The Cavβ1 Subunit Regulates Gene Expression in Muscle Progenitor Cells

Abstract

CaVβ subunits are traditionally considered constituents of CaV complexes (CaV1or2, CaVβ, and CaV α2/δ), where they localize at the plasma membrane and serve to regulate channel expression and gating properties. Recent publications also show CaVβ subunit localization in the nucleus. This phenomenon has been observed under a variety of conditions (different cell types, β subunit isoforms, co-expressed proteins, etc). However, the mechanisms responsible for CaVβ subunit nuclear shuttling, as well as a physiological role for this nuclear localization, remain major questions. Ongoing work in our laboratory has shown that muscle progenitor cells (myoblasts) express CaVβ1 protein (but not CaV1 subunits) in both the cytoplasm and nucleus and that the loss of CaVβ1 expression impairs proliferation in these cells. To better understand the mechanisms that link CaVβ1 nuclear localization with control of proliferation, we have conducted large-scale screening experiments designed to identify which genes are directly regulated by CaVβ1, as well as its protein binding partners. To test if CaVβ1 may regulate gene expression, we conducted microarray experiments on RNA extracted from wild type, heterozygous, and CaVβ1 -null mouse primary myoblasts. A number of gene transcripts were found to be differentially regulated based on the relative amount of CaVβ1 expression. To identify specific CaVβ1 target genes, we performed chromatin immunoprecipitation -on-a-chip experiments to locate which promoter regions CaVβ1 bound to across the entire mouse genome. Nuclear binding partners of CaVβ1 were screened using affinity purification of CaVβ1a-YFP from myoblast nuclear fractions coupled with mass spectrometry. Finally, the importance of CaVβ1 in embryonic myogenesis was explored in CaVβ1 -null mice. Our results support the idea of CaVβ subunits acting as transcription factors and regulating gene expression independently from CaV's, and suggest these functions may be particularly important to progenitor cell growth.