Orai Coordinates Multiple Regulatory Processes via its N-Terminal Conserved Domain

Abstract

Among store-operated Ca2+ entry (SOCE) pathways the Ca2+ release-activated Ca2+ (CRAC) channel is best characterized. STIM1 and Orai1 have been identified as the molecular key components of the CRAC channel. STIM1 acts as a Ca2+ sensor in the ER. Upon store-depletion STIM1 proteins oligomerize and translocate close to the plasma membrane. Thereby Orai proteins, representing pore-forming subunits in the plasma membrane, are activated by coupling to STIM1 multimers. Here we focused especially on the role of Orai N-termini. It is noticeable that this cytosolic strand includes an among all Orai proteins conserved region close to the first TM domain, which contains positively charged as well as hydrophobic amino acids. Up to now the role of these residues has not been clarified. Moreover this N-terminal conserved region functions not only as a binding partner for STIM1, but also for CaM and CRACR2A. The series of positively charged amino acids further resembles a PIP2 binding domain. With respect to these overlapping regions, we examined the role of positively charged and hydrophobic amino acids by generating point mutants. Single point mutations of positively charged amino acids reduced fast inactivation, possibly due to impairment of CaM binding. Replacement of hydrophobic amino acids did not affect fast inactivation excluding the disturbance of CaM binding. However, store-operated currents of these hydrophobic amino acid mutants were significantly increased, suggesting a possible connection with lipid-mediated processes. Mag. Lackner Barbara is a Recipient of a DOC/DOC-fFORTE-fellowship of the Austrian Academy of Sciences at the Institute of Biophysics at the Johannes-Kepler University of Linz.