Leucine-Rich Repeat Containing 10 (Lrrc10) Protein is a Novel Regulator of Cardiac Cav1.2 L-Type Calcium Channels

Leucine-rich repeat containing 10 (LRRC10) is a cardiac-specific protein that plays a critical role in cardiac function. We have demonstrated that the Lrrc10-null (Lrrc10-/-) mice develop dilated cardiomyopathy. Our recent data indicate that Lrrc10-/- cardiomyocytes exhibit reduced L-type Ca2+ channel (LTCC) current (ICa,L). However, it is unclear how LRRC10 regulates ICa,L in the heart. To investigate the role of LRRC10 in the regulation of LTCCs, we co-expressed the Myc tagged LRRC10 (LRRC10-Myc), heamagglutinin tagged Cav1.2 (Cav1.2-HA) and the auxiliary Cavβ2C subunit in HEK293 cells and performed co-immunoprecipitation (co-IP) on lysates using either anti-HA, anti-Myc antibody or control IgG. Western blot analysis demonstrated that Cav1.2 and LRRC10 associated with one another without the co-expression of Cavβ2C subunit. Also, the Cavβ2C and LRRC10 did not co-IP with one another suggesting that the LRRC10 may directly interact with Cav1.2 subunit. We then tested if a single point mutation H150A or triple point mutations Y104A, W127A and H150A would alter putative functional interaction sites in the LRRC10 and investigated if these mutations disrupt LRRC10 association with Cav1.2. Both LRRC10 mutants did not associate with Cav1.2. Additionally, co-IP analysis using mouse ventricular homogenates demonstrated that LRRC10 and Cav1.2 subunit are associated with one another. Finally, whole-cell patch clamp experiments performed in ventricular myocytes from Lrrc10-/- mice demonstrated a significant reduction in the ICa,L density (−2.5 0.2 pA/pF) and delayed inactivation, compared to WT myocytes (−6 0.6 pA/pF). In summary, we demonstrate that the LRRC10 and Cav1.2 subunit of LTCC may directly interact with one another and that mutations in LRRC10 residues, likely important for protein-protein interactions, disrupts this association. We conclude that LRRC10 is a novel and essential regulator of the LTCC function in ventricular myocytes.