Bcl-2 and IP3 compete for the ligand-binding domain of IP3Rs modulating Ca2+ signaling output
Bcl-2 and IP3 compete for the ligand-binding domain of IP3Rs modulating Ca2+ signaling output
复制标题
DOI：
10.1007/s00018-019-03091-8
复制
发表时间：
2019-10-01
影响因子：
8
通讯作者：
Bultynck, Geert
中科院分区：
文献类型：
--
作者：
Ivanova, Hristina;Wagner, Larry E.;Bultynck, Geert
Bcl-2蛋白通过直接靶向和抑制IP 3受体（IP 3R）（一种主要的细胞内Ca 2+释放通道）而成为细胞内Ca 2+动力学的关键调节剂。在这里，我们证明了这种抑制作用发生在基础的条件下，但不是高IP 3R活性，因为过表达和纯化的Bcl-2（或其BH 4结构域）可以抑制由低浓度的激动剂或IP 3引起的IP 3R功能，而不能减弱对高浓度的激动剂或IP 3。令人惊讶的是，Bcl-2仍然能够抑制IP（3）R1通道，该通道缺乏包含先前鉴定的Bcl-2结合位点的残基（a.a. 1380-1408），位于ARM 2结构域，调节区的一部分。使用大量的计算，生物化学和生物物理方法，我们证明Bcl-2，更具体地说，其BH 4结构域结合IP（3）R1的配体结合结构域（LBD）。与这一发现一致，LBD和Bcl-2（或其BH 4结构域）之间的相互作用对IP 3和腺苷酸A（IP 3R的配体）敏感。反之亦然，Bcl-2的BH 4结构域抵消了IP 3与LBD的结合。总的来说，我们的工作揭示了Bcl-2在LBD水平影响IP 3R活性的新机制。这允许精确调节Bcl-2对IP（3）Rs的抑制特性，其可调节到细胞中的IP 3信号传导水平。
Bcl-2 proteins have emerged as critical regulators of intracellular Ca2+ dynamics by directly targeting and inhibiting the IP3 receptor (IP3R), a major intracellular Ca2+-release channel. Here, we demonstrate that such inhibition occurs under conditions of basal, but not high IP3R activity, since overexpressed and purified Bcl-2 (or its BH4 domain) can inhibit IP3R function provoked by low concentration of agonist or IP3, while fails to attenuate against high concentration of agonist or IP3. Surprisingly, Bcl-2 remained capable of inhibiting IP(3)R1 channels lacking the residues encompassing the previously identified Bcl-2-binding site (a.a. 1380-1408) located in the ARM2 domain, part of the modulatory region. Using a plethora of computational, biochemical and biophysical methods, we demonstrate that Bcl-2 and more particularly its BH4 domain bind to the ligand-binding domain (LBD) of IP(3)R1. In line with this finding, the interaction between the LBD and Bcl-2 (or its BH4 domain) was sensitive to IP3 and adenophostin A, ligands of the IP3R. Vice versa, the BH4 domain of Bcl-2 counteracted the binding of IP3 to the LBD. Collectively, our work reveals a novel mechanism by which Bcl-2 influences IP3R activity at the level of the LBD. This allows for exquisite modulation of Bcl-2's inhibitory properties on IP(3)Rs that is tunable to the level of IP3 signaling in cells.