Interactome analysis reveals endocytosis and membrane recycling of EpCAM during differentiation of embryonic stem cells and carcinoma cells

Min Pan,Vera Kohlbauer,Alexandra Blancke Soares,Henrik Schinke,Yuanchi Huang,Gisela Kranz,Tanja Quadt,Matthias Hachmeister,Olivier Gires

doi:10.1016/j.isci.2021.103179

Abstract

SummaryTransmembrane epithelial cell adhesion molecule (EpCAM) is expressed in epithelia, carcinoma, teratoma, and embryonic stem cells (ESCs). EpCAM displays spatiotemporal patterning during embryogenesis, tissue morphogenesis, cell differentiation, and epithelial-to-mesenchymal transition (EMT) in carcinomas. Potential interactors of EpCAM were identified in murine F9 teratoma cells using a stable isotope labeling with amino acids in cell culture-based proteomic approach (n = 77, enrichment factor >3, p value ≤ 0.05). Kyoto Encyclopedia of Genes and Genomes and gene ontology terms revealed interactions with regulators of endosomal trafficking and membrane recycling, which were further validated for Rab5, Rab7, and Rab11. Endocytosis and membrane recycling of EpCAM were confirmed in mF9 cells, E14TG2α ESC, and Kyse30 carcinoma cells. Reduction of EpCAM during mesodermal differentiation and TGFβ-induced EMT correlated with enhanced endocytosis and block or reduction of recycling in ESCs and esophageal carcinoma cells. Hence, endocytosis and membrane recycling are means of regulation of EpCAM protein levels during differentiation of ESC and EMT induction in carcinoma cells.

Highlights

Epithelial cell adhesion molecule (EpCAM) was initially identified as an antigen expressed on colon carcinoma cells that induced a humoral response in mice (Herlyn et al, 1979; Koprowski et al, 1979)
Potential interactors of epithelial cell adhesion molecule (EpCAM) were identified in murine F9 teratoma cells using a stable isotope labeling with amino acids in cell culture-based proteomic approach (n = 77, enrichment factor >3, p value % 0.05)
Endocytosis and membrane recycling of EpCAM were confirmed in mF9 cells, E14TG2a embryonic stem cells (ESCs), and Kyse30 carcinoma cells

Summary

Introduction

Epithelial cell adhesion molecule (EpCAM) was initially identified as an antigen expressed on colon carcinoma cells that induced a humoral response in mice (Herlyn et al, 1979; Koprowski et al, 1979). EpCAM is composed of an extracellular domain, a single type I transmembrane domain, and a short intracellular domain (Balzar et al, 1999) Functional implications of this transmembrane glycoprotein range from cell adhesion and junction (Ladwein et al, 2005; Litvinov et al, 1994a, 1994b; Wu et al, 2013), migration and morphogenesis (Gaiser et al, 2012; Maghzal et al, 2010, 2013; Slanchev et al, 2009), tissue integrity (Gaston et al, 2021; Guerra et al, 2012; Kozan et al, 2015; Lei et al, 2012; Salomon et al, 2017; Sivagnanam et al, 2008), proliferation (Munz et al, 2004; Osta et al, 2004), and signal transduction (Chaves-Perez et al, 2013; Maetzel et al, 2009) to differentiation and stem cell pluripotency (Gonzalez et al, 2009; Huang et al, 2011; Lu et al, 2010, 2013; Ng et al, 2009; Sarrach et al, 2018). The functions of EpICD can be controlled at various levels, including its initial cleavage and nuclear translocation (Denzel et al, 2009) and its degradation by the proteasome (Huang et al, 2019)

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