Molecular heterogeneity of calcium channel β-subunits in canine and human heart: evidence for differential subcellular localization

Abstract

Multiple Ca 2+ channel β-subunit (Ca v β) isoforms are known to differentially regulate the functional properties and membrane trafficking of high-voltage-activated Ca 2+ channels, but the precise isoform expression pattern of Ca v β subunits in ventricular muscle has not been fully characterized. Using sequence data from the Human Genome Project to define the intron/exon structure of the four known Ca v β genes, we designed a systematic RT-PCR strategy to screen human and canine left ventricular myocardial samples for all known Ca v β isoforms. A total of 18 different Ca v β isoforms were detected in both canine and human ventricles including splice variants from all four Ca v β genes. Six of these isoforms have not previously been described. Western blots of ventricular membrane fractions and immunocytochemistry demonstrated that all four Ca v β subunit genes are expressed at the protein level, and the Ca v β subunits show differential subcellular localization with Ca v β 1b , Ca v β 2 , and Ca v β 3 predominantly localized to the T-tubule sarcolemma, whereas Ca v β 1a and Ca v β 4 are more prevalent in the surface sarcolemma. Coexpression of the novel Ca v β 2c subunits (Ca v β 2cN1 , Ca v β 2cN2 , Ca v β 2cN4 ) with the pore-forming α 1C (Ca v 1.2) and Ca v α 2 δ subunits in HEK 293 cells resulted in a marked increase in ionic current and Ca v β 2c isoform-specific modulation of voltage-dependent activation. These results demonstrate a previously unappreciated heterogeneity of Ca v β subunit isoforms in ventricular myocytes and suggest the presence of different subcellular populations of Ca 2+ channels with distinct functional properties.