Cloning of a retinally abundant regulator of G-protein signaling (RGS-r/RGS16): genomic structure and chromosomal localization of the human gene¹

Abstract

Regulators of G-protein signaling (RGS) constitute a family of GTPase-activating proteins with varying tissue-specific expression patterns and G-protein alpha subunit specificities. Here, we describe the molecular cloning of the human RGS-r/RGS16 cDNA, encoding a predicted polypeptide of 23 kDa that shows 86% identity to mouse RGS-r. Northern blot analysis shows that, like the mouse Rgs-r message, hRGS-r mRNA is abundantly expressed in retina, with lower levels of expression in most other tissues examined. Characterization of the genomic organization of the hRGS-r gene shows that it consists of five exons and four introns. We have also mapped the human RGS-r /RGS16 gene to chromosome 1q25–1q31 by fluorescence in situ hybridzation. Analysis of human ESTs reveals that at least five members of the RGS gene family map to chromosome 1q, suggesting that at least part of the RGS family arose through gene duplication. The chromosomal location, retinal abundance, and presumed function of the human RGS-r protein in desensitizing photoreceptor signaling make the RGS-r/RGS16 locus a candidate for mutations responsible for retinitis pigmentosa with para-arteriolar preservation of retinal pigment epithelium (RP-PPRE or RP12), an autosomal recessive disorder previously mapped to 1q31.

Introduction

Photons, odors, hormones, neurotransmitters, and other sensory stimuli activate signal transduction cascades via heterotrimeric G-protein-coupled receptors [reviewed in Hamm and Gilchrist (1996)]. Signaling by G-protein-coupled receptors is achieved by regulation of the activation state of its coupled heterotrimeric G-protein. In the GDP-bound state, the G_α subunit is inactive and forms a heterotrimeric complex with G_βγ subunits. The binding of agonist causes a conformational change in the receptor allowing it to be converted to an active state that promotes G_α to release GDP and to bind GTP. Once activated, the heterotrimeric G-protein dissociates into the GTP-bound G_α and G_βγ subunits that function as effector molecules to modulate cellular enzymes and ion channnels. Signal transduction continues until GTP is hydrolyzed and the subunits reassociate, thereby preventing further receptor signaling. In vertebrate photoreceptors, the intrinsic GTPase activity of the transducin alpha-subunit is stimulated by its effector, cGMP phosphodiesterase (Arshavsky and Bownds, 1992). However, in vitro experiments with purified G_α transducin subunits and cGMP phosphodiesterase have shown that the GTP hydrolysis rate observed in vitro cannot account for the rapid desensitization of the photoresponse observed in vivo (Angleson and Wensel, 1993). This discrepancy has provoked the search for factors that might accelerate the GTPase activity of the G_α subunits — so-called GAPs or `GTPase activating proteins'.

Recently, several groups have identified a highly conserved group of proteins called RGSs, for regulators of G-protein signaling, that function as GAPs for G_α subunits [reviewed in Dohlman and Thorner (1997)]. RGS proteins are defined by a conserved, tripartite domain that is presumed to function in desensitizing G-protein mediated signaling pathways by accelerating the intrinsic GTP hydrolysis activity of specific G-protein alpha subunits (Berman et al., 1996; Hunt et al., 1996; Watson et al., 1996); in addition, RGS proteins can also act as competitive inhibitors of effector protein binding to G_α subunits (Hepler et al., 1997). At present, the mammalian RGS family consists of 17 members: GAIP (De Vries et al., 1995), RGS1 through RGS15 (Koelle and Horvitz, 1996; Siderovski et al., 1996; Druey et al., 1996) and RGS-r (Chen et al., 1996).

In this paper, we describe the cloning of the human RGS-r cDNA, its mRNA expression in different adult tissues, and the structural characterization and chromosomal localization of the hRGS-r/hRGS16 gene to human chromosome 1q25–31.