Characterization of Japanese flounder (Paralichthys olivaceus) Caspase1 involved in extracellular ATP-mediated immune signaling in fish

Abstract

Caspase1 is a member of inflammatory Caspases that play important roles in the innate immune system. Although several teleost caspase1 genes have been identified, their partner proteins and implication in extracellular ATP-mediated immune signaling in fish are still very limited. Here we identified and characterized a caspase1 gene, named JfCaspase1, from Japanese flounder Paralichthys olivaceus. JfCaspase1 mRNA was constitutively expressed in all examined normal tissues with high expression in skin and gills and moderate expression in the enriched Japanese flounder head kidney macrophages (HKMs) and peripheral blood leukocytes (PBLs). JfCaspase1 was initially down-regulated but significantly up-regulated at the later stage upon LPS and poly(I:C) challenges in the HKMs. JfCaspase1 was also up-regulated in the Japanese flounder immune-related tissues including head kidney, gill and spleen by bacterial challenge with Edwardsiella tarda. JfCaspase1 protein is comprised of 384 amino acid residues with a calculated molecular mass of 43.75 kDa and is phylogenetically close to fish Caspase1 proteins. JfCaspase1 was co-immunopercipitated with Japanese flounder apoptosis-associated speck-like protein (ASC) when co-expressed in HeLa cells, suggesting that there is a potential interaction between the two proteins. In addition, we showed that extracellular ATP, a potent signaling molecule in activating innate immune response, rapidly up-regulates JfCaspase1 expression and enhances its enzymatic activity both in the HKMs and PBLs. Our findings indicated that the inflammatory JfCaspase1 interacted with ASC protein is implicated in the extracellular ATP-mediated immune signaling in fish.