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Mediated through the combined action of STIM proteins and Orai channels, store-operated Ca2+ entry (SOCE) functions ubiquitously among different cell types. The existence of multiple STIM and Orai genes has made it difficult to assign specific roles of each STIM and Orai homolog in mediating Ca2+ signals. Using CRISPR/Cas9 gene editing tools, we generated cells with both STIM or all three Orai homologs deleted and directly monitored store Ca2+ and Ca2+ signals. We found that unstimulated, SOCE null KO cells still retain 50~70% of ER Ca2+ stores of wildtype (wt) cells. After brief exposure to store-emptying conditions, acute refilling of ER Ca2+ stores was totally blocked in KO cells. However, after 24 h in culture, stores were eventually refilled. Thus, SOCE is critical for immediate refilling of ER Ca2+ but is dispensable for the maintenance of long-term ER Ca2+ homeostasis. Using the Orai null background triple Orai-KO cells, we examined the plasma membrane translocation properties of a series of truncated STIM1 variants. FRET analysis reveals that, even though PM tethering of STIM1 expedites the activation of STIM1 by facilitating its oligomerization, migration, and accumulation in ER-PM junctions, it is not required for the conformational switch, oligomerization, and clustering of STIM1. Even without overt puncta formation at ER-PM junctions, STIM11–491 and STIM11–666 could still rescue SOCE when expressed in STIM KO cells. Thus, ER-PM trapping and clustering of STIM molecules only facilitates the process of SOCE activation, but is not essential for the activation of Orai channels.Electronic supplementary materialThe online version of this article (10.1007/s00424-018-2165-5) contains supplementary material, which is available to authorized users.

We report, in this letter, a simple thermal oxidation approach to growing large-scale arrays of aligned tungsten oxide nanorods on planar substrates. By passing a current of ∼50 A through a tungsten spiral coil in a vacuum of ∼3 × 10−2 Torr, single-crystalline WO2.9 nanorods, ∼20 nm in diameter and ∼400 nm long, were deposited on planar substrates, e.g. Si(001), Si(111), SiO2/Si, glass, and Ag-coated silicon substrates at temperatures below 200°C. The nanorods were vertically aligned on the substrates. The structure, optical properties and alignment of the nanorods was investigated by x-ray diffraction and transmission electron microscopes, micro-Raman spectrometer, and scanning electron microscope and x-ray pole figure measurement, respectively. The present study provides an easy way to grow large-scale arrays of aligned tungsten oxide nanorods at relatively low temperatures, which might also be applicable to the growth of other metal oxides.