USP21 deubiquitylates Nanog to regulate protein stability and stem cell pluripotency.

Xingyu Liu,Yuhan Chen,Yiling Zhang,Yuan Zhang,Yuying Yao,Chuanchun Han,Huiguo Ding,Ping Wang,Lingqiang Zhang,Wenyi Wei,Xin Zhang,Chanjuan Wang,Jing Zhang,Yun Zhai

doi:10.1038/sigtrans.2016.24

Abstract

The homeobox transcription factor Nanog has a vital role in maintaining pluripotency and self-renewal of embryonic stem cells (ESCs). Stabilization of Nanog proteins is essential for ESCs. The ubiquitin–proteasome pathway mediated by E3 ubiquitin ligases and deubiquitylases is one of the key ways to regulate protein levels and functions. Although ubiquitylation of Nanog catalyzed by the ligase FBXW8 has been demonstrated, the deubiquitylase that maintains the protein levels of Nanog in ESCs yet to be defined. In this study, we identify the ubiquitin-specific peptidase 21 (USP21) as a deubiquitylase for Nanog, but not for Oct4 or Sox2. USP21 interacts with Nanog protein in ESCs in vivo and in vitro. The C-terminal USP domain of USP21 and the C-domain of Nanog are responsible for this interaction. USP21 deubiquitylates the K48-type linkage of the ubiquitin chain of Nanog, stabilizing Nanog. USP21-mediated Nanog stabilization is enhanced in mouse ESCs and this stabilization is required to maintain the pluripotential state of the ESCs. Depletion of USP21 in mouse ESCs leads to Nanog degradation and ESC differentiation. Overall, our results demonstrate that USP21 maintains the stemness of mouse ESCs through deubiquitylating and stabilizing Nanog.

Highlights

Embryonic stem cells (ESCs) and induced pluripotent stem cells are pluripotent stem cell lines that have the ability to self-renew and differentiate into multiple lineages.[1]
We found that ubiquitin-specific peptidase 21 (USP21) significantly upregulated Nanog levels, whereas other DUBs had little to no effect on the Nanog expression levels (Figures 1a–c)
The effect of USP21 on Nanog is specific since overexpression of USP21 seemed to have little effect on Sox[2] and Oct[4] (Figure 2b), two wellcharacterized transcription factors in stem cells

Summary

Introduction

Embryonic stem cells (ESCs) and induced pluripotent stem cells are pluripotent stem cell lines that have the ability to self-renew and differentiate into multiple lineages.[1]. Previous studies have demonstrated that transcription factors are needed to activate and/or suppress Nanog expression. In this regard, Oct[4] and Sox[2] have been shown to be the main regulators of Nanog expression in mESCs.[11,12,13] Oct[4], Sox[2] and Nanog often co-occupy a substantial portion of their target genes and collaborate to form regulatory circuitry consisting of autoregulatory and feed-forward loops. Epigenetic factors, including Wdr[5], Mof and Ezh[2], can modulate Nanog transcription in ESCs.[14,15,16]

Methods

Results

Conclusion