A genetically encoded ratiometric calcium sensor enables quantitative measurement of the local calcium microdomain in the endoplasmic reticulum

作者：Chen Luo, Huiyu Wang, Qi Liu, Wenting He, Lin Yuan, Pingyong Xu · 发表于：Biophysics Reports · 年份：2019 · DOI：10.1007/s41048-019-0082-6 · 被引用次数：26 · 研究领域：Ion Channels and Receptors、Calcium signaling and nucleotide metabolism、Cellular transport and secretion

The local Ca 2+ release from the heterogeneously distributed endoplasmic reticulum (ER) calcium store has a critical role in calcium homeostasis and cellular function. However, single fluorescent protein-based ER calcium probes experience challenges in quantifying the ER calcium store in differing live cells, and intensity-based measurements make it difficult to detect local calcium microdomains in the ER. Here, we developed a genetically encoded ratiometric ER calcium indicator (GCEPIA1-SNAP ER ) that can detect the real-time ER calcium store and local calcium microdomains in live cells. GCEPIA1-SNAP ER was located in the lumen of the ER and showed a linear, reversible and rapid response to changes in the ER calcium store. The GCEPIA1-SNAP ER probe effectively monitored the depletion of the ER calcium store by TG or starvation treatment, and through its use we identified heterogeneously distributed calcium microdomains in the ER which were correlated with the distribution of STIM1 clusters upon ER calcium store depletion. Lastly, GCEPIA1-SNAP ER can be used to detect the ER calcium store by high-throughput flow cytometry and confers the ability to study the function of calcium microdomains of the ER.