TY - JOUR
T1 - Cross-talk between N-terminal and C-terminal domains in stromal interaction molecule 2 (STIM2) determines enhanced STIM2 sensitivity
AU - Emrich, Scott M.
AU - Yoast, Ryan E.
AU - Xin, Ping
AU - Zhang, Xuexin
AU - Pathak, Trayambak
AU - Nwokonko, Robert
AU - Gueguinou, Maxime F.
AU - Subedi, Krishna P.
AU - Zhou, Yandong
AU - Ambudkar, Indu S.
AU - Hempel, Nadine
AU - Machaca, Khaled
AU - Gill, Donald L.
AU - Trebak, Mohamed
N1 - Publisher Copyright:
© 2019 American Society for Biochemistry and Molecular Biology Inc.. All rights reserved.
PY - 2019/4/19
Y1 - 2019/4/19
N2 - Store-operated Ca2+ entry (SOCE) is a ubiquitous pathway for Ca2+ influx across the plasma membrane (PM). SOCE is mediated by the endoplasmic reticulum (ER)-associated Ca2+-sensing proteins stromal interaction molecule 1 (STIM1) and STIM2, which transition intoan active conformation in response toERCa2+store depletion, thereby interacting with and gating PM-associated ORAI1 channels. Although structurally homologous, STIM1 and STIM2 generate distinct Ca2+ signatures in response to varying strengths of agonist stimulation. The physiological functions of these Ca2+ signatures, particularly under native conditions, remain unclear. To investigate the structural properties distinguishing STIM1 and STIM2 activation of ORAI1 channels under native conditions, here we used CRISPR/Cas9 to generate STIM1-/-, STIM2-/-, and STIM1/2-/- knockouts in HEK293 and colorectalHCT116cells.Weshow that depending on cell type, STIM2 can significantly sustain SOCE in response to maximal store depletion. Utilizing the SOCE modifier 2-aminoethoxydiphenyl borate (2-APB), we demonstrate that 2-APB-activated store-independent Ca2+ entry is mediated exclusively by endogenous STIM2. Using variants that either stabilize or disrupt intramolecular interactions of STIM C termini, we show that the increased flexibility of the STIM2 C terminus contributes to its selective store-independent activation by 2-APB. However,STIM1 variants with enhanced flexibility in the C terminus failed to supportitsstore- independentactivation.STIM1/STIM2chimericconstructs indicated that coordination between N-terminal sensitivity and C-terminal flexibility is required for specific store-independent STIM2 activation. Our results clarify the structural determinants underlying activation of specificSTIMisoforms, insights that are potentially useful for isoform-selective drug targeting.
AB - Store-operated Ca2+ entry (SOCE) is a ubiquitous pathway for Ca2+ influx across the plasma membrane (PM). SOCE is mediated by the endoplasmic reticulum (ER)-associated Ca2+-sensing proteins stromal interaction molecule 1 (STIM1) and STIM2, which transition intoan active conformation in response toERCa2+store depletion, thereby interacting with and gating PM-associated ORAI1 channels. Although structurally homologous, STIM1 and STIM2 generate distinct Ca2+ signatures in response to varying strengths of agonist stimulation. The physiological functions of these Ca2+ signatures, particularly under native conditions, remain unclear. To investigate the structural properties distinguishing STIM1 and STIM2 activation of ORAI1 channels under native conditions, here we used CRISPR/Cas9 to generate STIM1-/-, STIM2-/-, and STIM1/2-/- knockouts in HEK293 and colorectalHCT116cells.Weshow that depending on cell type, STIM2 can significantly sustain SOCE in response to maximal store depletion. Utilizing the SOCE modifier 2-aminoethoxydiphenyl borate (2-APB), we demonstrate that 2-APB-activated store-independent Ca2+ entry is mediated exclusively by endogenous STIM2. Using variants that either stabilize or disrupt intramolecular interactions of STIM C termini, we show that the increased flexibility of the STIM2 C terminus contributes to its selective store-independent activation by 2-APB. However,STIM1 variants with enhanced flexibility in the C terminus failed to supportitsstore- independentactivation.STIM1/STIM2chimericconstructs indicated that coordination between N-terminal sensitivity and C-terminal flexibility is required for specific store-independent STIM2 activation. Our results clarify the structural determinants underlying activation of specificSTIMisoforms, insights that are potentially useful for isoform-selective drug targeting.
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U2 - 10.1074/jbc.RA118.006801
DO - 10.1074/jbc.RA118.006801
M3 - Article
C2 - 30824535
AN - SCOPUS:85064865284
SN - 0021-9258
VL - 294
SP - 6318
EP - 6332
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 16
ER -