Transcription Factor NF-κB Regulates Expression of Pore-forming Ca2+ Channel Unit, Orai1, and Its Activator, STIM1, to Control Ca2+ Entry and Affect Cellular Functions

Abstract

Alterations of cytosolic Ca 2ϩ activity participate in the regulation of a wide variety of cellular functions including excitation-contraction coupling, exocytosis, migration, cell proliferation, and cell death (1-4). Cytosolic Ca 2ϩ is increased by release of Ca 2ϩ from intracellular stores and/or Ca 2ϩ entry across the cell membrane (5). Ca 2ϩ release from intracellular stores results in the stimulation of Ca 2ϩ release-activated Ca 2ϩ channel (CRAC) 2 (6, 7), which consists of the pore forming units Orai1, -2, and/or -3 (8 -10) and the endoplasmic reticulum-located regulatory subunit STIM1 or -2 (11-13). The stimulation of the channel leads to the inward current I CRAC and the store-operated Ca 2ϩ entry (SOCE). Recent observations uncovered the powerful stimulation of I CRAC and SOCE by the serum and glucocorticoid-inducible kinase SGK1 (14), a kinase stimulated by growth factors and involved in stress response (15) and regulation of cell survival (16). SGK1 is partially effective through phosphorylation of the ubiquitin ligase Nedd4-2 (neuronal precursor cells expressed developmentally down-regulated). Nedd4-2 ubiquitinates Orai1, thus preparing the channel protein for degradation (14). The effect of Nedd4-2 on Orai1 parallels that of Nedd4-2 on the epithelial Na ϩ channel ENaC (16, 17). The phosphorylation of Nedd4-2 leads to binding of the ubiquitin ligase to the protein 14-3-3, which prevents the interaction with the channel protein (18). Accordingly, SGK1 enhances Orai1 protein abundance in the cell membrane (14). STIM is similarly regulated by ubiquitination (19). However, the effect of SGK1 on Orai1 protein abundance is only in part explained by Nedd4-2-dependent protein degradation. Therefore, further experiments were performed to explore whether SGK1, in addition, stimulates Orai1 and/or STIM1 expression. As a matter of fact, RT-PCR revealed an increase of Orai1 and STIM1 transcript levels after expression of constitutively active SGK1. Thus, further experiments were performed to uncover the transcription factor involved. Previously, SGK1 has been shown to foster nuclear translocation and activation of nuclear factor B (NF-B) (20 -22). Accordingly, this study explored the putative involvement of NF-B subunits p65 (RELA), p50 (NFKB1), and p52 (NFKB2) in the regulation of Orai1 and STIM1 expression. EXPERIMENTAL PROCEDURES

Impairment of hepatic nuclear factor-4α binding to theStim1promoter contributes to high glucose-induced upregulation of STIM1 expression in glomerular mesangial cells

“…Because HNF4␣ expression is tissue specific (11,23,41), our results suggest that transcription factor-regulated STIM1 expression might be cell context dependent. Indeed, we failed to observe an NF-B effect on STIM1 expression in MCs (data not shown), as previously described in mast cell and pulmonary vascular endothelial cells (7,12). HNF4␣ is generally thought as a transcriptional activator and is known to activate a wide variety of genes involved in glucose, fatty acid, cholesterol, and amino acid metabolism (18,20,31,45).…”

“…Little is known about the regulation of the abundance of STIM1, which could also affect channel function. In this regard, the transcription factor NF-B has been recently reported to promote STIM1 protein expression in mast cells and in vascular endothelial cells (7,12). In the present study, we showed that HNF4␣, a nuclear transcription factor, regulated STIM1 expression by functioning as a repressor in MCs.…”

Cooperative Signaling via Transcription Factors NF-κB and AP1/c-Fos Mediates Endothelial Cell STIM1 Expression and Hyperpermeability in Response to Endotoxin

“…It has been shown recently that the serum and glucocorticoid-inducible kinase SGK1 controls constitutive STIM1 transcription via NF-B signaling in bone marrow-derived mast cells (50). However, the transcriptional mechanism of STIM1 expression in response to inflammatory stimuli in ECs was not studied.…”

Serum- and glucocorticoid-inducible kinase SGK1 regulates reorganization of actin cytoskeleton in mast cells upon degranulation

“…The process of F-actin disassembly was shown to be dependent on the increase of intracellular Ca 2ϩ (16,23). On the other hand, a powerful regulator of Ca 2ϩ entry and exocytosis in MCs is the serum-and glucocorticoidinducible kinase SGK1 (9,10,33). SGK1-deficient (sgk1 Ϫ/Ϫ ) MCs have a decreased FcεRI-dependent Ca 2ϩ entry and degranulation, and sgk1 Ϫ/Ϫ mice are thus protected against anaphylactic reaction, pointing to a strongly impaired MC function in vivo (33).…”

“…SGK1 is partially effective through phosphorylation of the ubiquitin ligase Nedd4-2 (neuronal precursor cells expressed developmentally downregulated), which ubiquitinates Orai1, thus preparing the channel protein for degradation (9). Moreover, SGK1 activates the transcription factor NF-B, which is required for Orai1 and STIM1 transcription (10). However, although regulation of Ca 2ϩ -dependent actin cytoskeleton dynamics is a crucial step for MC degranulation, nothing is known about the role of SGK1 in the process of actin restructuring.…”