Comprehensive transcriptome-wide characterisation of hepatopancreas in response to acute low-temperature stress in Macrobrachium rosenbergii

The giant freshwater prawn, Macrobrachium rosenbergii , is an economically important aquaculture species that exhibits intrinsically low-temperature tolerance, rendering it susceptible to low-temperature stress, which impairs survival, growth, and productivity. Although thermal transcriptomics has been studied in aquatic species, ncRNA regulation under acute low-temperature stress remains poorly defined in M. rosenbergii . In this study, we present the first comprehensive whole-transcriptome profiling of the hepatopancreas in M. rosenbergii under acute low-temperature stress. The treatment group was subjected to a cooling regime from 26.0 ± 0.5 °C to 13.5 ± 0.5 °C at a rate of 1.5 °C/h, followed by maintenance at 13.5 ± 0.5 °C for 12-h, whereas the control group was held at a constant 26.0 ± 0.5 °C for 36-h. Sampling occurred at 36-h and 44.3-h after fasting onset in the NHG and LHG, respectively, resulting in an 8.3-h difference. Each condition included three biological replicates derived from independent culture tanks. We performed whole-transcriptome sequencing of the hepatopancreas to profile coding and ncRNA expression under acute low-temperature stress. FDR-controlled analysis identified 439 mRNAs, 23 lncRNAs, and 13 miRNAs, no circRNAs were significant. Exploratory analysis using a relaxed threshold identified 2,250 mRNAs, 80 lncRNAs, 2 circRNAs, and 23 miRNAs. Exploratory results were used only for hypothesis generation. Functional enrichment analysis showed that DEmRNAs were substantially enriched in immune-related pathways and metabolic pathways. Correlation analysis between DElncRNAs and DEmRNAs identified 1,436 mRNAs that were potentially targeted by the 80 DElncRNAs, forming a total of 8,729 lncRNA-mRNA interaction pairs. Integration of miRNA-mRNA and lncRNA-mRNA interactions enabled the construction of a mRNA-bridged putative lncRNA–miRNA–mRNA association network, consisting of 2 miRNAs, 2 mRNAs and 8 lncRNAs, this network comprised 8 lncRNA-mRNA pairs and 4 miRNA-mRNA pairs. This network, generated based on bioinformatic algorithms, remains to be experimentally verified. Within the constructed interaction network, we predicted candidate genes including Acat2 , Alg-2 , Atg18a , eIF3i , Kpc2 , Larp4B , maf-S , Megf8 , mRpL21 , Nmnat , Ptpmeg , Pus10 , Rab7 , Slmap , spartin , Tim23 , Srrm1 , Usp7 , and Wee1 were identified as potentially associated with acute low-temperature adaptation in M. rosenbergii . RT-qPCR on the RNA-seq samples confirmed technical concordance (mRNA R² = 0.8046, lncRNA R² = 0.953 and miRNA R² = 0.9124). Because survival was low (~13%), these profiles mainly represent acute low-temperature-tolerant survivors rather than the general population response. This study provides a comprehensive transcriptomic landscape of coding and ncRNAs in the hepatopancreas of M. rosenbergii under acute low-temperature stress. Our findings suggest that acute low-temperature stress not only alters mRNA expression but also profoundly reshapes the expression patterns and interaction networks of lncRNAs, circRNAs, and miRNAs. It is plausible that acute low‑temperature stress orchestrates coordinated reprogramming of immune, apoptotic, and metabolic pathways through integrated ncRNA‑mediated regulatory networks, and that this process could unveil a complex molecular framework for acute low-temperature adaptation. These results, particularly the candidate genes and regulatory networks identified, provide a resource for future functional studies and may guide selective breeding programs aimed at enhancing acute low-temperature tolerance in this economically important species.