Leukemia inhibitory factor activates cardiac L-type Ca²⁺ channels via phosphorylation of serine 1829 in the rabbit Ca_v1.2 subunit

We have previously reported that leukemia inhibitory factor (LIF) gradually increased cardiac L-type Ca²⁺ channel current (I _CaL), which peaked at 15 minutes in both adult and neonatal rat cardiomyocytes, and this increase was blocked by the mitogen-activated protein kinase kinase inhibitor PD98059. This study investigated the molecular basis of LIF-induced augmentation of I_CaL in rodent cardiomyocytes. LIF induced phosphorylation of a serine residue in the α_1c subunit (Ca_v1.2) of L-type Ca²⁺ channels in cultured rat cardiomyocytes, and this phosphorylation was inhibited by PD98059. When constructs encoding either a wild-type or a carboxyl-terminal-truncated rabbit Ca_v1.2 subunit were transfected into HEK293 cells, LIF induced phosphorylation of the resultant wild-type protein but not the mutant protein. Cotransfection of constitutively active mitogen-activated protein kinase kinase also resulted in phosphorylation of the Ca_v1.2 subunit in the absence of LIF stimulation. In in-gel kinase assays, extracellular signal-regulated kinase phosphorylated a glutathione S-transferase fusion protein of the carboxyl-terminal region of Ca_v1.2 (residues 1700 through 1923), which contains the consensus sequence Pro-Leu-Ser-Pro. A point mutation within this consensus sequence, which results in a substitution of alanine for serine at residue 1829 (S1829A), was sufficient to abolish the LIF-induced phosphorylation. LIF increased I_CaL in HEK cells transfected with wild-type Ca_v1.2 but not with the mutated version. These results provide direct evidence that LIF phosphorylates the sefine residue at position 1829 of the Ca_v1.2 subunit via the actions of extracellular signal-regulated kinase and that this phosphorylation increases I_CaL in cardiomyocytes.