Meishan is a well-established local Chinese breed known for its high fecundity, strong immune response and high meat quality. However, the molecular mechanism of immune regulation during the development of Meishan pigs still remains unclear. Here, we performed the transcriptional sequencing of spleen tissues from Meishan pigs at different development stages. In total, 10,268 lncRNAs were identified, including 1254 novel lncRNAs and 9014 known lncRNAs. Time series analysis revealed that genes of the up-regulated module were enriched in pathways associated with transport, immunity, and histone acetylation modifications, while genes of the down-regulated module were enriched in DNA metabolic process and cell cycle. Weighted gene co-expression network analysis (WGCNA) showed the functional linkage between mRNAs and lncRNAs, indicating that lncRNAs are important regulatory elements of mRNAs. Notably, a lncRNA-miRNA-mRNA competing endogenous RNA (ceRNA) network that contained 3 mRNAs (AKT3, CBL and PTK2B), 17 lncRNAs and 67 miRNAs were screened out, which probably plays a critical role in immune regulation of Meishan pigs. Our findings not only revealed the transcriptome profile of spleen development, but also provide novel insights into the mechanism of lncRNA-miRNA-mRNA axis in the immune response in Meishan pigs.

The primary pathogen causing post-weaning diarrhea in piglets is Escherichia coli F18 (E. coli F18), hence it is essential to investigate the mechanism governing E. coli F18 resistance in native pig breeds. Based on the previous RNA-seq results of the duodenum from E. coli F18-resistant and -susceptible Meishan piglets, CXCL11, an important functional gene, was preliminarily screened. In this investigation, in order to further examine the expression regulation mechanism of E. coli F18 in intestinal porcine epithelial cells (IPEC-J2) against E. coli F18 infection, CXCL11 gene expression on IPEC-J2 cells infected by E. coli F18 was detected, which was significantly downregulated (p < 0.01). Secondly, the overexpression on the IPEC-J2 cell line was successfully structured, and a relative quantification method of the PILIN, bacteria enumeration, and immunofluorescence assay indicated that the CXCL11 overexpression significantly reduced the ability of E. coli F18 to interact with IPEC-J2 in vitro. The promoter region of the CXCL11 gene was predicted to contain a CpG island (−619 ~ −380 bp) of which 13 CpG sites in the sequencing region were methylated to varying degrees, and the methylation level of one CPG site (mC-7) positively linked negatively with the expression of the CXCL11 gene (p < 0.05). Meanwhile, a dual luciferase assay detected the mutation of the mC-7 site that significantly inhibited the luciferase activity of the CXCL11 gene promoter (p < 0.01). Transcription factor prediction and expression verification indicated that mC-7 is located in the OSR1-binding domain, and that its expression level is related to E. coli F18 susceptibility. We speculated that methylation modification of the mC-7 site of the CpG island in the promoter region of the CXCL11 gene might inhibit the binding of transcription factor OSR1 with the mC-7 site, and then affect its expression level to regulate the susceptibility to E. coli F18.