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Role of the microtubule cytoskeleton in the function of the store-operated Ca2+ channel activator STIM1.

ABSTRACT: We examined the role of the microtubule cytoskeleton in the localization and store-operated Ca(2+) entry (SOCE) function of the endoplasmic reticulum (ER) Ca(2+) sensor stromal interaction molecule 1 (STIM1) in HEK 293 cells. STIM1 tagged with an enhanced yellow fluorescent protein (EYFP-STIM1) exhibited a fibrillar localization that colocalized with endogenous alpha-tubulin. Depolymerization of microtubules with nocodazole caused a change from a fibrillar EYFP-STIM1 localization to one that was similar to that of the ER. Treatment of HEK 293 cells with nocodazole had a detrimental impact on SOCE and the associated Ca(2+) release-activated Ca(2+) current (I(CRAC)). This inhibition was significantly reversed in cells overexpressing EYFP-STIM1, implying that the primary inhibitory effect of nocodazole is related to STIM1 function. Surprisingly, nocodazole treatment alone induced significant SOCE and I(CRAC) in cells expressing EYFP-STIM1, and this was accompanied by an increase in EYFP-STIM1 fluorescence near the plasma membrane. We conclude that microtubules play a facilitative role in the SOCE signaling pathway by optimizing the localization of STIM1.

SUBMITTER: Smyth JT

PROVIDER: S-EPMC2440341 | biostudies-literature | 2007 Nov

REPOSITORIES: biostudies-literature

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Role of the microtubule cytoskeleton in the function of the store-operated Ca2+ channel activator STIM1.

Smyth Jeremy T JT DeHaven Wayne I WI Bird Gary S GS Putney James W JW

Journal of cell science 20071009 Pt 21

We examined the role of the microtubule cytoskeleton in the localization and store-operated Ca(2+) entry (SOCE) function of the endoplasmic reticulum (ER) Ca(2+) sensor stromal interaction molecule 1 (STIM1) in HEK 293 cells. STIM1 tagged with an enhanced yellow fluorescent protein (EYFP-STIM1) exhibited a fibrillar localization that colocalized with endogenous alpha-tubulin. Depolymerization of microtubules with nocodazole caused a change from a fibrillar EYFP-STIM1 localization to one that was ...[more]

PMID: 17925382

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Project description:1,4,5-trisphosphate (IP(3))-dependent Ca(2+) signaling regulates gonad function, fertility, and rhythmic posterior body wall muscle contraction (pBoc) required for defecation in Caenorhabditis elegans. Store-operated Ca(2+) entry (SOCE) is activated during endoplasmic reticulum (ER) Ca(2+) store depletion and is believed to be an essential and ubiquitous component of Ca(2+) signaling pathways. SOCE is thought to function to refill Ca(2+) stores and modulate Ca(2+) signals. Recently, stromal interaction molecule 1 (STIM1) was identified as a putative ER Ca(2+) sensor that regulates SOCE. We cloned a full-length C. elegans stim-1 cDNA that encodes a 530-amino acid protein with approximately 21% sequence identity to human STIM1. Green fluorescent protein (GFP)-tagged STIM-1 is expressed in the intestine, gonad sheath cells, and spermatheca. Knockdown of stim-1 expression by RNA interference (RNAi) causes sterility due to loss of sheath cell and spermatheca contractile activity required for ovulation. Transgenic worms expressing a STIM-1 EF-hand mutant that constitutively activates SOCE in Drosophila and mammalian cells are sterile and exhibit severe pBoc arrhythmia. stim-1 RNAi dramatically reduces STIM-1GFP expression, suppresses the EF-hand mutation-induced pBoc arrhythmia, and inhibits intestinal store-operated Ca(2+) (SOC) channels. However, stim-1 RNAi surprisingly has no effect on pBoc rhythm, which is controlled by intestinal oscillatory Ca(2+) signaling, in wild type and IP(3) signaling mutant worms, and has no effect on intestinal Ca(2+) oscillations and waves. Depletion of intestinal Ca(2+) stores by RNAi knockdown of the ER Ca(2+) pump triggers the ER unfolded protein response (UPR). In contrast, stim-1 RNAi fails to induce the UPR. Our studies provide the first detailed characterization of STIM-1 function in an intact animal and suggest that SOCE is not essential for certain oscillatory Ca(2+) signaling processes and for maintenance of store Ca(2+) levels in C. elegans. These findings raise interesting and important questions regarding the function of SOCE and SOC channels under normal and pathophysiological conditions.

Dataset Information

Role of the microtubule cytoskeleton in the function of the store-operated Ca2+ channel activator STIM1.

Publications

Role of the microtubule cytoskeleton in the function of the store-operated Ca2+ channel activator STIM1.

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