New Modes of GPCR Signalling

ABSTRACT
Sequential Activation of Orai1 Subunits by Different Numbers of STIM1
Leads to Multistate Opening of the CRAC Channel
Zhengzheng Li 1, Lin Liu 2, Yongqiang Deng 1, Wei Ji 1, Wen Du 1, Pingyong Xu 1 Liangyi
Chen 1, and Tao Xu 1,2
1National Key Laboratory of Biomacromolecules, Institute of Biophysics, Chinese
Academy of Sciences, Beijing 100101, P. R. China.
2College of Life Science and Technology, Huazhong University of Science and
Technology, Wuhan 430074, P. R. China.
STIM1 and Orai1 are essential proteins of the Ca 2+ -release-activated Ca 2+ (CRAC)
channel which mediates store-operated Ca 2+ entry. STIM1 is localized at the
endoplasmic reticulum (ER) and functions as the sensor of ER luminal Ca 2+
concentration, and Orai1 is the channel pore subunit at the plasma membrane. STIM1
aggregates underneath the plasma membrane after store depletion and activates the
CRAC channel by directly binding to Orai1. It remains a challenge to determine how
many STIM1 molecules are required to open a CRAC channel. Here, by linking
different numbers of Orai1 subunits with functional cytoplasmic domains of STIM1
(residues 336-485, designated S domains), we provide evidence that each CRAC
channel can be fully activated by eight S domains. Further experiments revealed that
two S domains can fully activate one Orai1 subunit and the CRAC channel can
progressively open as more Orai1 subunits are activated. Our data suggest that full
activation of one CRAC channel complex requires eight STIM1 molecules, and the
activation of the channel is not an “all-or-none” process but undergoes multiple
intermediate states corresponding to different numbers of Orai1 subunits being
activated.