Characterization of orange-spotted grouper (Epinephelus coioides) ASC and caspase-1 involved in extracellular ATP-mediated immune signaling in fish

Xin Zhang; Zetian Liu; Chen Li; Ya Zhang; Liqun Wang; Jingguang Wei; Qiwei Qin

doi:10.1016/j.fsi.2019.12.023

Characterization of orange-spotted grouper (Epinephelus coioides) ASC and caspase-1 involved in extracellular ATP-mediated immune signaling in fish

Fish Shellfish Immunol. 2020 Feb:97:58-71. doi: 10.1016/j.fsi.2019.12.023. Epub 2019 Dec 16.

Authors

Xin Zhang¹, Zetian Liu¹, Chen Li¹, Ya Zhang¹, Liqun Wang¹, Jingguang Wei², Qiwei Qin³

Affiliations

¹ Joint Laboratory of Guangdong Province and Hong Kong Region on Marine Bioresource Conservation and Exploitation, College of Marine Sciences, South China Agricultural University, Guangzhou, 510642, China.
² Joint Laboratory of Guangdong Province and Hong Kong Region on Marine Bioresource Conservation and Exploitation, College of Marine Sciences, South China Agricultural University, Guangzhou, 510642, China. Electronic address: weijg@scau.edu.cn.
³ Joint Laboratory of Guangdong Province and Hong Kong Region on Marine Bioresource Conservation and Exploitation, College of Marine Sciences, South China Agricultural University, Guangzhou, 510642, China; Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao, 266000, China. Electronic address: qinqw@scau.edu.cn.

PMID: 31837409
DOI: 10.1016/j.fsi.2019.12.023

Abstract

Apoptosis-associated speck-like protein containing a CARD domain (ASC) is a critical adaptor molecule in multiple inflammasome protein complexes that mediate inflammation and host defense. Caspase-1 is a member of inflammatory caspases that play important roles in the innate immune system. However, few studies have been performed in lower vertebrates such as teleosts and implications of extracellular ATP-mediated immune signalling in fish. Here we identified and characterized novel ASC and caspase-1 genes (namely EcASC and EcCaspase-1) from the orange-spotted grouper (Epinephelus coioides). EcASC and EcCaspase-1 encode 204- and 388-aa proteins which shared 55.34% and 72.89% identity with those in Siniperca chuatsi and Perca flavescens, respectively. EcASC contained a PYRIN domain (aa 5-82) and CARD domain (aa 107-201). EcCaspase1 contained a CARD domain (aa 1-88) and a CASc domain (aa 127-376). Both EcASC and EcCaspase-1 were distributed in all tissues tested in the healthy grouper. The expression of EcASC and EcCaspase-1 was significantly upregulated in response to ATP infection. Subcellular localization analysis showed that EcCaspase-1 exhibited a clear distribution in both cytoplasm and nucleus. In contrast, EcASC was observed in the cytoplasm as speck-like structures, which are called "pyroptosomes". EcCaspase-1 co-localized with the spot-like protein (EcASC). Overexpression of EcASC and EcCaspase-1 inhibited NF-κB activation and promoted P53 activation in grouper spleen (GS) cells. Extracellular ATP was an effective signaling molecule that activates the innate immune response, rapidly upregulating the expression of EcASC and EcCaspase1, and enhancing their promotion of proinflammatory cytokine expression in GS cells. Both EcASC and EcCaspase-1 promoted ATP-induced apoptosis. Our results suggested that the interactions of inflammatory EcCaspase-1 with EcASC proteins were associated with extracellular ATP-mediated immune signaling in fish.

Keywords: ASC; ATP; Caspase-1; Grouper; Immune; Interaction.

MeSH terms

Adenosine Triphosphate / immunology*
Animals
Bass / genetics
Bass / immunology*
CARD Signaling Adaptor Proteins / genetics
CARD Signaling Adaptor Proteins / immunology*
Caspase 1 / genetics
Caspase 1 / immunology*
Cloning, Molecular
Fish Diseases / immunology
Gene Expression Regulation
Immunity, Innate*
Phylogeny
Signal Transduction / immunology*

Substances

CARD Signaling Adaptor Proteins
Adenosine Triphosphate
Caspase 1