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2-Aminoethoxydiphenyl Borate
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Donald L. Gill – One of the best experts on this subject based on the ideXlab platform.
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Molecular mechanisms underlying inhibition of STIM1-Orai1-mediated Ca2+ entry induced by 2-Aminoethoxydiphenyl Borate.
Pflugers Archiv : European journal of physiology, 2016Co-Authors: Yandong Zhou, Guolin Ma, Lian He, Lijuan Zhou, Shuce Zhang, Shenyuan L Zhang, Donald L. GillAbstract:Store-operated Ca2+ entry (SOCE) mediated by STIM1 and Orai1 is crucial for Ca2+ signaling and homeostasis in most cell types. 2-Aminoethoxydiphenyl Borate (2-APB) is a well-described SOCE inhibitor, but its mechanisms of action remain largely elusive. Here, we show that 2-APB does not affect the dimeric state of STIM1, but enhances the intramolecular coupling between the coiled-coil 1 (CC1) and STIM-Orai-activating region (SOAR) of STIM1, with subsequent reduction in the formation of STIM1 puncta in the absence of Orai1 overexpression. 2-APB also inhibits Orai1 channels, directly inhibiting Ca2+ entry through the constitutively active, STIM1-independent Orai1 mutants, Orai1-P245T and Orai1-V102A. When unbound from STIM1, the constitutively active Orai1-V102C mutant is not inhibited by 2-APB. Thus, we used Orai1-V012C as a tool to examine whether 2-APB can also inhibit the coupling between STIM1 and Orai1. We reveal that the functional coupling between STIM1 and Orai1-V102C is inhibited by 2-APB. This inhibition on coupling is indirect, arising from 2-APB’s action on STIM1, and it is most likely mediated by functional channel residues in the Orai1 N-terminus. Overall, our findings on this two-site inhibition mediated by 2-APB provide new understanding on Orai1-activation by STIM1, important to future drug design.
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Mechanism of Activation of Store-Operated Calcium Entry by 2-Aminoethoxydiphenyl Borate
Biophysical Journal, 2014Co-Authors: Yandong Zhou, Youjun Wang, Xizhuo Wang, Lucas Occhiena, Daniel Y. Chung, Donald L. GillAbstract:Store-operated Ca2+ entry (SOCE) is mediate by STIM-induced activation of Orai channels. The small molecule, 2-Aminoethoxydiphenyl Borate (2-APB), is known to have a biphasic effect on SOCE; lower 2-APB (≤10 μM) enhances, while higher levels of 2-APB (50-100 μM) are strongly inhibitory following a transient activation. But the mechanism by which 2-APB activates SOCe is still elusive. The effects of 2-APB were examined on coupling between various Orai1 mutants and STIM1, C-terminal (ct) STIM1 fragments, or the STIM-Orai activating region (SOAR) of STIM1, using a combination of imaging and whole-cell patch clamp analysis. Orai1 coupling with STIM1ct, STIM1ct-4EA and SOAR can be transiently activated by 2-APB at low levels (10 µM). The activation of SOCE by low 2-APB is induced by its action on the SOAR-Orai1 complex. We examined which domains/residues in Orai1 or in STIM1 are essential for the activating effect of 2-APB. Adjustment of cytosolic pH upward using nigericin abolished the action of 2-APB. In contrast, lower pH potentiated the activating effect of 2-APB. Our results reveal that the Orai1 C-terminus and N-terminus function in a concerted manner to mediate the activating effect of 2-APB on SOCE. Mutational analysis reveals that the negatively charged residues within the Orai1 C-terminal region are not required for the 2-APB-induced SOCE activation. Our studies provide new insights into the mechanism of 2-APB-induced activation of coupling between STIM and Orai and the triggering of SOCE.
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Modification of phospholipase C-γ-induced Ca2+ signal generation by 2-Aminoethoxydiphenyl Borate
Biochemical Journal, 2003Co-Authors: Kartik Venkatachalam, Krystyna E. Rys-sikora, Li Ping He, Fei Zheng, Donald L. GillAbstract:The mechanisms by which Ca(2+)-store-release channels and Ca(2+)-entry channels are coupled to receptor activation are poorly understood. Modification of Ca(2+) 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 Ca(2+) store release was induced by 2-APB (EC(50) 10-20 microM). At 75 microM, 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 (InsP(3)) receptors (InsP(3)Rs) or phospholipase C-gamma2 (PLC-gamma2). Like the BCR, 2-APB activated transfected TRPC3 (canonical transient receptor potential) channels, which acted as sensors for PLC-gamma2-generated diacylglycerol in DT40 cells. The action of 2-APB on InsP(3)Rs and TRPC3 channels was prevented by PLC-inhibition, and required PLC-gamma2 catalytic activity. However, unlike BCR activation, no increased InsP(3) level could be measured in response to 2-APB. Also, calyculin A-induced cytoskeletal reorganization prevented 2-APB-induced InsP(3)R and TRPC3-channel activation, but not that induced by the BCR. 2-APB still activated TRPC3 channels in DT40 cells with fully depleted Ca(2+) stores, indicating its action was not via Ca(2+) release. Significantly, 2-APB-induced InsP(3)R and TRPC3 activation was prevented in DT40 knockout cells devoid of the BCR- and PLC-gamma2-coupled adaptor/kinases, Syk, Lyn, Btk or BLNK. The results suggest that 2-APB activates Ca(2+) signals in lymphocytes by initiating and enhancing coupling between components of the BCR-PLC-gamma2 complex and both Ca(2+)-entry and Ca(2+)-release channels.
