Na+/Ca2+ exchangers and Orai channels jointly refill endoplasmic reticulum (ER) Ca2+ via ER nanojunctions in vascular endothelial cells View Full Text


Ontology type: schema:ScholarlyArticle      Open Access: True


Article Info

DATE

2017-05-11

AUTHORS

Cristiana M. L. Di Giuro, Niroj Shrestha, Roland Malli, Klaus Groschner, Cornelis van Breemen, Nicola Fameli

ABSTRACT

We investigated the role of Na+/ Ca2+ exchange (NCX) in the refilling of endoplasmic reticulum (ER) Ca2+ in vascular endothelial cells under various conditions of cell stimulation and plasma membrane (PM) polarization. Better understanding of the mechanisms behind basic ER Ca2+ content regulation is important, since current hypotheses on the possible ultimate causes of ER stress point to deterioration of the Ca2+ transport mechanism to/from ER itself. We measured [Ca2+]i 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 Ca2+ ATPase (SERCA), Na+/ K+ ATPase (NKA)) involved in the Ca2+ communication between the extracellular space and the ER. Following histamine-stimulated ER Ca2+ release, blockade of NCX Ca2+-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 Ca2+ 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 Ca2+ 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 Ca2+ control unit during ER refilling with extracellular Ca2+. More... »

PAGES

1287-1299

Identifiers

URI

http://scigraph.springernature.com/pub.10.1007/s00424-017-1989-8

DOI

http://dx.doi.org/10.1007/s00424-017-1989-8

DIMENSIONS

https://app.dimensions.ai/details/publication/pub.1085391974

PUBMED

https://www.ncbi.nlm.nih.gov/pubmed/28497275


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