Ong Hwei Ling, Liu Xibao, Tsaneva-Atanasova Krasimira, Singh Brij B, Bandyopadhyay Bidhan C, Swaim William D, Russell James T, Hegde Ramanujan S, Sherman Arthur, Ambudkar Indu S
Secretory Physiology Section, Gene Therapy and Therapeutics Branch, NIDCR, National Institutes of Health, Bethesda, Maryland 20892, USA.
J Biol Chem. 2007 Apr 20;282(16):12176-85. doi: 10.1074/jbc.M609435200. Epub 2007 Feb 12.
STIM1 (stromal interacting molecule 1), an endoplasmic reticulum (ER) protein that controls store-operated Ca(2+) entry (SOCE), redistributes into punctae at the cell periphery after store depletion. This redistribution is suggested to have a causal role in activation of SOCE. However, whether peripheral STIM1 punctae that are involved in regulation of SOCE are determined by depletion of peripheral or more internal ER has not yet been demonstrated. Here we show that Ca(2+) depletion in subplasma membrane ER is sufficient for peripheral redistribution of STIM1 and activation of SOCE. 1 microM thapsigargin (Tg) induced substantial depletion of intracellular Ca(2+) stores and rapidly activated SOCE. In comparison, 1 nM Tg induced slower, about 60-70% less Ca(2+) depletion but similar SOCE. SOCE was confirmed by measuring I(SOC) in addition to Ca(2+), Mn(2+), and Ba(2+) entry. Importantly, 1 nM Tg caused redistribution of STIM1 only in the ER-plasma membrane junction, whereas 1 microM Tg caused a relatively global relocalization of STIM1 in the cell. During the time taken for STIM1 relocalization and SOCE activation, 1 nM Bodipy-fluorescein Tg primarily labeled the subplasma membrane region, whereas 1 microM Tg labeled the entire cell. The localization of Tg in the subplasma membrane region was associated with depletion of ER in this region and activation of SOCE. Together, these data suggest that peripheral STIM1 relocalization that is causal in regulation of SOCE is determined by the status of [Ca(2+)] in the ER in close proximity to the plasma membrane. Thus, the mechanism involved in regulation of SOCE is contained within the ER-plasma membrane junctional region.
基质相互作用分子1（STIM1）是一种内质网（ER）蛋白，可控制钙库操纵的Ca²⁺内流（SOCE），在钙库耗竭后重新分布到细胞周边的点状结构中。这种重新分布被认为在SOCE的激活中起因果作用。然而，参与SOCE调节的外周STIM1点状结构是由外周内质网还是更多内部内质网的耗竭所决定，尚未得到证实。在这里，我们表明质膜下内质网中的Ca²⁺耗竭足以使STIM1在外周重新分布并激活SOCE。1微摩尔毒胡萝卜素（Tg）可诱导细胞内Ca²⁺储存大量耗竭并迅速激活SOCE。相比之下，1纳摩尔Tg诱导的Ca²⁺耗竭较慢，减少约60 - 70%，但SOCE相似。除了Ca²⁺、Mn²⁺和Ba²⁺内流外，通过测量I(SOC)证实了SOCE。重要的是，1纳摩尔Tg仅导致STIM1在内质网 - 质膜连接处重新分布，而1微摩尔Tg导致STIM1在细胞内相对整体重新定位。在STIM1重新定位和SOCE激活所需的时间内，1纳摩尔Bodipy - 荧光素Tg主要标记质膜下区域，而1微摩尔Tg标记整个细胞。Tg在质膜下区域的定位与该区域内质网的耗竭和SOCE的激活相关。总之，这些数据表明在SOCE调节中起因果作用的外周STIM1重新定位是由紧邻质膜的内质网中[Ca²⁺]的状态决定的。因此，SOCE调节所涉及的机制包含在内质网 - 质膜连接区域内。
