Identification and Characterization of Immune-Associated MicroRNAs in Silver Carp (Hypophthalmichthys molitrix) Responding to Aeromonas veronii and LPS Stimulation

Abstract

The ubiquitous Gram-negative bacterial pathogen Aeromonas veronii (A. veronii) can easily cause inflammatory reactions in aquatic organisms, resulting in high mortality and huge economic losses. MicroRNAs (miRNAs) participate in immune regulation and have certain conserved properties. MiRNAs are involved in the immune responses of a variety of teleost fish infected with bacteria, whereas there is no related report in silver carp (Hypophthalmichthys molitrix). Therefore, we identified the expression profiles of miRNA in silver carp stimulated by A. veronii and LPS. Among them, the quantity of differentially expressed miRNAs (DEmiRNAs) obtained in the silver carp challenge group was 73 (A. veronii) and 90 (LPS). The GO enrichment and analysis of KEGG pathways have shown that the predicted target genes are mainly associated with lipid metabolism and the immune response in silver carp. This indicates the possibility that miRNAs play a role in regulating immune-related pathways. In addition, a total of eight DEmiRNAs validated the accuracy of the sequencing result via quantitative real-time PCR (qRT-PCR). Finally, we selected the silver carp head kidney macrophage cells (HKCs) as model cells and proved that miR-30b-5p can regulate the inflammatory response in silver carp HKCs. This study lays the foundation for exploring miRNA regulation in silver carp during pathogenic bacterial infection. In addition, it provides a reference for the future development of non-coding RNA antibacterial drugs.

Poly (I:C)-Induced microRNA-30b-5p Negatively Regulates the JAK/STAT Signaling Pathway to Mediate the Antiviral Immune Response in Silver Carp (Hypophthalmichthys molitrix) via Targeting CRFB5

“…With the development of the research, the changes in the miR-30 family’s immune response during environmental stress and the development of teleost fish have been reported [ 26 ]. Additionally, miR-30b-5p plays a broad regulatory role in Gram-negative bacterial infections in silver carp according to our earlier studies [ 27 ]. However, there are few reports on the antiviral effect of the miR-30 family in teleost fish, and its exact role in the innate immunity is still unknown to a great extent.…”

“…Total RNA was extracted using Trizol (Sangon, Shanghai, China) following the manufacturer’s instructions. As previously described [ 27 ], the quality and concentration of RNA were assessed by employing a bioanalyzer (Agilent Bioanalyzer 2100, Pleasanton, CA, USA), and samples with RNA Intensity Number (RIN) values greater than 8 were selected for subsequent experiments. To evaluate the mRNA and mature miR-30b-5p expression, reverse transcription was used to convert RNA (unified 500 ng) to cDNA using the miRNA 1st-Strand cDNA Synthesis Kit (by stem-loop) (Vazyme, Nanjing, China) and the PrimeScript™ RT reagent Kit with gDNA Eraser (Takara, Tokyo, Japan), respectively.…”

Downregulation of miR-1388 Regulates the Expression of Antiviral Genes via Tumor Necrosis Factor Receptor (TNFR)-Associated Factor 3 Targeting Following poly(I:C) Stimulation in Silver Carp (Hypophthalmichthys molitrix)

“…Nevertheless, our comprehension of the regulatory mechanisms governing miRNAs in fish remains limited, impeding further advancements in this field of study. In previous investigations, our research team discovered the involvement of miR-1388 in regulating the innate immunity of silver carp [ 25 ]. In the present study, we aimed to elucidate the specific role played by miR-1388 in poly(I:C) stimulation, thereby furnishing a solid theoretical foundation for understanding the intricate network of miRNA-mediated host–pathogen interactions.…”