Abstract
Serum and glucocorticoid-regulated kinase 1 (SGK1) encodes a phosphatidylinositol 3-kinase-dependent serine/threonine kinase that is rapidly induced in response to cellular stressors and is an important cell survival signal. Previous studies have suggested that an increase in cytoplasmic Ca(2+) concentration ([Ca(2+)]c) is required for increased SGK1 expression, but the subcellular source of Ca(2+) regulating SGK1 transcription remains uncertain. Activation of endoplasmic reticulum stress (ERS) with thapsigargin (TG) increased SGK1 mRNA and protein expression in MDA-MB-231 cells. Intracellular Ca(2+) imaging revealed that store-operated Ca(2+) entry played a prominent role in SGK1 induction by TG. Neither ERS nor release of Ca(2+) from the ER was sufficient to activate SGK1. Prolonged elevation of intracellular Ca(2+) levels, however, triggered cell death with a much greater proportion of the cells undergoing necrosis rather than apoptosis. A relative increase in the percentage of cells undergoing necrosis was observed in cells expressing a short hairpin RNA targeted to the SGK1 gene. Necrotic cell death evoked by cytoplasmic Ca(2+) overloading was associated with persistent hyperpolarization of the inner mitochondrial membrane and a modest increase in calpain activation, but did not involve detectable caspase 3 or caspase 7 activation. The effects of cytoplasmic Ca(2+) overloading on mitochondrial membrane potential were significantly reduced in cells expressing SGK1 compared with SGK1-depleted cells. Our findings indicate that store-operated Ca(2+) entry regulates SGK1 expression in epithelial cells and suggest that SGK1-dependent cytoprotective signaling involves effects on maintaining mitochondrial function.
Highlights
Mechanisms underlying SGK1 activation are incompletely understood in epithelial cells
Our results suggest that store-operated Ca2ϩ entry and subsequent calmodulin-dependent protein kinase II (CaMKII) activity regulate SGK1 induction in MDA-MB-231 cells
Induction of endoplasmic reticulum stress (ERS) in MDA-MB-231 cells was confirmed by measuring for the presence the GRP78 (78-kDa glucose-regulated protein), known as immunoglobulin heavychain binding protein (BiP), a member of the 70-kDa heat shock protein family and a protein chaperone within the ER lumen
Summary
Mechanisms underlying SGK1 activation are incompletely understood in epithelial cells. Store-operated Ca2؉ entry regulates SGK1 expression in epithelial cells and suggest that SGK1-dependent cytoprotective signaling involves effects on maintaining mitochondrial function. The activation of SGK1 by insulin was inhibited by BAPTA and calmodulin antagonists [16] These studies indicate an important role of Ca2ϩ signaling pathways in SGK1 gene expression and kinase activity. Our results suggest that store-operated Ca2ϩ entry and subsequent calmodulin-dependent protein kinase II (CaMKII) activity regulate SGK1 induction in MDA-MB-231 cells. We discovered that necrotic cell death produced by a sustained increase in intracellular Ca2ϩ was attenuated by SGK1 prosurvival signals associated with maintenance of the mitochondrial membrane potential
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