Signaling pathways of duck RIG-I in gene-edited DF1 chicken cells

Hyeong Ju Ryu; Si Eun Kim; Ki Hyun Kim; Ji Hoon Park; Heesu Jeong; Ismail Shaleh; Tae Sub Park

doi:10.1016/j.psj.2024.104739

Signaling pathways of duck RIG-I in gene-edited DF1 chicken cells

Poult Sci. 2024 Dec 25;104(2):104739. doi: 10.1016/j.psj.2024.104739. Online ahead of print.

Authors

Hyeong Ju Ryu¹, Si Eun Kim², Ki Hyun Kim³, Ji Hoon Park⁴, Heesu Jeong⁵, Ismail Shaleh⁶, Tae Sub Park⁷

Affiliations

¹ Graduate School of International Agricultural Technology and Institute of Green-Bio Science and Technology, Seoul National University, Pyeongchang-gun, Gangwon-do 25354, South Korea. Electronic address: rue970430@snu.ac.kr.
² Graduate School of International Agricultural Technology and Institute of Green-Bio Science and Technology, Seoul National University, Pyeongchang-gun, Gangwon-do 25354, South Korea. Electronic address: kse517@snu.ac.kr.
³ Graduate School of International Agricultural Technology and Institute of Green-Bio Science and Technology, Seoul National University, Pyeongchang-gun, Gangwon-do 25354, South Korea. Electronic address: wldms3748@snu.ac.kr.
⁴ Graduate School of International Agricultural Technology and Institute of Green-Bio Science and Technology, Seoul National University, Pyeongchang-gun, Gangwon-do 25354, South Korea. Electronic address: dasom8857166@snu.ac.kr.
⁵ Graduate School of International Agricultural Technology and Institute of Green-Bio Science and Technology, Seoul National University, Pyeongchang-gun, Gangwon-do 25354, South Korea. Electronic address: heesu0302@snu.ac.kr.
⁶ Graduate School of International Agricultural Technology and Institute of Green-Bio Science and Technology, Seoul National University, Pyeongchang-gun, Gangwon-do 25354, South Korea; Department of Biology, Faculty of Mathematics and Natural Sciences, IPB University, Bogor 16680, Indonesia. Electronic address: ismailshaleh@snu.ac.kr.
⁷ Graduate School of International Agricultural Technology and Institute of Green-Bio Science and Technology, Seoul National University, Pyeongchang-gun, Gangwon-do 25354, South Korea; Institute of Green-Bio Science and Technology, Seoul National University, Pyeongchang-gun, Gangwon-do 25354, South Korea. Electronic address: taesubpark@snu.ac.kr.

PMID: 39733732
DOI: 10.1016/j.psj.2024.104739

Abstract

Retinoic acid inducible gene I (RIG-I) is an innate immune RNA sensor which can detect viral infection such as influenza viruses. Duck but not chicken has an RIG-I gene. However, the immune responses could be induced in chicken cells by transferring the duck RIG-I transgene. However, effects of other pathogen-recognition receptor (PRR) genes such as Toll-like receptor 3 (TLR3) and melanoma differentiation-associated protein 5 (MDA5) could not be ruled out. In this study, we knocked out TLR3 and MDA5 genes using gene-editing protocol, and stably transferred the duck RIG-I transgene into TLR3/MDA5 double knockout (KO) chicken DF1 cells. We investigated the antiviral responses induced by duck RIG-I in chicken cells. Duck RIG-I induced the expression of interferon-stimulated genes (ISGs) and inflammatory cytokines such as interferon regulatory factor 7 (IRF7), interferon β (IFNβ), Mx1, and protein kinase R1 (PKR1) after treatment with polyinosinic: polycytidylic acid (poly I:C) in TLR3/MDA5 double KO DF1 cells. Additionally, to examine the duck RIG-I signaling cascade, we knocked out mitochondrial antiviral-signaling protein (MAVS), which encodes an antiviral signaling factor in innate immunity. Duck RIG-I in TLR3/MDA5/MAVS triple KO DF1 cells did not activate downstream expression of ISGs. Finally, to analyze the global signaling pathways of duck RIG-I in chicken cells, next-generation sequencing of total mRNAs with and without poly I:C treatment was conducted. In conclusion, duck RIG-I mediated antiviral signaling independently of TLR3 and MDA5, and MAVS induced and stimulated ISGs by duck RIG-I in chicken cells.

Keywords: Chicken; Duck RIG-I; Immune response; PRRs; Signal pathways.