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- Title
Na /Ca exchangers and Orai channels jointly refill endoplasmic reticulum (ER) Ca via ER nanojunctions in vascular endothelial cells.
- Authors
Di Giuro, Cristiana; Shrestha, Niroj; Malli, Roland; Groschner, Klaus; Breemen, Cornelis; Fameli, Nicola
- Abstract
We investigated the role of Na/ Ca exchange (NCX) in the refilling of endoplasmic reticulum (ER) Ca in vascular endothelial cells under various conditions of cell stimulation and plasma membrane (PM) polarization. Better understanding of the mechanisms behind basic ER Ca content regulation is important, since current hypotheses on the possible ultimate causes of ER stress point to deterioration of the Ca transport mechanism to/from ER itself. We measured [Ca] temporal changes by Fura-2 fluorescence under experimental protocols that inhibit a host of transporters (NCX, Orai, non-selective transient receptor potential canonical (TRPC) channels, sarco/endoplasmic reticulum Ca ATPase (SERCA), Na/ K ATPase (NKA)) involved in the Ca communication between the extracellular space and the ER. Following histamine-stimulated ER Ca release, blockade of NCX Ca-influx mode (by 10 μM KB-R7943) diminished the ER refilling capacity by about 40%, while in Orai1 dominant negative-transfected cells NCX blockade attenuated ER refilling by about 60%. Conversely, inhibiting the ouabain sensitive NKA (10 nM ouabain), which may be localized in PM-ER junctions, increased the ER Ca releasable fraction by about 20%, thereby supporting the hypothesis that this process of privileged ER refilling is junction-mediated. Junctions were observed in the cell ultrastructure and their main parameters of membrane separation and linear extension were (9.6 ± 3.8) nm and (128 ± 63) nm, respectively. Our findings point to a process of privileged refilling of the ER, in which NCX and store-operated Ca entry via the stromal interaction molecule (STIM)-Orai system are the sole protagonists. These results shed light on the molecular machinery involved in the function of a previously hypothesized subplasmalemmal Ca control unit during ER refilling with extracellular Ca.
- Subjects
SODIUM-calcium exchange; ENDOPLASMIC reticulum; VASCULAR endothelial cells; CALCIUM channels; TRP channels
- Publication
Pflügers Archiv: European Journal of Physiology, 2017, Vol 469, Issue 10, p1287
- ISSN
0031-6768
- Publication type
Article
- DOI
10.1007/s00424-017-1989-8