Characterization of the Mechanisms Involved in Maintenance of Apical Trafficking and Polarity in Intestinal Epithelial Cells

Abstract

The apical recycling endosome supported by myosin Vb (MYO5B) acts as the main decision point for both the biosynthetic and endocytic pathways as they traffic proteins to and from the apical surface in polarized cells. Mutations in MYO5B cause microvillus inclusion disease (MVID), the main symptom of which is chronic, unremitting diarrhea. Utilizing this disease as a pathophysiologic model, we were able to unravel the mechanisms involved in MYO5B dependent apical trafficking in enterocytes. We determined that mutations in MYO5B disrupt normal apical trafficking and establishment of polarity in enterocytes, through Rab8a and Rab11a coupled pathways. This results in global changes in polarity at the villus tips that may account for deficits in apical absorption, loss of microvilli, aberrant junctions, and losses in transcellular ion transport pathways, leading to the MVID clinical phenotype of secretory diarrhea. In tandem, microvilli assembly requires MYO5B-coupled interaction with Rab8a, while loss of Rab11a interaction with MYO5B induced microvillus inclusions. Concomitantly, we were able to determine that mutations in MYO5B affect polarity in stomach parietal cells, enterocytes of the large intestine, and hepatocytes. These novel findings suggest that the effects of MYO5B mutations in MVID patients are not isolated to the small intestine. Rather alterations in polarity are observed across many tissue types. Recently, mutations in STX3 have been implicated as the cause of Atypical MVID and in these patients’ enterocytes polarity is disrupted and apical microvilli are loss. We also report that Rab11a regulates apical intestinal enterocyte polarity, and demonstrate that Rab11a loss results in a decrease in microvilli length, lateral microvilli formation, mistrafficking of apically trafficked proteins, and mislocalization of STX3. We have identified one patient with unstable MYO5B who has displayed relapsing and remitting symptoms diarrheal symptoms for 21 years. Since, the morphological characteristics in MVID overlaps with other diarrheal syndromes we theorize that mutations in MYO5B and other apical trafficking component genes might account for populations of patients with celiac disease who fail to respond to gluten-free diet and/or similar enteropathies.