PUBLICATION
CRISPR/Cas9-mediated knockout of tnf-α1 in zebrafish reduces disease resistance after Edwardsiella piscicida bacterial infection
- Authors
- Nadarajapillai, K., Jung, S., Sellaththurai, S., Ganeshalingam, S., Kim, M.J., Lee, J.
- ID
- ZDB-PUB-231202-5
- Date
- 2023
- Source
- Fish & shellfish immunology 144: 109249 (Journal)
- Registered Authors
- Keywords
- CRISPR/Cas9, E. piscicida, Knock-out, Zebrafish: tnf- α1
- MeSH Terms
-
- Animals
- Bacterial Proteins/genetics
- CRISPR-Cas Systems
- Cytokines/genetics
- Disease Resistance/genetics
- Edwardsiella*/physiology
- Enterobacteriaceae Infections*
- Fish Diseases*
- Tumor Necrosis Factor-alpha/genetics
- Tumor Necrosis Factor-alpha/metabolism
- Zebrafish
- PubMed
- 38040136 Full text @ Fish Shellfish Immunol.
Citation
Nadarajapillai, K., Jung, S., Sellaththurai, S., Ganeshalingam, S., Kim, M.J., Lee, J. (2023) CRISPR/Cas9-mediated knockout of tnf-α1 in zebrafish reduces disease resistance after Edwardsiella piscicida bacterial infection. Fish & shellfish immunology. 144:109249.
Abstract
Tumor necrosis factor (TNF) is an important cytokine involved in immune responses to bacterial infections in vertebrates, including fish. Although Tnf-α is a well-studied cytokine, there are contradictory findings about Tnf-α function following bacterial infection. In this study, we analyzed the expression and function of the Tnf-α-type I isoform (Tnf-α1) in zebrafish by knockout experiments using the CRISPR/Cas9 gene-editing tool. The open reading frame of tnf-α1 encodes a 25.82 kDa protein with 234 amino acids (aa). The expression of tnf-α1 in the early stages of zebrafish was observed from the 2-cell stage. Adult zebrafish spleens showed the highest expression of tnf-α1. To evaluate the function of Tnf-α1, an 8 bp deletion in the target region, resulting in a short truncated protein of 55 aa, was used to create the tnf-α1 knockout mutant. The pattern of downstream gene expression in 7-day larvae in wild-type (WT) and tnf-α1 knockout fish was examined. We also verified the fish mortality rate after Edwardsiella piscicida challenge and found that it was much higher in tnf-α1 knockout fish than in WT fish. Additionally, downstream gene expression analyses after E. piscicida exposure revealed a distinct expression pattern in tnf-α1 knockout fish compared to that in WT fish. Overall, our study using tnf-α1 deletion in zebrafish confirmed that Tnf-α1 is critical for immune regulation during bacterial infection.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping