MicroRNA profiling and functional validation reveal Poly I:C-responsive miRNAs potentially regulate immune pathways in the giant freshwater prawn, Macrobrachium rosenbergii

Abstract

MicroRNAs (miRNAs) are important regulators of gene expression in immune responses. In this study, we examined miRNA expression in M. rosenbergii following stimulation with polyinosinic:polycytidylic acid (Poly I:C), a synthetic analog of viral double-stranded RNA (dsRNA). Small RNA libraries were prepared from control and Poly I:C-injected prawns and sequenced using high-throughput sequencing. A total of 137 differentially expressed miRNAs (DEMs) were identified. Target gene prediction and enrichment analysis showed that these miRNAs are involved in several immune-related pathways, including the ubiquitin–proteasome pathway (UPP), endocytosis, autophagy, and transforming growth factor-β (TGF-β) signaling pathway. Time-course analysis revealed distinct expression patterns of selected miRNAs after Poly I:C injection. Dual-luciferase reporter assays confirmed that mro-miR-7689-3p and mro-miR-9014 directly target nejire ( nej ) and autophagy-related gene 9 ( atg9 ), respectively. These results provide new insights into the role of miRNAs in the antiviral immunity of M. rosenbergii and may support the development of RNA-based strategies for disease control in prawn aquaculture.