L-type Ca2+ channel C terminus calmodulin (CaM)-binding domains are molecular determinants for Ca2+-CaM-dependent increases in L-type Ca2+ current (ICa), and a CaM-binding IQ domain mimetic peptide (IQmp) increases L-type Ca2+ channel current by promoting a gating mode with prolonged openings (mode 2), suggesting the intriguing possibility that CaM-binding domains are 'auto-agonist' signalling molecules. In order to test the breadth of this concept, we studied the effect of a second C terminus CaM-binding domain (CB) mp (CBmp), in conjunction with IQmp, on single L-type Ca2+ channel currents in excised cell membrane patches from rabbit ventricular myocytes. Here we show that both CBmp and IQmp are agonist ligands that non-additively increase L-type Ca2+ channel opening probability (Po) by inducing mode 2 gating. CBmp and IQmp agonist effects were lost under conditions favouring calcification of CaM (Ca2+-CaM, 150 nM free Ca2+ and 10-20 μM CaM), but persisted in the presence of CaM (0-20 μM) under conditions adverse to Ca2+-CaM (20 mM BAPTA), indicating that CaM-binding domains increase L-type Ca2+ channel Po by a low Ca2+-CaM activity mechanism. Increasing Ca2+-CaM in the bath (cytosol) reduced the efficacy of CBmp and IQmp signals with Ba2+ as charge carrier, suggesting that CaM binding motifs target a site outside of the pore region. We measured the combined effects of CBmp and Ca2+-CaM-dependent protein kinase II (CaMKII) on L-type Ca2+ channels by using an engineered Ca2+-CaM-independent form of CaMKII that remains active under low Ca2+-CaM conditions, permissive for CBmp signalling. CBmp and CaMKII increased L-type Ca2+ channel Po in a non-additive manner, suggesting that low and high Ca2+-CaM-dependent L-type Ca2+ channel facilitation pathways converge upon a common signalling mechanism.
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