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  • A novel chimeric aequorin fused with caveolin-1 reveals a sphingosine kinase 1-regulated Ca²⁺ microdomain in the caveolar compartment.

A novel chimeric aequorin fused with caveolin-1 reveals a sphingosine kinase 1-regulated Ca²⁺ microdomain in the caveolar compartment.

Biochimica et biophysica acta (2015-04-22)
Ilari Pulli, Tomas Blom, Christoffer Löf, Melissa Magnusson, Alessandro Rimessi, Paolo Pinton, Kid Törnquist
ABSTRACT

Caveolae are plasma membrane invaginations enriched in sterols and sphingolipids. Sphingosine kinase 1 (SK1) is an oncogenic protein that converts sphingosine to sphingosine 1-phosphate (S1P), which is a messenger molecule involved in calcium signaling. Caveolae contain calcium responsive proteins, but the effects of SK1 or S1P on caveolar calcium signaling have not been investigated. We generated a Caveolin-1-Aequorin fusion protein (Cav1-Aeq) that can be employed for monitoring the local calcium concentration at the caveolae ([Ca²⁺]cav). In HeLa cells, Cav1-Aeq reported different [Ca²⁺] as compared to the plasma membrane [Ca²⁺] in general (reported by SNAP25-Aeq) or as compared to the cytosolic [Ca²⁺] (reported by cyt-Aeq). The Ca²⁺ signals detected by Cav1-Aeq were significantly attenuated when the caveolar structures were disrupted by methyl-β-cyclodextrin, suggesting that the caveolae are specific targets for Ca²⁺ signaling. HeLa cells overexpressing SK1 showed increased [Ca²⁺]cav during histamine-induced Ca²⁺ mobilization in the absence of extracellular Ca²⁺ as well as during receptor-operated Ca²⁺ entry (ROCE). The SK1-induced increase in [Ca²⁺]cav during ROCE was reverted by S1P receptor antagonists. In accordance, pharmacologic inhibition of SK1 reduced the [Ca²⁺]cav during ROCE. S1P treatment stimulated the [Ca²⁺]cav upon ROCE. The Ca²⁺ responses at the plasma membrane in general were not affected by SK1 expression. In summary, our results show that SK1/S1P-signaling regulates Ca²⁺ signals at the caveolae. This article is part of a Special Issue entitled: 13th European Symposium on Calcium.

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Product Description

Sigma-Aldrich
L-Glutamine, meets USP testing specifications, suitable for cell culture, 99.0-101.0%, from non-animal source
Sigma-Aldrich
L-Glutamine