Ivanova Hristina, Luyten Tomas, Decrock Elke, Vervliet Tim, Leybaert Luc, Parys Jan B, Bultynck Geert
KU Leuven, Laboratory of Molecular and Cellular Signaling, Department of Cellular and Molecular Medicine and, Herestraat 49, BE-3000, Leuven, Belgium.
Ghent University, Physiology Group, Department of Basic Medical Sciences, BE-9000, Ghent, Belgium.
Cell Calcium. 2017 Mar;62:41-46. doi: 10.1016/j.ceca.2017.01.010. Epub 2017 Jan 25.
Ca signalling plays an important role in various physiological processes in vertebrates. In mammals, the highly conserved anti-apoptotic B-cell lymphoma-2 (Bcl-2) protein is an important modulator of the inositol 1,4,5-trisphosphate receptor (IPR), i.e. the main intracellular Ca - release channel located at the endoplasmic reticulum (ER). The Bcl-2 Homology (BH) 4 domain of Bcl-2 (BH4-Bcl-2) is a critical determinant for inhibiting IPRs, by directly targeting a region in the modulatory domain of the receptor (domain 3). In this paper, we aimed to track the evolutionary history of IPR regulation by the BH4 domain of Bcl-2 orthologues from different classes of vertebrates, including Osteichthyes, Amphibia, Reptilia, Aves and Mammalia. The high degree of conservation of the BH4 sequences correlated with the ability of all tested peptides to bind to the domain 3 of mouse IPR1 in GST-pull downs and their overall ability to inhibit IP-induced Ca release (IICR) in permeabilized cells. Nevertheless, the BH4 domains differed in their potency to suppress IICR. The peptide derived from X. laevis was the least potent inhibitor. We identified a critical residue in BH4-Bcl-2 from H. sapiens, Thr7, which is replaced by Gly7 in X. laevis. Compared to the wild type X. laevis BH4-Bcl-2, a "humanized" version of the peptide (BH4-Bcl-2 Gly7Thr), displayed increased IPR-inhibitory properties. Despite the differences in the inhibitory efficiency, our data indicate that the BH4 domain of Bcl-2 orthologues from different classes of vertebrates can act as a binding partner and inhibitor of IPR channels.
钙信号传导在脊椎动物的各种生理过程中起着重要作用。在哺乳动物中，高度保守的抗凋亡B细胞淋巴瘤-2（Bcl-2）蛋白是肌醇1,4,5-三磷酸受体（IPR）的重要调节剂，即位于内质网（ER）的主要细胞内钙释放通道。Bcl-2的Bcl-2同源（BH）4结构域（BH4-Bcl-2）是抑制IPR的关键决定因素，它直接靶向受体调节结构域（结构域3）中的一个区域。在本文中，我们旨在追踪不同类脊椎动物（包括硬骨鱼纲、两栖纲、爬行纲、鸟纲和哺乳纲）中Bcl-2直系同源物的BH4结构域对IPR调节的进化历程。BH4序列的高度保守性与所有测试肽在GST下拉实验中与小鼠IPR1结构域3结合的能力及其在通透细胞中抑制IP诱导的钙释放（IICR）的总体能力相关。然而，BH4结构域在抑制IICR的效力上存在差异。源自非洲爪蟾的肽是效力最低的抑制剂。我们在智人的BH4-Bcl-2中鉴定出一个关键残基，苏氨酸7，在非洲爪蟾中它被甘氨酸7取代。与野生型非洲爪蟾BH4-Bcl-2相比，该肽的“人源化”版本（BH4-Bcl-2 Gly7Thr）表现出增强的IPR抑制特性。尽管抑制效率存在差异，但我们的数据表明，不同类脊椎动物中Bcl-2直系同源物的BH4结构域可作为IPR通道的结合伴侣和抑制剂。
