Background: As a chronic relapsing disease, probiotic treatment of ulcerative colitis can avoid resistance and side effects of conventional treatment, but its mechanism of action is unknown. This study investigated the therapeutic effect and mechanism of action of potential probiotics Lactiplantibacillus plantarum HNU082 (Lp082) on dextran sulfate sodium-induced colitis in mice.Results: The results showed that the intake of Lp082 increased body weight, water intake, food intake, colon length, and reduced disease activity index, immune organ index, inflammatory markers, and histopathological score. Lp082 strengthened the biological barrier by improving the diversity and composition of gut microbiota, increasing the short chain fatty acids (SCFAs) producing bacteria and SCFAs content, and enhancing the metabolic pathway of SCFAs. Lp082 improved the chemical barrier by increasing the goblet cells and mucin-2 and decreasing the ICAM-1 and VCAM. Lp082 enhanced the mechanical barrier by increasing the mRNA expression of ZO-1, ZO-2, occludin and decreasing claudin-1 and claudin-2. Lp082 strengthened the immune barrier by reducing the content of IL-1β, IL-6, TNF-α, MPO, IFN-γ and increasing the IL-10, TGF-β1, and TGF-β2. Lp082 also affected gene expression in mice, including that it inhibited the gene expression of IL-1β, IL-1α, Ereg, Cdc7, and others. Thus inhibited the biological processes such as IL-1β production, IL-6 production, acute inflammatory response, and promoted the expression of genes Isg15, Prg2, Abcc2, thus boosting the biological processes such as IL-10 production and drug transport. The differentially expressed genes in DSS group were primarily enriched in the disease-related KEGG pathways but were mainly enriched in the KEGG pathway associated with platelet activated autophagy in animals in Lp082 group. Lp082 also inhibited the mRNA expression of NF-κB1, NF-κB2, COX-2, iNOS, Toll-4, and others, thus suppressing the NK-KB pathways.Conclusions: Shotgun metagenomic and transcriptome analysis revealed that Lp082 could relieve ulcerative colitis, protect the intestinal mucosal barrier through SCFAs, regulate gene expression and signaling pathways. The study provides new clues to reveal the complex mechanisms of the gut barriers and genes in relieving ulcerative colitis.