Involvement of trp-2 protein in store-operated influx of calcium in fibroblasts.
Mammalian homologues of the Drosophila melanogaster transient receptor potential (trp) gene have been proposed to encode store-operated channels. This assertion essentially stays on the fact that expression of different trp proteins produces trans-membrane cation currents. However, the selectivity of the expressed channels and their mode of activation, in particular, their dependence to store depletion appears to be quite variable. In the present work, we adopted an anti-sense strategy to study this question in transfected Chinese hamster ovary cells expressing the rat neurotensin receptor (CHO-NTR cells), a cellular model characterized by its very large store-dependent entry of Ca(2+). We identified different trp transcripts by RT-PCR, the trp-1 and trp-2 transcripts being by far the most abundant. CHO-NTR cells were then transfected with a mouse trp-2 anti-sense construct (CHO-NTR-TRP2AS cells). We showed that in these cells, trp-2 mRNA was suppressed in comparison with cells transfected with a control plasmid. The store-operated entry of Ca(2+) was evaluated after store depletion by an IP(3)-dependent mechanism (neurotensin stimulation) or by direct inhibition of the endoplasmic reticulum Ca(2+)ATPase (thapsigargin stimulation). In both cases, store-dependent entry of Ca(2+) was largely reduced in CHO-NTR-TRP2AS cells in comparison with control cells, suggesting that trp-2 protein might constitute a functional subunit of store-operated channels.
Gailly
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Département de Physiologie et de Pharmacologie, Université Catholique de Louvain (UCL 5540), Av. Hippocrate 55, 1200 Brussels, Belgium. gailly@fymu.ucl.ac.be
Colson-Van Schoor
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D000818:Animals; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects; D006224:Cricetinae; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism*; D015870:Gene Expression; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D009496:Neurotensin / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D018028:Receptors, Neurotensin / Q000235:genetics / Q000378:metabolism; D050052:TRPC Cation Channels; D019284:Thapsigargin / Q000494:pharmacology; D014162:Transfection
D008565:Membrane Proteins; D012333:RNA, Messenger; D018028:Receptors, Neurotensin; D050052:TRPC Cation Channels; C113139:Trp2 protein, vertebrate; C491858:Trpc2 protein, mouse; C491857:Trpc2 protein, rat; D009496:Neurotensin; D019284:Thapsigargin; D015544:Inositol 1,4,5-Trisphosphate; D002118:Calcium