AP-1B-mediated protein sorting regulates polarity and proliferation of intestinal epithelial cells in mice

Abstract

Background & aims: In epithelial cells, protein sorting mechanisms regulate localization of plasma membrane proteins that generate and maintain cell polarity. The clathrin-adaptor protein (AP) complex AP-1B is expressed specifically in polarized epithelial cells, where it regulates basolateral sorting of membrane proteins. However, little is known about its physiological significance.

Methods: We analyzed the intestinal epithelia of mice deficient in Ap1m2 (Ap1m2(-/-) mice), which encodes the AP-1B μ1B subunit, and compared it with 129/B6/CD1 littermates (controls). Notch signaling was inhibited by intraperitoneal injection of dibenzazepine, and β-catenin signaling was inhibited by injection of IWR1. Intestinal tissue samples were collected and analyzed by immunofluorescence analysis.

Results: Ap1m2(-/-) mice developed intestinal epithelial cell hyperplasia. The polarity of intestinal epithelial cells was disrupted, as indicated by the appearance of ectopic microvilli-like structures on the lateral plasma membrane and mislocalization of basolateral membrane proteins, including the low-density lipoprotein receptor and E-cadherin. The E-cadherin-β-catenin complex therefore was disrupted at the adherens junction, resulting in nuclear translocation of β-catenin. This resulted in up-regulation of genes regulated by β-catenin/transcription factor 4 (Tcf4) complex, and increased the proliferation of intestinal epithelial cells.

Conclusions: AP-1B is required for protein sorting and polarization of intestinal cells in mice. Loss of AP-1B in the intestinal epithelia results in mislocalization of E-cadherin, activation of β-catenin/Tcf4 complex, proliferation, and hyperplasia.

Keywords: AJ; AP; Apical and Basolateral; BMP; BrdU; Gene Regulation; LDLR; MDCK; Madin–Darby canine kidney; TGF; TGN; Tight Junction; ZO-1; adaptor protein; adherens junction; bone morphogenic protein; bromodeoxyuridine; low-density lipoprotein receptor; trans-Golgi network; transforming growth factor; zonula occludens-1.