Li Yingxiang, Zhang Linlin, Qu Tao, Li Li, Zhang Guofan
University of Chinese Academy of Sciences, Beijing, China.
Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao, China.
PLoS One. 2016 Jan 4;11(1):e0146049. doi: 10.1371/journal.pone.0146049. eCollection 2016.
Genomic and transcriptomic studies have revealed a sophisticated and powerful apoptosis regulation network in oyster, highlighting its adaptation to sessile life in a highly stressful intertidal environment. However, the functional molecular basis of apoptosis remains largely unexplored in oysters. In this study, we focused on a representative apoptotic gene encoding voltage-dependent anion channel 2 (VDAC2), a porin that abounds at the mitochondrial outer membrane. This is the first report on the identification and characterization of a VDAC gene in the Pacific oyster, Crassostrea gigas (CgVDAC2). The full length of CgVDAC2 was 1,738 bp with an open reading frame of 843 bp that encoded a protein of 281 amino acids. A four-element eukaryotic porin signature motif, a conserved ATP binding motif, and a VKAKV-like sequence were identified in the predicted CgVDAC2. Expression pattern analysis in different tissues and developmental stages as well as upon infection by ostreid herpesvirus 1 revealed the energy supply-related and immunity-related expression of CgVDAC2. CgVDAC2 was co-localized with mitochondria when it was transiently transfected into HeLa cells. Overexpression of CgVDAC2 in HEK293T cells suppressed the UV irradiation-induced apoptosis by inhibiting the pro-apoptotic function of CgBak. RNA interference induced reduction in CgVDAC2 expression showed a promoted apoptosis level upon UV light irradiation in hemocytes. The yeast two-hybrid system and co-immunoprecipitation assay indicated a direct interaction between CgVDAC2 and the pro-apoptotic protein CgBak. This study revealed the function of VDAC2 in oyster and provided new insights into its involvement in apoptosis modulation and host defense in mollusks.
基因组和转录组研究揭示了牡蛎中一个复杂而强大的细胞凋亡调控网络，突出了其在高度应激的潮间带环境中对固着生活的适应性。然而，牡蛎细胞凋亡的功能分子基础在很大程度上仍未被探索。在本研究中，我们聚焦于一个具有代表性的凋亡基因，该基因编码电压依赖性阴离子通道2（VDAC2），一种存在于线粒体外膜的孔蛋白。这是关于太平洋牡蛎（Crassostrea gigas，CgVDAC2）中VDAC基因的鉴定和表征的首次报道。CgVDAC2的全长为1738 bp，开放阅读框为843 bp，编码一个由281个氨基酸组成的蛋白质。在预测的CgVDAC2中鉴定出了一个四元件真核孔蛋白特征基序、一个保守的ATP结合基序和一个类似VKAKV的序列。在不同组织和发育阶段以及感染牡蛎疱疹病毒1后的表达模式分析揭示了CgVDAC2与能量供应相关和免疫相关的表达。当CgVDAC2瞬时转染到HeLa细胞中时，它与线粒体共定位。在HEK293T细胞中过表达CgVDAC2通过抑制CgBak的促凋亡功能抑制了紫外线照射诱导的细胞凋亡。RNA干扰诱导的CgVDAC2表达降低显示血细胞在紫外线照射后凋亡水平升高。酵母双杂交系统和免疫共沉淀分析表明CgVDAC2与促凋亡蛋白CgBak之间存在直接相互作用。本研究揭示了VDAC2在牡蛎中的功能，并为其参与软体动物的细胞凋亡调节和宿主防御提供了新的见解。
