Roles of autophagy-related proteins in the pathogeneis of Crohn's disease

Crohn's disease (CD) is a chronic, inflammatory disease of the gastrointestinal tract that affects millions of people worldwide. Its physiology is complex and is thought to result from interactions between genetic predispositions and environmental triggers under the influence of immune responses. Genome-wide association scans (GWAS) have identified a large number of genes associated with CD, but the increased risk imparted by each of them is low. This suggests that the environmental component of CD is crucial for disease development. We examined the CD-associated risk allele (rs2241880-G) of Atg16L1 and the immune response protein HMGB1 during CD to attempt to understand how this gene variant combines with environmental triggers and immune responses to lead to CD.;We generated HCT 116 cells deficient in Atg16L1 (Atg16L1Delta/Delta ) or homozygous for the CD-associated risk allele (Atg16L1 G/G) and used them to show that Atg16L1 has a role in bacterial entry and loss of Atg16L1 delays invasion by Salmonella typhimurium. We also demonstrated that Atg16L1 is important for lipidation of the mAtg8 family members GABARAP and GATE-16, in addition to LC3, and that LC3 interacts with microtubules in an Atg16L1-dependent manner. The importance of Atg16L1 to vesicular trafficking was evident as altered transferrin receptor trafficking and iron delivery in cells deficient in Atg16L1. These data demonstrate that Atg16L1 has non-autophagic functions that contribute to host-microbe interactions and non-autophagic vesicular trafficking.;We also demonstrated that the CD-associated risk allele of Atg16L1 is associated with a decrease in an isoform of Atg16L1 composed entirely of the WD repeat region of the protein. The decreased levels of the truncated Atg16L1 isoform are due, at least in part, to decreased translational efficiency. This isoform binds microtubules and modulates their dynamic instability. Decreased levels are associated with altered transferrin receptor trafficking and decreased iron availability in IEC. Since rs2241880-G does not affect basal autophagy, it likely contributes to development of CD through alterations in microtubule-mediated vesicular trafficking, rather than alterations in autophagy.;Finally, we examined inflammatory processes in intestinal epithelial cells (IEC) during colitis in mouse models and in inflammatory bowel disease (IBD) using HMGB1f/f, Vil-CRE mice and samples from human patients. These showed that calpain activation is a key feature of this inflammation and that autophagy is compromised during IBD by calpain-mediated cleavage of beclin-1, which converts it from a pro-autophagic to a pro-apoptotic protein. HMGB1 protects beclin-1 from this cleavage and so controls the cell fate of IEC during this inflammatory process.;This work has provided new clues about how genetic predispositions and inflammatory processes contribute to CD. We propose a model for CD development wherein iron deficiency coupled with calpain activation could lead to sustained gastrointestinal damage, decreased epithelial repair, and CD.