The regulators of G-protein signaling (RGS) proteins are important regulatory and structural components of G-protein coupled receptor complexes. RGS proteins are GTPase activating proteins (GAPs) of Gi-and Gq-class Galpha proteins, and thereby accelerate signaling kinetics and termination. Here, we mapped the chromosomal positions of all 21 Rgs genes in mouse, and determined human RGS gene structures using genomic sequence from partially assembled bacterial artificial chromosomes (BACs) and Celera fragments. In mice and humans, 18 of 21 RGS genes are either tandemly duplicated or tightly linked to genes encoding other components of G-protein signaling pathways, including Galpha, Ggamma, receptors (GPCR), and receptor kinases (GPRK). A phylogenetic tree revealed seven RGS gene subfamilies in the yeast and metazoan genomes that have been sequenced. We propose that similar systematic analyses of all multigene families from human and other mammalian genomes will help complete the assembly and annotation of the human genome sequence.