Bcl-xL Reduces Chinese Giant Salamander Iridovirus-Induced Mitochondrial Apoptosis by Interacting with Bak and Inhibiting the p53 Pathway.

Yiqun Li,Yuding Fan,Yan Meng,Jingjing Zhang,Mingyang Xue,Ge Lin,Yong Zhou,Wenzhi Liu,Lingbing Zeng,Nan Jiang

doi:10.3390/v13112224

Yiqun Li, Yuding Fan + Show 8 more

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https://doi.org/10.3390/v13112224

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Abstract

Chinese giant salamander iridovirus (GSIV) infection could lead to mitochondrial apoptosis in this animal, a process that involves B-cell lymphoma-2 (BCL-2) superfamily molecules. The mRNA expression level of Bcl-xL, a crucial antiapoptotic molecule in the BCL-2 family, was reduced in early infection and increased in late infection. However, the molecular mechanism remains unknown. In this study, the function and regulatory mechanisms of Chinese giant salamander (Andrias davidianus) Bcl-xL (AdBcl-xL) during GSIV infection were investigated. Western blotting assays revealed that the level of Bcl-xL protein was downregulated markedly as the infection progressed. Plasmids expressing AdBcl-xL or AdBcl-xL short interfering RNAs were separately constructed and transfected into Chinese giant salamander muscle cells. Confocal microscopy showed that overexpressed AdBcl-xL was translocated to the mitochondria after infection with GSIV. Additionally, flow cytometry analysis demonstrated that apoptotic progress was reduced in both AdBcl-xL-overexpressing cells compared with those in the control, while apoptotic progress was enhanced in cells silenced for AdBcl-xL. A lower number of copies of virus major capsid protein genes and a reduced protein synthesis were confirmed in AdBcl-xL-overexpressing cells. Moreover, AdBcl-xL could bind directly to the proapoptotic molecule AdBak with or without GSIV infection. In addition, the p53 level was inhibited and the mRNA expression levels of crucial regulatory molecules in the p53 pathway were regulated in AdBcl-xL-overexpressing cells during GSIV infection. These results suggest that AdBcl-xL plays negative roles in GSIV-induced mitochondrial apoptosis and virus replication by binding to AdBak and inhibiting p53 activation.

Highlights

The B-cell lymphoma-2 (BCL-2) superfamily, which comprises both proapoptotic molecules and antiapoptotic molecules, are proteins characterized by the existence of short conserved sequence regions (the BCL-2 homology (BH) motifs), playing crucial roles in determining whether the cells survive or die [1,2]
43% of overall sequence identities with the Bcl-xL homolog from amphibian and fish species, including Xenopus laevis, X. tropicalis, Ctenopharyngodon idella, Clarias magur, Danio rerio, Epinephelus coioides, and Gadus morhua (Figure 1)
The results of the present study suggest that AdBcl-xL can interact directly with AdBak under giant salamander iridovirus (GSIV) infection or in normal conditions

Summary

Introduction

The B-cell lymphoma-2 (BCL-2) superfamily, which comprises both proapoptotic molecules and antiapoptotic molecules, are proteins characterized by the existence of short conserved sequence regions (the BCL-2 homology (BH) motifs), playing crucial roles in determining whether the cells survive or die [1,2]. The ratio of antagonist (antiapoptotic) to agonist (proapoptotic) molecules dictates whether a cell responds to a proximal apoptotic stimulus [3]. Bcl-xL is an antiapoptotic molecule and shares conserved BH motifs and a. In mammals, when cells suffer apoptotic stimulus, Bak (BCL2 antagonist/killer 1) and Bax (BCL2-associated X, apoptosis regulator) are activated and oligomerize on the mitochondrial outer membrane (MOM) to permeabilize the MOM by forming pores [6]. Bcl-xL is present on the MOM to prevent the oligomerization of proapoptotic molecules (Bax/Bak) and control the balance of the mitochondrial membrane potential (MMP) [7].

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