Identification of long non-coding RNA MSTRG.5748.1 and MSTRG.7894.1 from Megalobrama amblycephala and their potential roles in innate immunity

Megalobrama amblycephala is one of the most economically important freshwater fish in China, and the bacterial septicemia caused by Aeromonas hydrophila is a serious threat to the breeding industry of M. amblycephala. Unfortunately, the characterization of long noncoding RNA (lncRNA) in response to A. hydrophila infection has not been performed in M. amblycephala. To better understand the biological significance of lncRNA in the immune system, we identified two lncRNA, named MSTRG.5748.1 and MSTRG.7894.1, as playing critical roles in the antibacterial response of M. amblycephala. After separating the nucleus and cytoplasm of the hepatocytes from M. amblycephala, cellular localization of MSTRG.5748.1 and MSTRG.7894.1 was performed to predict their functions. The results showed that MSTRG.5748.1 was mainly expressed in the nucleus, suggesting that its functions are mostly to regulate the expression of downstream genes through epistasis and transcription. MSTRG.7894.1 existed in both the nucleus and cytoplasm, which indicated that it has many regulatory modes. qPCR analysis showed that MSTRG.5748.1 and MSTRG.7894.1 were expressed in the immune-related organs of M. amblycephala, and significantly changed in the liver after A. hydrophila infection. RNA-seq analysis revealed that differentially expressed genes (DEGs) were mainly enriched in antigen processing and presentation via MHC class I, RIG-I-like receptor (RLR) signaling pathway, and IFN-related pathway, and a large number of pathwayrelated genes were significantly regulated after lncRNA overexpression in muscle cell of M. amblycephala. Overexpression of MSTRG.5748.1 and MSTRG.7894.1 significantly inhibited the expression of STING and IFN, significantly upregulated muscle cell viability, and promoted cell proliferation by targeting STING and IFN.