Store-operated Ca2+ Entry Is Suppressed During Mitosis Due To Phosphorylation Of The Endoplasmic Reticulum Ca2+ Sensor Stim1

Abstract

When ER Ca2+ stores are depleted due to physiological Ca2+ release or pharmacological perturbation, Ca2+ influx via plasma membrane Ca2+ channels is activated by a process known as store-operated Ca2+ entry (SOCE). The current associated with SOCE is Ca2+ release-activated Ca2+ current (Icrac). SOCE involves Orai Ca2+ influx channels and STIM ER Ca2+ sensors. When ER Ca2+ stores are full, STIM1 is localized throughout the ER membrane; however, ER Ca2+ store depletion induces rearrangement of STIM1 into punctate structures near the plasma membrane where it activates Orai channels. Interestingly, mitosis is the only known physiological situation in which Ca2+ store depletion is dissociated from SOCE or Icrac activation. Identification of the molecular components of the SOCE signaling pathway has facilitated analysis of the mechanism underlying mitotic SOCE suppression. We found that in mitotic HeLa cells, an enhanced yellow fluorescent protein-tagged STIM1 (eYFP-STIM1) did not rearrange into puncta in response to Ca2+ store depletion and accordingly, SOCE was not activated. We hypothesized that mitosis-specific phosphorylation of STIM1 may underlie the block of STIM1 rearrangement and SOCE suppression. To this end, the phospho-specific MPM-2 antibody recognized eYFP-STIM1 immunoprecipitated from mitotic but not interphase cells. MPM-2 recognizes phosphorylated serine or threonine followed by proline, and human STIM1 contains 10 instances of S/T-P, all located in the cytoplasmic, C-terminus. STIM1 truncation mutants indicate that at least 2 sites within the C-terminus account for the mitosis-specific phosphorylation. Individual phosphorylation site mutants are being created to identify specific phosphorylated residues and to determine the functional consequences of phosphorylation during mitosis. Suppression of SOCE during mitosis may be an important signaling event, because mitotic processes such as chromosome separation and cytokinesis are exquisitely sensitive to small changes in cytoplasmic Ca2+.