Dynamic Assembly of a Membrane Signaling Complex Enables Selective Activation of NFAT by Orai1

Abstract

SummaryNFAT-dependent gene expression is essential for the development and function of the nervous, immune, and cardiovascular systems and kidney, bone, and skeletal muscle [1]. Most NFAT protein resides in the cytoplasm because of extensive phosphorylation, which masks a nuclear localization sequence. Dephosphorylation by the Ca2+-calmodulin-activated protein phosphatase calcineurin triggers NFAT migration into the nucleus [2, 3]. In some cell types, NFAT can be activated by Ca2+ nanodomains near open store-operated Orai1 and voltage-gated Ca2+ channels in the plasma membrane [4, 5]. How local Ca2+ near Orai1 is detected and whether other Orai channels utilize a similar mechanism remain unclear. Here, we report that the paralog Orai3 fails to activate NFAT. Orai1 is effective in activating gene expression via Ca2+ nanodomains because it participates in a membrane-delimited signaling complex that forms after store depletion and brings calcineurin, via the scaffolding protein AKAP79, to calmodulin tethered to Orai1. By contrast, Orai3 interacts less well with AKAP79 after store depletion, rendering it ineffective in activating NFAT. A channel chimera of Orai3 with the N terminus of Orai1 was able to couple local Ca2+ entry to NFAT activation, identifying the N-terminal domain of Orai1 as central to Ca2+ nanodomain-transcription coupling. The formation of a store-dependent signaling complex at the plasma membrane provides for selective activation of a fundamental downstream response by Orai1.

Distinct Structural Domains of Caveolin-1 Independently Regulate Ca2+ Release-Activated Ca2+ Channels and Ca2+ Microdomain-Dependent Gene Expression

“…Although this is a general concern with overexpression studies, we do not think a gross displacement of endogenous scaffolding proteins has occurred under our conditions. CRAC channels activate the NFAT pathway in RBL-1 cells through recruitment of the scaffolding protein AKAP79 (12). Despite expression of recombinant caveolin-1, the NFAT pathway remained functional.…”

“…Because the same trigger of local Ca 2ϩ through CRAC channels activates both transcription factors in RBL cells (5-7), we reasoned that mechanisms must exist to tunnel the Ca 2ϩ microdomain down one pathway and not the other. Ca 2ϩ microdomains activate NFAT and c-fos indirectly, through recruitment of AKAP79 and calcineurin (12) and of Syk and Stat5 (6), respectively. We therefore posited that a large plasma membrane scaffolding protein that interacts with numerous signaling molecules might determine which transcription factor is activated by the local Ca 2ϩ signal.…”

Calcium signaling and cell proliferation

“…When normal epithlelial and breast cancer tissue is compared, down-regulation of Orai3 It was demonstrated that Orai1, but not its paralog Orai3, can activate NFAT [97]. In this case, temporal and spatial regulation mechanisms of these proteins are crucial for activation or recruitment of proteins to the PM.…”

“…In this case, temporal and spatial regulation mechanisms of these proteins are crucial for activation or recruitment of proteins to the PM. Orai1 participates in a membrane signaling complex involving calcineurin, which binds through CaM tethering to Orai1 by AKAP79 protein [97]. This complex= is activated after Ca 2+ store depletion and then activates gene expression via Ca 2+ microdomains [97].…”

A STIM1-dependent ‘trafficking trap’ mechanism regulates Orai1 plasma membrane residence and Ca2+ influx levels

“…5D,J), and importantly show that the translocation of intracellular Orai1 to the plasma membrane following store depletion is functionally coupled to increased levels of Ca 2+ influx through SOCE. (Courjaret and Machaca, 2014;Dolmetsch et al, 1997;Kar et al, 2012Kar et al, , 2014Parekh, 2009). SOCE levels depend on the stoichiometry of STIM1 and Orai1 in plasma membrane clusters (Hoover and Lewis, 2011), and as such on the levels of Orai1 at the plasma membrane.…”