α5β1 Integrin Engagement Increases Large Conductance, Ca2+-activated K+ Channel Current and Ca2+ Sensitivity through c-src-mediated Channel Phosphorylation*

Large conductance, calcium-activated K+ (BK) channels are important regulators of cell excitability and recognized targets of intracellular kinases. BK channel modulation by tyrosine kinases, including focal adhesion kinase and c-src, suggests their potential involvement in integrin signaling. Recently, we found that fibronectin, an endogenous α5β1 integrin ligand, enhances BK channel current through both Ca2+- and phosphorylation-dependent mechanisms in vascular smooth muscle. Here, we show that macroscopic currents from HEK 293 cells expressing murine BK channel α-subunits (mSlo) are acutely potentiated following α5β1 integrin activation. The effect occurs in a Ca2+-dependent manner, 1–3 min after integrin engagement. After integrin activation, normalized conductance-voltage relations for mSlo are left-shifted at free Ca2+ concentrations ≥1 μm. Overexpression of human c-src with mSlo, in the absence of integrin activation, leads to similar shifts in mSlo Ca2+ sensitivity, whereas overexpression of catalytically inactive c-src blocks integrin-induced potentiation. However, neither integrin activation nor c-src overexpression potentiates current in BK channels containing a point mutation at Tyr-766. Biochemical tests confirmed the critical importance of residue Tyr-766 in integrin-induced channel phosphorylation. Thus, BK channel activity is enhanced by α5β1 integrin activation, likely through an intracellular signaling pathway involving c-src phosphorylation of the channel α-subunit at Tyr-766. The net result is increased current amplitude, enhanced Ca2+ sensitivity, and rate of activation of the BK channel, which would collectively promote smooth muscle hyperpolarization in response to integrin-extracellular matrix interactions.