Journal of fish pathology (한국어병학회지)

The Korean Society of Fish Pathology (한국어병학회)
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Evaluation of availability of various internal ribosome entry sites (IRESs) for bicistronic expression in different fish cell lines

  • Kyung Min Lee (Department of Aquatic Life Medicine, Pukyong National University) ;
  • Ki Hong Kim (Department of Aquatic Life Medicine, Pukyong National University)
  • Received : 2023.06.01
  • Accepted : 2023.06.12
  • Published : 2023.06.30
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Abstract

The internal ribosome entry site (IRES) elements from various RNA viruses are widely used to express proteins in bicistronic or multicistronic ways in mammalian cells. However, research on the application of IRESs in fish cells has been poorly conducted. In this study, to evaluate the availability of various viral IRESs in fish cells based on a bicistronic vector system, the translation activity of IRESs from viruses of invertebrates (aphid lethal paralysis virus ALPV; cricket paralysis virus, CrPV; and Plautia stali intestine virus, PSIV), of mammals (encephalomyocarditis virus, EMCV), and of fish (infectious pancreatic necrosis virus, IPNV; marine birnavirus, MABV; and snakehead retrovirus, SnRV) was analyzed in various fish cell lines originated from salmonid (chinook salmon embryonic cells, CHSE-214), cyprinid (Epithelioma papulosum cyprinid, EPC), and flatfish (Hirame natural embryonic cells, HINAE). Translation mediated by EMCV IRES was shown in all cell lines, but the activity was weak in CHSE-214 and HINAE cells while high activity was shown in EPC cells, suggesting that although EMCV IRES can be broadly used in various fish cells, the translational activity can be varied according to different cell lines. PSIV IRES showed moderate activity in EPC cells and low activity in CHSE-214 and HINAE cells. CrPV IRES showed weak activity in EPC and HINAE cells and no activity in CHSE-214 cells. In the case of ALPV IRES, green fluorescence was observed only in EPC cells but the activity was very weak. These results suggest that dicistroviral IRESs analyzed in this study would not be the best option for protein expression in various fish cells. The translation activity of SnRV IRES in fish cells was first verified in EPC cells, though the activity was very weak. The 5'UTR of MABV segment A showed high translation activities in all examined fish cell lines, suggesting that IRES of MABV can be used to develop multicistronic expression systems or cap-independent RNA-based translation systems in a wide range of fish cells.

Keywords

Acknowledgement

This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Korea government (MSIT) (No. NRF-2021M3E5E60 26104).

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