A key role for STIM1 in store operated calcium channel activation in airway smooth muscle

BACKGROUND: Control of cytosolic calcium plays a key role in airway myocyte function. Changes in intracellular Ca(2+ )stores can modulate contractile responses, modulate proliferation and regulate synthetic activity. Influx of Ca(2+ )in non excitable smooth muscle is believed to be predominantly through store operated channels (SOC) or receptor operated channels (ROC). Whereas agonists can activate both SOC and ROC in a range of smooth muscle types, the specific trigger for SOC activation is depletion of the sarcoplasmic reticulum Ca(2+ )stores. The mechanism underlying SOC activation following depletion of intracellular Ca(2+ )stores in smooth muscle has not been identified. METHODS: To investigate the roles of the STIM homologues in SOC activation in airway myocytes, specific siRNA sequences were utilised to target and selectively suppress both STIM1 and STIM2. Quantitative real time PCR was employed to assess the efficiency and the specificity of the siRNA mediated knockdown of mRNA. Activation of SOC was investigated by both whole cell patch clamp electrophysiology and a fluorescence based calcium assay. RESULTS: Transfection of 20 nM siRNA specific for STIM1 or 2 resulted in robust decreases (>70%) of the relevant mRNA. siRNA targeted at STIM1 resulted in a reduction of SOC associated Ca(2+ )influx in response to store depletion by cyclopiazonic acid (60%) or histamine but not bradykinin. siRNA to STIM2 had no effect on these responses. In addition STIM1 suppression resulted in a more or less complete abrogation of SOC associated inward currents assessed by whole cell patch clamp. CONCLUSION: Here we show that STIM1 acts as a key signal for SOC activation following intracellular Ca(2+ )store depletion or following agonist stimulation with histamine in human airway myocytes. These are the first data demonstrating a role for STIM1 in a physiologically relevant, non-transformed endogenous expression cell model.