Campylobacter jejuni (C. jejuni) infections are of increasing importance worldwide. As a typical mucosal pathogen, the interaction of C. jejuni with mucins is a prominent step in the colonisation of mucosal surfaces. Despite recent advances in understanding the interaction between bacterial pathogens and host mucins, the mechanisms of mucin glycosylation during intestinal C. jejuni infection remain largely unclear. This prompted us to identify relevant regulatory networks that are concerted by miRNAs and could play a role in the mucin modification and interaction.
We firstly used a human intestinal in vitro model, in which we observed altered transcription of MUC2 and TFF3 upon C. jejuni NCTC 11168 infection. Using a combined approach consisting of in silico analysis together with in vitro expression analysis, we identified the conserved miRNAs miR-125a-5p and miR-615-3p associated with MUC2 and TFF3. Further pathway analyses showed that both miRNAs appear to regulate glycosyltransferases, which are related to the KEGG pathway ‘Mucin type O-glycan biosynthesis’. To validate the proposed interactions, we applied an in vivo approach utilising a well-established secondary abiotic IL-10−/− mouse model for infection with C. jejuni 81-176. In colonic tissue samples, we confirmed infection-dependent aberrant transcription of MUC2 and TFF3. Moreover, two predicted glycosyltransferases, the sialyltransferases ST3GAL1 and ST3GAL2, exhibited inversely correlated transcriptional levels compared to the expression of the identified miRNAs miR-125a-5p and miR-615-3p, respectively. In this study, we mainly focused on the interaction between miR-615-3p and ST3GAL2 and were able to demonstrate their molecular interaction using luciferase reporter assays and RNAi. Detection of ST3GAL2 in murine colonic tissue by immunofluorescence demonstrated reduced intensity after C. jejuni 81-176 infection and was thus consistent with the observations made above.
We report here for the first time the regulation of glycosyltransferases by miRNAs during murine infection with C. jejuni 81-176. Our data suggest that mucin type O-glycan biosynthesis is concerted by the interplay of miRNAs and glycosyltransferases, which could determine the shape of intestinal glycosylated proteins during infection.