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Using Zebrafish Models of Human Influenza A Virus Infections to Screen Antiviral Drugs and Characterize Host Immune Cell Responses
Published on: January 20, 2017
The discrepancy function of NLRC5 isoforms in antiviral and antibacterial immune responses
Lu Cao1, Xiao Man Wu2, Yi Wei Hu2
1State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan, Hubei Province 430072, China; University of Chinese Academy of Sciences, Beijing, 100039, China.
Abstract:
NOD-like receptors (NLRs) are a family of intracellular pattern recognition receptors (PRRs) that play critical roles in innate immunity against pathogens infection. NLRC5, the largest member of NLR family, has been characterized as a regulator of innate immunity and MHC class I expression. Alternative splicing of NLRC5 is only reported in human and zebrafish. However, the function of NLRC5 isoforms in the innate immune responses remains unknown. In the present study, we report the functional characterization of zfNLRC5a and zfNLRC5d, two splicing isoforms of zebrafish NLRC5. zfNLRC5a and zfNLRC5d are generated by exon skipping, and whose alternative splicing sites exist in the region of LRRs. Fluorescence microscopy showed that zfNLRC5 isoforms were located throughout the entire cell including nuclear staining. The expression of zfNLRC5 isoform was inducible in response to bacterial and viral infections. During SVCV infection, the in vitro and in vivo studies found that zfNLRC5d overexpression increased protection against viral infection; however zfNLRC5a overexpression had no significant effect on antiviral activity. Interestingly, zfNLRC5 isoforms but not zfNLRC5 were involved in transcriptional regulation of TLRs and NF-κB signaling. Overexpression of zfNLRC5 isoforms also contributed to negative regulation of antibacterial immune response, with the decreased expression of nfkbiaa (IκBα). All together, these results firstly demonstrate the function of NLRC5 isoforms in antiviral and antibacterial immune responses both in vitro and in vivo.
Insights
Zebrafish NLRC5 isoforms, zfNLRC5a and zfNLRC5d, regulate innate immunity. zfNLRC5d enhances antiviral defense, while both isoforms impact TLRs, NF-κB signaling, and antibacterial responses.
Area of Science:
- Immunology
- Molecular Biology
- Genetics
Background:
- NOD-like receptors (NLRs) are crucial for innate immunity.
- NLRC5 regulates immunity and MHC class I expression.
- NLRC5 alternative splicing is known in humans and zebrafish, but isoform functions are unclear.
Purpose of the Study:
- To functionally characterize zebrafish NLRC5 (zfnlrc5) splicing isoforms, zfNLRC5a and zfNLRC5d.
- To investigate the role of these isoforms in innate immune responses against viral and bacterial infections.
Main Methods:
- Generated and characterized zfNLRC5a and zfNLRC5d isoforms via exon skipping.
- Utilized fluorescence microscopy for subcellular localization.
- Assessed isoform expression upon bacterial and viral challenge.
- Performed in vitro and in vivo studies to evaluate antiviral and antibacterial activities.
- Analyzed transcriptional regulation of Toll-like receptors (TLRs) and NF-κB signaling pathways.
Main Results:
- zfNLRC5a and zfNLRC5d are generated by alternative splicing in the LRR region.
- Isoforms are expressed throughout the cell, including the nucleus, and are inducible by infection.
- zfNLRC5d overexpression conferred protection against viral infection (SVCV), while zfNLRC5a had no significant effect.
- Both isoforms, but not full-length zfNLRC5, modulated TLRs and NF-κB signaling.
- Overexpression of zfNLRC5 isoforms negatively regulated antibacterial immunity by decreasing nfkbiaa (IκBα) expression.
Conclusions:
- This study provides the first functional characterization of NLRC5 isoforms in zebrafish.
- zfNLRC5d plays a protective role in antiviral immunity.
- NLRC5 isoforms are involved in both antiviral and antibacterial innate immune responses, influencing TLR and NF-κB signaling pathways.
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