information processing. We have cloned a diverse set of snail Go subunits, i.e. Gao' Gai' Gas' and Gaq , as well as a G{3 subunit [8,9,10] All of these Lymnaea subunits appear to be remarkably similar to their mammalian counterparts (76-82% amino acid sequence identity). Our findings indicate that the G protein family is yet more elaborate, implying that additional members remain to be discovered

Methods

Results

Conclusion