Clathrin-Mediated Endocytosis Regulates a Balance between Opposing Signals to Maintain the Pluripotent State of Embryonic Stem Cells

Abstract

Summary: Endocytosis is implicated in the maintenance of embryonic stem cell (ESC) pluripotency, although its exact role and the identity of molecular players remain poorly understood. Here, we show that the clathrin heavy chain (CLTC), involved in clathrin-mediated endocytosis (CME), is vital for maintaining mouse ESC (mESC) pluripotency. Knockdown of Cltc resulted in a loss of pluripotency accompanied by reduced E-cadherin (E-CAD) levels and increased levels of transforming growth factor β (TGF-β) and extracellular signal-regulated kinase (ERK) signaling. We demonstrate that both E-CAD and TGF-β receptor type 1 (TGF-βR1) are internalized through CME in mESCs. While E-CAD is recycled, TGF-βR1 is targeted for lysosomal degradation thus maintaining inverse levels of these molecules. Finally, we show that E-CAD interacts with ERK, and that the decreased pluripotency upon CME loss can be rescued by inhibiting TGF-βR, MEK, and GSK3β, or overexpressing E-CAD. Our results demonstrate that CME is critical for balancing signaling outputs to regulate ESC pluripotency, and possibly cell fate choices in early development. : Subramanyam and colleagues demonstrate that clathrin-mediated endocytosis (CME) is necessary to maintain ESC pluripotency by regulating the trafficking of E-CAD and TGF-βR1. Loss of CME results in a loss of pluripotency characterized by decreased E-CAD and enhanced TGF-βR1 and MEK signaling. Expression of pluripotency markers in ESCs lacking Cltc can be restored by inhibiting TGF-βR1 or MEK or by overexpressing E-CAD. Keywords: embryonic stem cells, pluripotency, trafficking, E-cadherin, clathrin, recycling, TGF-β, epithelial