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Published on: March 24, 2017
Nlrc5 ablation interferes with MHC-I gene expression and immune cell migration
K A S N Shanaka1, K P Madushani2, Myoung-Jin Kim3
1Department of Marine Life Sciences & Center for Genomic Selection in Korean Aquaculture, Jeju National University, Jeju, 63243, Republic of Korea; Marine Life Research Institute, Jeju National University, Jeju, 63333, Republic of Korea.
Abstract:
Nucleotide-binding domain and leucine-rich repeat-containing receptors (NLRs) have complex and diverse functions. Studies in mammalian models have indicated a clear role of NLRC5 in the regulation of MHC-I gene expression. The nlrc5 KO zebrafish model was generated using CRISPR/Cas9 technology to understand its function with respect to VHSV. The VHSV infection experiment demonstrated higher mortality in nlrc5 KO fish with higher VHSV copy numbers. Virus-infected pathology, such as tissue swelling, hemorrhage, and eye bulging, was significantly higher in the nlrc5 KO fish. Expressional analysis of ifn system genes indicated the significant upregulation of ifnφ1. Gene expression analysis during VHSV infection indicated a significant downregulation of MHC-I genes in the nlrc5 KO model compared to the wild-type (WT). Owing to the impact of MHC-I, the viral copy number may increase with higher mortality under VHSV infections. For neutrophil migration analysis, a nlrc5 KO and a neutrophil-labeled model were developed (nlrc5-/- Tg(mpx:mcherry)). Injury was generated in the caudal fin, and the injury site was stimulated with poly I: C. The number of neutrophils were reduced in the nlrc5 KO fish. Levels of critical cytokines responsible for neutrophil migration were significantly reduced in nlrc5 KO fish during VHSV infection. The data from the current study in zebrafish reconfirm the observations in mammalian models, and neutrophil migration analysis suggests that Nlrc5 may be associated with inflammatory activation in the presence of VHSV.
Insights
The NLRC5 gene is crucial for immune response in zebrafish, as its absence leads to increased mortality and severe pathology during viral infections. This highlights NLRC5
Area of Science:
- Immunology
- Genetics
- Zebrafish models
Background:
- Nucleotide-binding domain and leucine-rich repeat-containing receptors (NLRs) have complex immune functions.
- NLRC5 is known to regulate MHC-I gene expression in mammals.
- The role of NLRC5 in viral infections, specifically VHSV, is not well understood.
Purpose of the Study:
- To investigate the function of NLRC5 in zebrafish against VHSV infection.
- To analyze the impact of NLRC5 on MHC-I gene expression and immune cell migration.
Main Methods:
- CRISPR/Cas9 technology was used to generate an nlrc5 knockout (KO) zebrafish model.
- VHSV infection experiments were conducted to assess mortality and viral load.
- Gene expression analysis of interferon (IFN) and MHC-I genes was performed.
- Neutrophil migration was analyzed in nlrc5 KO zebrafish under inflammatory conditions.
Main Results:
- nlrc5 KO zebrafish exhibited higher mortality rates and increased VHSV copy numbers.
- Significant upregulation of ifnφ1 and downregulation of MHC-I genes were observed in nlrc5 KO fish.
- Pathological signs of VHSV infection were more severe in nlrc5 KO fish.
- Neutrophil migration and levels of related cytokines were reduced in nlrc5 KO fish.
Conclusions:
- NLRC5 plays a critical role in the zebrafish immune response to VHSV infection.
- The absence of NLRC5 impairs MHC-I expression and neutrophil-mediated immunity.
- Zebrafish models are valuable for studying NLR functions and their implications in viral pathogenesis.
