CRACM1, CRACM2, and CRACM3 Are Store-Operated Ca2+ Channels with Distinct Functional Properties

Abstract

STIM1 in the endoplasmic reticulum and CRACM1 in the plasma membrane are essential molecular components for controlling the store-operated CRAC current. CRACM1 proteins multimerize and bind STIM1, and the combined overexpression of STIM1 and CRACM1 reconstitutes amplified CRAC currents. Mutations in CRACM1 determine the selectivity of CRAC currents, demonstrating that CRACM1 forms the CRAC channel's ion-selective pore, but the CRACM1 homologs CRACM2 and CRACM3 are less well characterized. Here, we show that both CRACM2 and CRACM3, when overexpressed in HEK293 cells stably expressing STIM1, potentiate I(CRAC) to current amplitudes 15-20 times larger than native I(CRAC). A nonconducting mutation of CRACM1 (E106Q) acts as a dominant negative for all three CRACM homologs, suggesting that they can form heteromultimeric channel complexes. All three CRACM homologs exhibit distinct properties in terms of selectivity for Ca(2+) and Na(+), differential pharmacological effects in response to 2-APB, and strikingly different feedback regulation by intracellular Ca(2+). Each of the CRAC channel proteins' specific functional features and the potential heteromerization provide for flexibility in shaping Ca(2+) signals, and their characteristic biophysical and pharmacological properties will aid in identifying CRAC-channel species in native cells that express them.

ORAI1-Mediated Calcium Influx in Lactation and in Breast Cancer

“…The ORAI isoforms differ in their calcium selectivity, conductance, and activation kinetics (42,43). TRPC1 is also implicated in store-operated calcium entry (25,26), in some cases with ORAI1 (44, 45), so we assessed its potential role in lactation.…”

Plasticity in Ca 2+ selectivity of Orai1/Orai3 heteromeric channel

“…The capability of heteromeric Orai assembly has been demonstrated biochemically (9,21,22). We initially evaluated the potential of Orai1 and Orai3 heteromerization by employing fluorescence resonance energy transfer (FRET) microscopy.…”

Store-Operated Calcium Channels: New Perspectives on Mechanism and Function

“…STIM1 was quickly identified as a Ca 2þ sensor for SOCE because it displayed an EF-handlike sequence on the predicted luminal side, and neutralization of acidic residues to reduce Ca 2þ binding by the EF hand elicited constitutive Ca 2þ entry independent of store depletion, in effect mimicking the store-depleted state (Liou et al 2005;Zhang et al 2005). Of the three Orai isoforms in mammals, Orai1 was most closely connected to the CRAC channel because a point mutation (R91W) in Orai1 led to a complete loss of I CRAC in human T cells (Feske et al 2005, Orai1 is the predominant isoform in these cells, and its properties (fast Ca 2þ -dependent inactivation, sensitivity to 2-aminoethyldiphenyl borate (2-APB), and predominant expression in human T cells) most closely match those of CRAC (Lis et al 2007). Orai1 was shown to be the pore-forming subunit of the CRAC channel by changes in I CRAC calcium selectivity caused by mutating highly conserved glutamates ( particularly E106D) in the trans-membrane domains Vig et al 2006a;Yeromin et al 2006).…”