LSD1-mediated demethylation of OCT4 safeguards pluripotent stem cells by maintaining the transcription of PORE-motif-containing genes

Songsong Dan,Xiaoyun Pan,Bo Kang,Xiaotao Duan,Jingchao Li,Shiqi She,Yanwen Zhou,Tong Su,Cheng Chen,Ying-Jie Wang,Xiaobing Zhang,Xinyu Chen,Yuelin Song,Xiao Pan,Wenjie Chen

doi:10.1038/s41598-021-89734-y

Songsong Dan, Xiaoyun Pan + Show 13 more

Open Access

https://doi.org/10.1038/s41598-021-89734-y

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Journal: Scientific Reports	Publication Date: May 13, 2021
Citations: 3	License type: open-access

Affiliation: Zhejiang University

Abstract

Reversible lysine methylation is essential for regulating histones and emerges to critically regulate non-histone proteins as well. Here we show that the master transcription factor OCT4 in pluripotent stem cells (PSCs) was methylated at multiple lysine residues. LSD1 that is highly expressed in PSCs can directly interact with and demethylate OCT4 at lysine 222 (K222) in the flexible linker region. Reduced LSD1 activity led to the methylation of OCT4-K222 that diminished the differentiation potential of PSCs while facilitating proteasome-independent degradation of OCT4 proteins. Furthermore, site-specifically replacing K222 with phenylalanine to mimic the constitutively methylated lysine promoted the ‘locked-in’ mode engagement of the OCT4 PORE-homodimers that tightly bind to and block the transcription of multiple PORE-motif-containing target genes regulating cell fate determination and cell junction organization, and thereby reducing the pluripotency of PSCs. Thus, LSD1-mediated demethylation of OCT4 plays a crucial role in restricting the ‘locked-in’ mode binding of OCT4 PORE-homodimers to the PORE-motif-containing genes and thereby maintaining their transcription to safeguard the pluripotency of PSCs.

Highlights

Reversible lysine methylation is essential for regulating histones and emerges to critically regulate non-histone proteins as well
Cell fate determination during embryogenesis depends on the properties of self-renewal and pluripotency possessed by pluripotent stem cells (PSCs) including embryonic stem cells (ESCs), embryonal carcinoma cells (ECCs) and induced pluripotent stem cells that have the potential to differentiate into three embryonic germ layers and are valuable for basic research, regenerative and translational medicine[1]
The intramolecular interaction between the murine Oct[4] linker residues and the POUH RK residues constrained the nonspecific binding of the POUH domain to random DNA sequences to ensure the specific recognition and binding of the OCT4-binding motifs, and it was speculated that post-translational modifications (PTMs) of critical residues in the OCT4 linker may impact on the mode of OCT4-DNA interactions[14]

Summary

Introduction

Reversible lysine methylation is essential for regulating histones and emerges to critically regulate non-histone proteins as well. Site- replacing K222 with phenylalanine to mimic the constitutively methylated lysine promoted the ‘locked-in’ mode engagement of the OCT4 PORE-homodimers that tightly bind to and block the transcription of multiple PORE-motif-containing target genes regulating cell fate determination and cell junction organization, and thereby reducing the pluripotency of PSCs. LSD1-mediated demethylation of OCT4 plays a crucial role in restricting the ‘locked-in’ mode binding of OCT4 PORE-homodimers to the PORE-motif-containing genes and thereby maintaining their transcription to safeguard the pluripotency of PSCs. Cell fate determination during embryogenesis depends on the properties of self-renewal and pluripotency possessed by pluripotent stem cells (PSCs) including embryonic stem cells (ESCs), embryonal carcinoma cells (ECCs) and induced pluripotent stem cells (iPSCs) that have the potential to differentiate into three embryonic germ layers (endoderm, mesoderm, and ectoderm) and are valuable for basic research, regenerative and translational medicine[1]. The demethylation of OCT4-K222 by LSD1 played an essential role in safeguarding the pluripotency of PSCs by restricting the ‘locked-in’ mode binding of OCT4 homodimers and allowing for the transcription of a group of PORE-motif-containing genes (PORE genes)

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