Michael J Berridge – One of the best experts on this subject based on the ideXlab platform.
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2-Aminoethoxydiphenyl Borate (2-APB) is a reliable blocker of store-operated Ca2+ entry but an inconsistent inhibitor of InsP3-induced Ca2+ release.
The FASEB Journal, 2020Co-Authors: Martin D. Bootman, Michael J Berridge, H. Llewelyn Roderick, Tony J. Collins, Lauren Mackenzie, Claire M. PeppiattAbstract:Since its introduction to Ca2+ signaling in 1997, 2-Aminoethoxydiphenyl Borate (2-APB) has been used in many studies to probe for the involvement of inositol 1,4,5-trisphosphate receptors in the generation of Ca2+ signals. Due to reports of some nonspecific actions of 2-APB, and the fact that its principal antagonistic effect is on Ca2+ entry rather than Ca2+ release, this compound may not have the utility first suggested. However, 2-APB has thrown up some interesting results, particularly with respect to store-operated Ca2+ entry in nonexcitable cells. These data indicate that although it must be used with caution, 2-APB can be useful in probing certain aspects of Ca2+ signaling.—Bootman, M. D., Collins, T. J., Mackenzie, L., Roderick, H. L., Berridge, M. J., Peppiatt, C. M. 2-Aminoethoxydiphenyl Borate (2-APB) is a reliable blocker of store-operated Ca2+ entry but an inconsistent inhibitor of InsP3-induced Ca2+ release.
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2-Aminoethoxydiphenyl Borate (2-APB) antagonises inositol 1,4,5-trisphosphate-induced calcium release, inhibits calcium pumps and has a use-dependent and slowly reversible action on store-operated calcium entry channels
Cell Calcium, 2003Co-Authors: Claire M. Peppiatt, Michael J Berridge, Martin D. Bootman, Tony J. Collins, Lauren Mackenzie, Stuart J. Conway, Andrew B. Holmes, H. Llewelyn RoderickAbstract:Abstract The action of 2-Aminoethoxydiphenyl Borate (2-APB) on Ca2+ signalling in HeLa cells and cardiac myocytes was investigated. Consistent with other studies, we found that superfusion of cells with 2-APB rapidly inhibited inositol 1,4,5-trisphosphate (InsP3)-mediated Ca2+ release and store-operated Ca2+ entry (SOC). In addition to abrogating hormone-evoked Ca2+ responses, 2-APB could antagonise Ca2+ signals evoked by a membrane permeant InsP3 ester. 2-APB also slowed the recovery of intracellular Ca2+ signals consistent with an effect on Ca2+ ATPases. The inhibitory action of 2-APB on InsP3 receptors (InsP3Rs), SOC channels and Ca2+ pumps persisted for several minutes after washout of the compound. Application of 2-APB to unstimulated cells had no effect on subsequent Ca2+ responses suggesting that it has a use-dependent action. Mitochondria in cells treated with 2-APB showed a rapid and slowly reversible swelling. 2-APB did not cause the mitochondria to depolarise, but it reduced the extent of mitochondrial calcium uptake. Although 2-APB has been demonstrated not to affect voltage-operated Ca2+ channels or ryanodine receptors, we found that it gave a concentration-dependent long-lasting inhibition of Ca2+ signalling in electrically-stimulated cardiac myocytes, where InsP3Rs and SOC channels do not play a significant role. Our data suggest that 2-APB has multiple cellular targets, a use-dependent action, is difficult to reverse and may affect Ca2+ signalling in cell types where InsP3 and SOC are not active.
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2-Aminoethoxydiphenyl Borate (2-APB) is a reliable blocker of store-operated Ca 2+ entry but an inconsistent inhibitor of InsP 3 -induced Ca 2+ release
The FASEB Journal, 2002Co-Authors: Lewis Mackenzie, H. Llewelyn Roderick, Martin D. Bootman, Claire M. Peppiatt, Tony J. Collins, Michael J BerridgeAbstract:Since its introduction to Ca2+ signaling in 1997, 2-Aminoethoxydiphenyl Borate (2-APB) has been used in many studies to probe for the involvement of inositol 1,4,5-trisphosphate receptors in the generation of Ca2+ signals. Due to reports of some nonspecific actions of 2-APB, and the fact that its principal antagonistic effect is on Ca2+ entry rather than Ca2+ release, this compound may not have the utility first suggested. However, 2-APB has thrown up some interesting results, particularly with respect to store-operated Ca2+ entry in nonexcitable cells. These data indicate that although it must be used with caution, 2-APB can be useful in probing certain aspects of Ca2+ signaling.
