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PcTRIM23 knockdown correlates with increased WSSV replication, hepatopancreatic apoptosis, and ROS accumulation in
Chao Peng1, Liye Shao1, Ronghua Wang1
1College of Life and Environmental Science, Hunan University of Arts and Science, Changde, 415000, China; Hunan Provincial Key Laboratory for Molecular Immunity Technology of Aquatic Animal Diseases, Changde, 415000, China; Hunan Provincial Key Laboratory for Health Aquaculture and Product Processing in Dongting Lake Area, Changde, 415000, China.
Abstract:
This study aimed to clone and identify the TRIM23 gene (designated as PcTRIM23) from Procambarus clarkii and investigate its immune function during white spot syndrome virus (WSSV) infection. The open reading frame (ORF) of PcTRIM23 was 1719 bp in length, encoding a 572-amino acid protein that contains a RING finger domain, two B-Box domains, a B-Box C-terminal domain (BBC) at the N-terminus and an ADP-ribosylation factor (ARF) domain at the C-terminus. PcTRIM23 exhibited the highest expression levels in the hepatopancreas and gills of P. clarkii, and its expression was significantly upregulated at 6-24 h following WSSV infection. After knocking down PcTRIM23 expression via RNA interference (RNAi), the copy number of WSSV and the transcriptional level of VP28 were markedly increased, showing a correlation between reduced PcTRIM23 expression and enhanced viral replication. Further studies revealed that PcTRIM23 knockdown induced hepatopancreatic cell apoptosis, as evidenced by the increased number of TUNEL-positive cells, mitochondrial vacuolization, the up-regulation of apoptosis-related genes including Cytochrome C (Cyt-C) and caspase-3, and the down-regulation of B-cell lymphoma/leukemia 2 (Bcl-2). In addition, PcTRIM23 knockdown led to the accumulation of reactive oxygen species (ROS) and the decreased expression of antioxidant-related genes such as Nrf2, Cu/Zn-SOD and CAT. In conclusion, our results suggest that PcTRIM23 is associated with a protective role against WSSV infection in P. clarkii, correlating with reduced cell apoptosis and enhanced antioxidant response. However, further rescue experiments are needed to establish a direct causal relationship. This study provides the first functional characterization of crayfish TRIM23 in antiviral immunity and lays a foundation for further investigation of its causal regulatory mechanisms.
Insights
The TRIM23 gene in crayfish (PcTRIM23) plays a crucial role in antiviral immunity against white spot syndrome virus (WSSV). Its knockdown increases viral replication, apoptosis, and oxidative stress, indicating a protective function.
Area of Science:
- Aquatic animal health
- Molecular biology
- Immunology
Background:
- White spot syndrome virus (WSSV) is a major pathogen affecting penaeid shrimp and crayfish.
- The tripartite motif-containing protein 23 (TRIM23) is involved in various cellular processes, including immunity.
- Understanding the role of TRIM23 in crayfish immunity is crucial for disease management.
Purpose of the Study:
- To clone and characterize the TRIM23 gene (PcTRIM23) from Procambarus clarkii.
- To investigate the immune function of PcTRIM23 during WSSV infection.
- To explore the molecular mechanisms underlying PcTRIM23's antiviral activity.
Main Methods:
- Gene cloning and sequence analysis of PcTRIM23.
- Quantitative real-time PCR (qRT-PCR) to analyze gene expression.
- RNA interference (RNAi) to knock down PcTRIM23 expression.
- TUNEL assay and Western blotting to assess apoptosis.
- Measurement of reactive oxygen species (ROS) and antioxidant gene expression.
Main Results:
- PcTRIM23 was cloned and characterized, showing typical TRIM23 domain structures.
- PcTRIM23 expression was upregulated in response to WSSV infection.
- Knockdown of PcTRIM23 led to increased WSSV replication, enhanced hepatopancreatic cell apoptosis, elevated ROS levels, and suppressed antioxidant responses.
- PcTRIM23 knockdown resulted in increased viral copy numbers and VP28 transcript levels.
Conclusions:
- PcTRIM23 plays a significant role in the antiviral immunity of Procambarus clarkii against WSSV.
- PcTRIM23 confers protection by inhibiting viral replication, reducing apoptosis, and enhancing antioxidant capacity.
- Further research is needed to elucidate the direct causal mechanisms of PcTRIM23's antiviral activity.
