Modification of phospholipase C-γ-induced Ca2+ signal generation by 2-aminoethoxydiphenyl borate

Hong Tao Ma, Kartik Venkatachalam, Krystyna E. Rys-Sikora, Li Ping He, Fei Zheng, Donald L. Gill

Research output: Contribution to journalArticle

23 Scopus citations

Abstract

The mechanisms by which Ca2+-store-release channels and Ca 2+-entry channels are coupled to receptor activation are poorly understood. Modification of Ca2+ signals by 2-aminoethoxydiphenyl borate (2-APB), suggests the agent may target entry channels or the machinery controlling their activation. In DT40 B-cells and Jurkat T-cells, complete Ca2+ store release was induced by 2-APB (EC50 10-20 μM). At 75 μM, 2-APB emptied stores completely in both lymphocyte lines, but had no such effect on other cells. In DT40 cells, 2-APB mimicked B-cell receptor (BCR) cross-linking, but no effect was observed in mutant DT40 lines devoid of inositol 1,4,5-trisphosphate (InsP3) receptors (InsP 3Rs) or phospholipase C-γ2 (PLC-γ2). Like the BCR, 2-APB activated transfected TRPC3 (canonical transient receptor potential) channels, which acted as sensors for PLC-γ2-generated diacylglycerol in DT40 cells. The action of 2-APB on InsP3Rs and TRPC3 channels was prevented by PLC-inhibition, and required PLC-γ2 catalytic activity. However, unlike BCR activation, no increased InsP3 level could be measured in response to 2-APB. Also, calyculin A-induced cytoskeletal reorganization prevented 2-APB-induced InsP3R and TRPC3-channel activation, but not that induced by the BCR. 2-APB still activated TRPC3 channels in DT40 cells with fully depleted Ca2+ stores, indicating its action was not via Ca2+ release. Significantly, 2-APB-induced InsP3R and TRPC3 activation was prevented in DT40 knockout cells devoid of the BCR- and PLC-γ2-coupled adaptor/kinases, Syk, Lyn, Btk or BLNK. The results suggest that 2-APB activates Ca2+ signals in lymphocytes by initiating and enhancing coupling between components of the BCR-PLC-γ2 complex and both Ca2+-entry and Ca2+-release channels.

Original languageEnglish (US)
Pages (from-to)667-676
Number of pages10
JournalBiochemical Journal
Volume376
Issue number3
DOIs
Publication statusPublished - Dec 15 2003

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All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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