Claire M. Peppiatt – One of the best experts on this subject based on the ideXlab platform.
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2-Aminoethoxydiphenyl Borate (2-APB) is a reliable blocker of store-operated Ca2+ entry but an inconsistent inhibitor of InsP3-induced Ca2+ release.
The FASEB Journal, 2020Co-Authors: Martin D. Bootman, Michael J Berridge, H. Llewelyn Roderick, Tony J. Collins, Lauren Mackenzie, Claire M. PeppiattAbstract:Since its introduction to Ca2+ signaling in 1997, 2-Aminoethoxydiphenyl Borate (2-APB) has been used in many studies to probe for the involvement of inositol 1,4,5-trisphosphate receptors in the generation of Ca2+ signals. Due to reports of some nonspecific actions of 2-APB, and the fact that its principal antagonistic effect is on Ca2+ entry rather than Ca2+ release, this compound may not have the utility first suggested. However, 2-APB has thrown up some interesting results, particularly with respect to store-operated Ca2+ entry in nonexcitable cells. These data indicate that although it must be used with caution, 2-APB can be useful in probing certain aspects of Ca2+ signaling.—Bootman, M. D., Collins, T. J., Mackenzie, L., Roderick, H. L., Berridge, M. J., Peppiatt, C. M. 2-Aminoethoxydiphenyl Borate (2-APB) is a reliable blocker of store-operated Ca2+ entry but an inconsistent inhibitor of InsP3-induced Ca2+ release.
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2-Aminoethoxydiphenyl Borate (2-APB) antagonises inositol 1,4,5-trisphosphate-induced calcium release, inhibits calcium pumps and has a use-dependent and slowly reversible action on store-operated calcium entry channels
Cell Calcium, 2003Co-Authors: Claire M. Peppiatt, Michael J Berridge, Martin D. Bootman, Tony J. Collins, Lauren Mackenzie, Stuart J. Conway, Andrew B. Holmes, H. Llewelyn RoderickAbstract:Abstract The action of 2-Aminoethoxydiphenyl Borate (2-APB) on Ca2+ signalling in HeLa cells and cardiac myocytes was investigated. Consistent with other studies, we found that superfusion of cells with 2-APB rapidly inhibited inositol 1,4,5-trisphosphate (InsP3)-mediated Ca2+ release and store-operated Ca2+ entry (SOC). In addition to abrogating hormone-evoked Ca2+ responses, 2-APB could antagonise Ca2+ signals evoked by a membrane permeant InsP3 ester. 2-APB also slowed the recovery of intracellular Ca2+ signals consistent with an effect on Ca2+ ATPases. The inhibitory action of 2-APB on InsP3 receptors (InsP3Rs), SOC channels and Ca2+ pumps persisted for several minutes after washout of the compound. Application of 2-APB to unstimulated cells had no effect on subsequent Ca2+ responses suggesting that it has a use-dependent action. Mitochondria in cells treated with 2-APB showed a rapid and slowly reversible swelling. 2-APB did not cause the mitochondria to depolarise, but it reduced the extent of mitochondrial calcium uptake. Although 2-APB has been demonstrated not to affect voltage-operated Ca2+ channels or ryanodine receptors, we found that it gave a concentration-dependent long-lasting inhibition of Ca2+ signalling in electrically-stimulated cardiac myocytes, where InsP3Rs and SOC channels do not play a significant role. Our data suggest that 2-APB has multiple cellular targets, a use-dependent action, is difficult to reverse and may affect Ca2+ signalling in cell types where InsP3 and SOC are not active.
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2-Aminoethoxydiphenyl Borate (2-APB) is a reliable blocker of store-operated Ca 2+ entry but an inconsistent inhibitor of InsP 3 -induced Ca 2+ release
The FASEB Journal, 2002Co-Authors: Lewis Mackenzie, H. Llewelyn Roderick, Martin D. Bootman, Claire M. Peppiatt, Tony J. Collins, Michael J BerridgeAbstract:Since its introduction to Ca2+ signaling in 1997, 2-Aminoethoxydiphenyl Borate (2-APB) has been used in many studies to probe for the involvement of inositol 1,4,5-trisphosphate receptors in the generation of Ca2+ signals. Due to reports of some nonspecific actions of 2-APB, and the fact that its principal antagonistic effect is on Ca2+ entry rather than Ca2+ release, this compound may not have the utility first suggested. However, 2-APB has thrown up some interesting results, particularly with respect to store-operated Ca2+ entry in nonexcitable cells. These data indicate that although it must be used with caution, 2-APB can be useful in probing certain aspects of Ca2+ signaling.