Activation of ERK1/2 by Store-Operated Calcium Entry in Rat Parotid Acinar Cells

Abstract

The regulation of intracellular Ca2+ concentration ([Ca2+]i) plays a critical role in a variety of cellular processes, including transcription, protein activation, vesicle trafficking, and ion movement across epithelial cells. In many cells, the activation of phospholipase C-coupled receptors hydrolyzes membrane phosphoinositides and produces the depletion of endoplasmic reticulum Ca2+ stores, followed by the sustained elevation of [Ca2+]i from Ca2+ entry across the plasma membrane via store-operated Ca2+ entry (SOCE). Ca2+ entry is also increased in a store-independent manner by arachidonate-regulated Ca2+ (ARC) channels. Using rat parotid salivary gland cells, we examined multiple pathways of Ca2+ entry/elevation to determine if they activated cell signaling proteins and whether this occurred in a pathway-dependent manner. We observed that SOCE activates extracellular signal-related kinases 1 and 2 (ERK1/2) to ∼3-times basal levels via a receptor-independent mechanism when SOCE was initiated by depleting Ca2+ stores using the endoplasmic reticulum Ca2+-ATPase inhibitor thapsigargin (TG). TG-initiated ERK1/2 phosphorylation increased as rapidly as that initiated by the muscarinic receptor agonist carbachol, which promoted an increase to ∼5-times basal levels. Notably, ERK1/2 phosphorylation was not increased by the global elevation of [Ca2+]i by Ca2+ ionophore or by Ca2+ entry via ARC channels in native cells, although ERK1/2 phosphorylation was increased by Ca2+ ionophore in Par-C10 and HSY salivary cell lines. Agents and conditions that blocked SOCE in native cells, including 2-aminoethyldiphenyl borate (2-APB), SKF96363, and removal of extracellular Ca2+, also reduced TG- and carbachol-stimulated ERK1/2 phosphorylation. TG-promoted ERK1/2 phosphorylation was blocked when SRC and Protein Kinases C (PKC) were inhibited, and it was blocked in cells pretreated with β-adrenergic agonist isoproterenol. These observations demonstrate that ERK1/2 is activated by a selective mechanism of Ca2+ entry (SOCE) in these cells, and suggest that ERK1/2 may contribute to events downstream of SOCE.