A Cluster of Three Novel Ca 2 Channel  Subunit Genes on Chromosome 19q13.4: Evolution and Expression Profile of the  Subunit Gene Family Daniel L. Burgess,* ,1 Lisa A. Gefrides,* Perry J. Foreman,* and Jeffrey L. Noebels * , † , ‡ * Department of Neurology, †Department of Molecular and Human Genetics, and ‡Division of Neuroscience, Baylor College of Medicine, One Baylor Plaza, Houston, Texas 77030 Received August 10, 2000; accepted November 7, 2000 The CACNG1 gene on chromosome 17q24 encodes an integral membrane protein that was originally iso- lated as the regulatory  subunit of voltage-dependent Ca 2 channels from skeletal muscle. The existence of an extended family of  subunits was subsequently demonstrated upon identification of CACNG2 (22q13), CACNG3 (16p12–p13), and CACNG4 and CACNG5 (17q24). In this study, we describe a cluster of three novel  subunit genes, CACNG6, CACNG7, and CACNG8, located in a tandem array on 19q13.4. Phylo- genetic analysis indicates that this array is paralogous to the cluster containing CACNG1, CACNG5, and CACNG4, respectively, on chromosome 17q24. We de- veloped sensitive RT-PCR assays and examined the expression profile of each member of the  subunit gene family, CACNG1–CACNG8. Analysis of 24 human tissues plus 3 dissected brain regions revealed that CACNG1 through CACNG8 are all coexpressed in fetal and adult brain and differentially transcribed among a wide variety of other tissues. The expression of dis- tinct complements of  subunit isoforms in different cell types may be an important mechanism for regu- lating Ca 2 channel function. © 2001 Academic Press INTRODUCTION Voltage-dependent Ca 2+ channels are expressed in a wide variety of tissues where they couple membrane excitation to critical cellular events such as muscle contraction, neurotransmitter and hormone release, neurite outgrowth, synaptogenesis, synaptic plasticity, and gene expression (Tsien and Tsien, 1990). In mus- cle, the subunit structure of the Ca 2+ channel exists as a heteromultimeric complex of  1 , ,  2 , and  sub- units associating with a 1:1:1:1 stoichiometry (Taka- hashi et al., 1987; reviewed by De Waard et al., 1996). The pore-forming  1 subunit contains the basic struc- tural elements required for voltage-sensing and gating, while the auxiliary ,  2 , and  subunits modulate these biophysical properties (Singer et al., 1991; De Waard et al., 1996). Both in vitro and in vivo analyses suggest that the  subunit alters channel activity by increasing steady state inactivation, thereby inhibiting Ca 2+ currents (Singer et al., 1991; Letts et al., 1998; Freise et al., 2000; Klugbauer et al., 2000; Sipos et al., 2000). Subsequent examination of Ca 2+ channel com- plexes in brain and other organs confirmed the bio- chemical association of  1 , , and  2  subunits but, surprisingly, did not identify any -like subunit mRNA or proteins in nonmuscle tissues (Ahlijanian et al., 1990; Jay et al., 1990; Witcher et al., 1993; Liu et al., 1996). Current genetic studies reveal that Ca 2+ channel subunits can be encoded by one of multiple, related loci. To date, 10 genes have been isolated and mapped that encode  1 subunit isoforms ( 1A - 1I ,  1S ), 4 that encode cytoplasmic  subunits ( 1 – 4 ) and 3 that en- code transmembrane  2  subunits ( 2  1 – 2  3 ). A single gene encoding the skeletal muscle  subunit (CACNG1) was isolated and localized to human chro- mosome 17q23 (Powers et al., 1993). Evidence for a second  subunit gene was obtained with the recent isolation of the molecular defect in the mouse stargazer mutant (stg). Stargazer homozygotes exhibit a complex neurological disorder consisting of ataxia, inner-ear defects, and epilepsy (Noebels et al., 1990). Positional cloning of two stargazer alleles identified a novel, neu- ronal specific gene encoding a 36-kDa protein with structural similarity to the Ca 2+ channel  1 subunit. The gene was designated Cacng2 (Letts et al., 1998). The existence of a single genetic isoform in neurons distinguished the  subunit from the  1 , , and  2  subunits that utilize genetic heterogeneity, coexpres- sion of multiple isoforms, and combinatorial diversity as mechanisms to generate a wide array of Ca 2+ chan- nel compositions and functional variants in the central nervous system (CNS) (Burgess and Noebels, 1999). Sequence data from this article have been deposited with the EMBL/GenBank Data Libraries under Accession Nos. (CACNG6) AF288386, (CACNG7) AF288387, and (CACNG8) AF288388. 1 To whom correspondence should be addressed. Telephone: (713) 798-5862. Fax: (713) 798-7528. E-mail: dburgess@bcm.tmc.edu. Genomics 71, 339 –350 (2001) doi:10.1006/geno.2000.6440, available online at http://www.idealibrary.com on 339 0888-7543/01 $35.00 Copyright © 2001 by Academic Press All rights of reproduction in any form reserved.