Molecular cloning, expression analysis, and functional characterization of an interleukin-15 like gene in common carp (Cyprinus carpio L.)

Front Immunol. 2024 Nov 19:15:1502847. doi: 10.3389/fimmu.2024.1502847. eCollection 2024.

Abstract

Interleukin-15 (IL-15) is a crucial cytokine involved in immune system regulation, which is produced by various cell types, including dendritic cells, monocytes, and macrophages. IL-15 plays a key role in the proliferation and activation of natural killer (NK) cells, CD8+ T cells, and memory CD8+ T cells, supporting their survival and enhancing their effector functions. Although IL-15 homologues in fish have been identified, their functions remain poorly understood. In this study, we cloned and investigated the bioactivities of an IL-15 homologue, referred to as IL-15 like (CcIL-15L), in common carp (Cyprinus carpio L.). An expression pattern analysis revealed that CcIL-15L was constitutively expressed in all examined tissues of healthy common carp, with the highest expression level observed in the intestine. Additionally, CcIL-15L expression was significantly up-regulated in the head kidney, spleen, gills, and intestine following Aeromonas hydrophila infection. In vitro, the recombinant protein CcIL-15L can significantly up-regulated the gene expression levels of pro-inflammatory cytokines (IL-1β, IL-6, IFN-γ, and TNF-α) and NK cell activation (perforin and Eomesa). We constructed a 3×FLAG eukaryotic expression vector and successfully expressed it in common carp by intramuscular injection. Additionally, the heterologous CcIL-15L protein was successfully overexpressed in vivo, and immune-related genes including CD4-1, CD8β2, TNF-α, and IgM showed significant induction in the head kidney and spleen. Furthermore, CcIL-15L overexpression reduced the bacterial loads after 24 h post-A. hydrophila infection in the liver, spleen, and kidney. Phagocytic and chemotaxis assays showed that rCcIL-15L could promoted the phagocytosis and chemotactic abilities of common carp HKLs. Our study provides a new perspective on the role for CcIL-15L in immunological functions in common carp.

Keywords: Cyprinus carpio; bioactivity analysis; chemotaxis; immune response; interleukin-15L; leukocytes; phagocytosis.

MeSH terms

  • Aeromonas hydrophila* / immunology
  • Animals
  • Carps* / genetics
  • Carps* / immunology
  • Cloning, Molecular*
  • Cytokines / genetics
  • Cytokines / metabolism
  • Fish Diseases* / genetics
  • Fish Diseases* / immunology
  • Fish Proteins* / genetics
  • Fish Proteins* / immunology
  • Fish Proteins* / metabolism
  • Gene Expression Profiling
  • Gene Expression Regulation
  • Gram-Negative Bacterial Infections / immunology
  • Gram-Negative Bacterial Infections / veterinary
  • Interleukin-15* / genetics
  • Interleukin-15* / immunology
  • Interleukin-15* / metabolism
  • Killer Cells, Natural / immunology
  • Killer Cells, Natural / metabolism
  • Phylogeny

Substances

  • Interleukin-15
  • Fish Proteins
  • Cytokines

Grants and funding

The author(s) declare financial support was received for the research, authorship, and/or publication of this article. This work was sponsored by the National Natural Science Foundation of China (Project No. 32002427), Henan Provincial Science and Technology Tackling Plan Project (Project No. 242102110070) and Henan Province Modern Agricultural Industrial Technology System Construction Project (Project No. HARS-22-16-T).