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GSIV serine/threonine kinase can induce apoptotic cell death via p53 and pro-apoptotic gene Bax upregulation in fish cells

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Abstract

Previous studies have shown that GSIV induces apoptotic cell death through upregulation of the pro-apoptotic genes Bax and Bak in Grouper fin cells (GF-1 cells). However, the role of viral genome-encoded protein(s) in this death process remains unknown. In this study, we demonstrated that the Giant seaperch iridovirus (GSIV) genome encoded a serine/threonine kinase (ST kinase) protein, and induced apoptotic cell death via a p53-mediated Bax upregulation approach and a downregulation of Bcl-2 in fish cells. The ST kinase expression profile was identified through Western blot analyses, which indicated that expression started at day 1 h post-infection (PI), increased up to day 3, and then decreased by day 5 PI. This profile indicated the role of ST kinase expression during the early and middle phases of viral replication. We then cloned the ST kinase gene and tested its function in fish cells. The ST kinase was transiently expressed and used to investigate possible novel protein functions. The transient expression of ST kinase in GF-1 cells resulted in apoptotic cell features, as revealed with Terminal deoxynucleotidyl transferase biotin-dUTP nick-end labeling (TUNEL) assays and Hoechst 33258 staining at 24 h (37 %) and 48 h post-transfection (PT) (49 %). Then, through studies on the mechanism of cell death, we found that ST kinase overexpression could upregulate the anti-stress gene p53 and the pro-apoptotic gene Bax at 48 h PT. Interestingly, this upregulation of p53 and Bax also correlated to alterations in the mitochondria function that induced loss of mitochondrial membrane potential (MMP) and activated the initiator caspase-9 and the effector caspase-3 in the downstream. Moreover, when the p53-dependent transcriptional downstream gene was blocked by a specific transcriptional inhibitor, it was found that pifithrin-α not only reduced Bax expression, but also averted cell death in GF-1 cells during the ST kinase overexpression. Taken altogether, these results suggested that aquatic GSIV ST kinase could induce apoptosis via upregulation of p53 and Bax expression, resulting in mitochondrial disruption, which activated a downstream caspases-mediated cell death pathway.

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Acknowledgments

The authors are grateful to Dr. S. C. Shi (Institute of Zoology and Department of Life Science, National Taiwan University, 1, Sec., 4, Roosevelt Rd., Taipei 10617, Taiwan, ROC) for providing the GF-1 cell line and to Dr. C. M. Wen (Department of Life Sciences, National University of Kaohsiung, Kaohsiung 811, Taiwan, ROC) for providing the GSIV K1 strain. This research was supported by Grants (NSC-99-2321-B-006-010-MY3; MOST 104-2313-B-006-003) awarded to Dr. Jainn-Ruey Hong from the National Science Council, Taiwan, Republic of China.

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Authors and Affiliations

  1. Laboratory of Molecular Virology and Biotechnology, Institute of Biotechnology, Department of Biotechnology and Bioindustry Sciences, National Cheng Kung University, No 1. University Road, Tainan City, 701, Taiwan, ROC

    Latif Reshi & Jiann-Ruey Hong

  2. Department of Life Sciences, College of Bioscience and Biotechnology, National Cheng Kung University, No. 1. University Road, Tainan City, 701, Taiwan, ROC

    Latif Reshi & Hao-Ven Wang

  3. Laboratory Department of Food Science and Technology, Chin Nan University of Pharmacy and Science, Tainan, 717, Taiwan, ROC

    Horng-Cherng Wu

  4. Laboratory of Marine Molecular Biology and Biotechnology, Institute of Cellular and Organismic Biology, Academia Sinica, Nankang, Taipei, 115, Taiwan, ROC

    Jen-Leih Wu

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  1. Latif Reshi
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Correspondence to Jiann-Ruey Hong.

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Reshi, L., Wu, HC., Wu, JL. et al. GSIV serine/threonine kinase can induce apoptotic cell death via p53 and pro-apoptotic gene Bax upregulation in fish cells. Apoptosis 21, 443–458 (2016). https://doi.org/10.1007/s10495-016-1219-4

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