Alpha5beta1 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 alpha5beta1 integrin ligand, enhances BK channel current through both Ca(2+)- and phosphorylation-dependent mechanisms in vascular smooth muscle. Here, we show that macroscopic currents from HEK 293 cells expressing murine BK channel alpha-subunits (mSlo) are acutely potentiated following alpha5beta1 integrin activation. The effect occurs in a Ca(2+)-dependent manner, 1-3 min after integrin engagement. After integrin activation, normalized conductance-voltage relations for mSlo are left-shifted at free Ca(2+) concentrations >or=1 microm. Overexpression of human c-src with mSlo, in the absence of integrin activation, leads to similar shifts in mSlo Ca(2+) 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 alpha5beta1 integrin activation, likely through an intracellular signaling pathway involving c-src phosphorylation of the channel alpha-subunit at Tyr-766. The net result is increased current amplitude, enhanced Ca(2+) sensitivity, and rate of activation of the BK channel, which would collectively promote smooth muscle hyperpolarization in response to integrin-extracellular matrix interactions.
Yang
Yan
Y
Department of Medical Pharmacology and Physiology, University of Missouri School of Medicine, Columbia, Missouri 65212, USA.
Wu
Xin
X
Gui
Peichun
P
Wu
Jianbo
J
Sheng
Jian-Zhong
JZ
Ling
Shizhang
S
Braun
Andrew P
AP
Davis
George E
GE
Davis
Michael J
MJ
D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D002460:Cell Line; D004789:Enzyme Activation / Q000187:drug effects; D005353:Fibronectins / Q000494:pharmacology; D006801:Humans; D039081:Integrin alpha5beta1 / Q000378:metabolism*; D015640:Ion Channel Gating / Q000187:drug effects*; D051037:Large-Conductance Calcium-Activated Potassium Channel alpha Subunits / Q000737:chemistry / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D010766:Phosphorylation / Q000187:drug effects; D019000:Phosphotyrosine / Q000378:metabolism; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism*; D011743:Pyrimidines / Q000494:pharmacology
C412373:AG 1879; D005353:Fibronectins; D039081:Integrin alpha5beta1; C465449:Kcnma1 protein, mouse; D051037:Large-Conductance Calcium-Activated Potassium Channel alpha Subunits; D011743:Pyrimidines; D019000:Phosphotyrosine; D016392:Proto-Oncogene Proteins pp60(c-src); D002118:Calcium