IP3 receptor isoforms differently regulate ER-mitochondrial contacts and local calcium transfer

Contact sites of endoplasmic reticulum (ER) and mitochondria locally convey calcium signals between the IP3 receptors (IP3R) and the mitochondrial calcium uniporter, and are central to cell survival. It remains unclear whether IP3Rs also have a structural role in contact formation and whether the different IP3R isoforms have redundant functions. Using an IP3R-deficient cell model rescued with each of the three IP3R isoforms and an array of super-resolution and ultrastructural approaches we demonstrate that IP3Rs are required for maintaining ER-mitochondrial contacts. This role is independent of calcium fluxes. We also show that, while each isoform can support contacts, type 2 IP3R is the most effective in delivering calcium to the mitochondria. Thus, these studies reveal a non-canonical, structural role for the IP3Rs and direct attention towards the type 2 IP3R that was previously neglected in the context of ER-mitochondrial calcium signaling

DPB162-AE, an inhibitor of store-operated Ca2+ entry, can deplete the endoplasmic reticulum Ca2+ store

Store-operated Ca2+ entry (SOCE), an important Ca2+ signaling pathway in non-excitable cells, regulates a variety of cellular functions. To study its physiological role, pharmacological tools, like 2-aminoethyl diphenylborinate (2-APB), are used to impact SOCE. 2-APB is one of the best characterized SOCE inhibitors. However, 2-APB also activates SOCE at lower concentrations, while it inhibits inositol 1,4,5-trisphosphate receptors (IP3Rs), sarco/endoplasmic reticulum Ca2+-ATPases (SERCAs) and other ion channels, like TRP channels. Because of this, 2-APB analogues that inhibit SOCE more potently and more selectively compared to 2-APB have been developed. The recently developed DPB162-AE is such a structural diphenylborinate isomer of 2-APB that selectively inhibits SOCE currents by blocking the functional coupling between STIM1 and Orai1. However, we observed an adverse effect of DPB162-AE on the ER Ca2+-store content at concentrations required for complete SOCE inhibition. DPB162-AE increased the cytosolic Ca2+ levels by reducing the ER Ca2+ store in cell lines as well as in primary cells. DPB162-AE did not affect SERCA activity, but provoked a Ca2+ leak from the ER, even after application of the SERCA inhibitor thapsigargin. IP3Rs partly contributed to the DPB162-AE-induced Ca2+ leak, since pharmacologically and genetically inhibiting IP3R function reduced, but not completely blocked, the effects of DPB162-AE on the ER store content. Our results indicate that, in some conditions, the SOCE inhibitor DPB162-AE can reduce the ER Ca2+-store content by inducing a Ca2+-leak pathway at concentrations needed for adequate SOCE inhibition.publisher: Elsevier articletitle: DPB162-AE, an inhibitor of store-operated Ca2+ entry, can deplete the endoplasmic reticulum Ca2+ store journaltitle: Cell Calcium articlelink: http://dx.doi.org/10.1016/j.ceca.2017.01.015 content_type: article copyright: © 2017 Elsevier Ltd. All rights reserved.status: publishe

DPB162-AE, an inhibitor of store-operated Ca2+ entry, can deplete the endoplasmic reticulum Ca2+ store

Store-operated Ca2+ entry (SOCE), an important Ca2+ signaling pathway in non-excitable cells, regulates a variety of cellular functions. To study its physiological role, pharmacological tools, like 2-aminoethyl diphenylborinate (2-APB), are used to impact SOCE. 2-APB is one of the best characterized SOCE inhibitors. However, 2-APB also activates SOCE at lower concentrations, while it inhibits inositol 1,4,5-trisphosphate receptors (IP3Rs), sarco/endoplasmic reticulum Ca2+-ATPases (SERCAs) and other ion channels, like TRP channels. Because of this, 2-APB analogues that inhibit SOCE more potently and more selectively compared to 2-APB have been developed. The recently developed DPB162-AE is such a structural diphenylborinate isomer of 2-APB that selectively inhibits SOCE currents by blocking the functional coupling between STIM1 and Orai1. However, we observed an adverse effect of DPB162-AE on the ER Ca2+-store content at concentrations required for complete SOCE inhibition. DPB162-AE increased the cytosolic Ca2+ levels by reducing the ER Ca2+ store in cell lines as well as in primary cells. DPB162-AE did not affect SERCA activity, but provoked a Ca2+ leak from the ER, even after application of the SERCA inhibitor thapsigargin. IP3Rs partly contributed to the DPB162-AE-induced Ca2+ leak, since pharmacologically and genetically inhibiting IP3R function reduced, but not completely blocked, the effects of DPB162-AE on the ER store content. Our results indicate that, in some conditions, the SOCE inhibitor DPB162-AE can reduce the ER Ca2+-store content by inducing a Ca2+-leak pathway at concentrations needed for adequate SOCE inhibition