IDDF2024-ABS-0381 Bacteroides fragilis toxin suppresses METTL3-mediated m6A modification in macrophage to promote inflammatory bowel disease

Background

Microbiota-host interactions are crucial in the progression of inflammatory bowel disease (IBD). Despite extensive research, the detailed mechanisms through which these interactions influence the disease's development and progression remain largely unclear. Understanding these intricate processes is essential for developing targeted therapies and improving patient outcomes.

Methods

Male C57BL/6 mice were used for all experiments. Enterotoxigenic Bacteroides fragilis (ETBF) was administered orally to induce colitis, while control groups received non-toxigenic Bacteroides fragilis. Both bacterial strains were cultured under anaerobic conditions before gavage. Post-treatment, intestinal tissues were collected, and macrophages were isolated using collagenase digestion followed by magnetic cell sorting (MACS). Total RNA from these macrophages was extracted, and m6A modifications were analyzed via m6A-specific antibodies and high-throughput sequencing. Quantitative reverse transcription PCR (qRT-PCR) and Western blot were employed to measure expression levels of METTL3, FOXD3, ITGA5, and inflammatory markers. The role of FOXD3 in regulating METTL3 transcription was assessed using specific inhibitors or siRNA.

Results

Here, we reveal that Enterotoxigenic Bacteroides fragilis (ETBF) induces a dramatic reduction of m6A modifications in intestinal inflammatory macrophages, contributing to the initiation of IBD progression. Mechanistically, we characterize B fragilis toxin (BFT), produced by ETBF, which reduces METTL3 transcription by inhibiting FOXD3 expression and promotes the expression of its target integrin subunit alpha 5 (ITGA5) expression by diminishing YTHDF2-dependent mRNA degradation. Targeting ITAG5 using Cilengitide significantly alleviated ETBF-induced colitis by decreasing the level of inflammatory factors in macrophages.

Conclusions

Our research offers valuable insights into how the gut microbiota modulates the m6A epitranscriptome in human macrophages. This modulation plays a significant role in the progression of inflammatory bowel disease (IBD). By elucidating these complex interactions, our findings contribute to a deeper understanding of IBD pathogenesis and highlight potential avenues for therapeutic intervention.