scholarly journals Two DNA binding domains of MGA act in combination to suppress ectopic activation of meiosis‐related genes in mouse embryonic stem cells

Stem Cells ◽  
2021 ◽  
Vol 39 (11) ◽  
pp. 1435-1446
Author(s):  
Kousuke Uranishi ◽  
Masataka Hirasaki ◽  
Yuka Kitamura ◽  
Yosuke Mizuno ◽  
Masazumi Nishimoto ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Dna Binding ◽  
Embryonic Stem ◽  
Mouse Embryonic Stem Cells ◽  
Dna Binding Domains ◽  
Binding Domains ◽  
Ectopic Activation

scholarly journals Two DNA binding domains of Mga act in combination to suppress ectopic activation of meiosis-related genes in mouse embryonic stem cells

2020 ◽  
Author(s):  
Kousuke Uranishi ◽  
Masataka Hirasaki ◽  
Yuka Kitamura ◽  
Yosuke Mizuno ◽  
Masazumi Nishimoto ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Dna Binding ◽  
Germ Cells ◽  
Molecular Mechanisms ◽  
Embryonic Stem ◽  
Mouse Embryonic Stem Cells ◽  
Dna Binding Domains ◽  
Binding Domains ◽  
Meiotic Entry

SUMMARYMouse embryonic stem cells (ESCs) have high potential for meiotic entry, like germ cells. Although the physiological meaning of this potential is not known, it is certain that a rigid safeguarding system is required to prevent ectopic onset of meiosis. PRC1.6, a non-canonical PRC1, is known for its suppression of precocious and ectopic meiotic onset in germ cells and ESCs, respectively, in which MGA has important roles in DNA binding as well as in constructing the complex as a scaffolding component. As a salient feature, MGA bears two distinct DNA-binding domains termed bHLHZ and T-box. However, how these features contribute to the functions of PRC1.6, particularly in the repression of meiotic genes, remains largely obscure. Here, we demonstrated that both DNA binding domains of Mga repress distinct sets of genes in murine ESCs, and substantial numbers of meiosis-related genes are included in both gene sets. In addition, our data demonstrated that both DNA binding domains of Mga, in particular bHLHZ, are crucially involved in repressing the expression of Meiosin, which plays essential roles in meiotic entry in collaboration with Stra8, revealing at least part of the molecular mechanisms that link negative and positive regulation of meiotic onset.

scholarly journals Structure-based discovery of NANOG variant with enhanced properties to promote self-renewal and reprogramming of pluripotent stem cells

2015 ◽  
Vol 112 (15) ◽  
pp. 4666-4671 ◽  
Author(s):  
Yohei Hayashi ◽  
Laura Caboni ◽  
Debanu Das ◽  
Fumiaki Yumoto ◽  
Thomas Clayton ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Dna Binding ◽  
Pluripotent Stem Cells ◽  
Biological Activities ◽  
Embryonic Stem ◽  
Dna Interaction ◽  
Mouse Embryonic Stem Cells ◽  
Self Renewal ◽  
Mutant Proteins

NANOG (from Irish mythology Tír na nÓg) transcription factor plays a central role in maintaining pluripotency, cooperating with OCT4 (also known as POU5F1 or OCT3/4), SOX2, and other pluripotency factors. Although the physiological roles of the NANOG protein have been extensively explored, biochemical and biophysical properties in relation to its structural analysis are poorly understood. Here we determined the crystal structure of the human NANOG homeodomain (hNANOG HD) bound to an OCT4 promoter DNA, which revealed amino acid residues involved in DNA recognition that are likely to be functionally important. We generated a series of hNANOG HD alanine substitution mutants based on the protein–DNA interaction and evolutionary conservation and determined their biological activities. Some mutant proteins were less stable, resulting in loss or decreased affinity for DNA binding. Overexpression of the orthologous mouse NANOG (mNANOG) mutants failed to maintain self-renewal of mouse embryonic stem cells without leukemia inhibitory factor. These results suggest that these residues are critical for NANOG transcriptional activity. Interestingly, one mutant, hNANOG L122A, conversely enhanced protein stability and DNA-binding affinity. The mNANOG L122A, when overexpressed in mouse embryonic stem cells, maintained their expression of self-renewal markers even when retinoic acid was added to forcibly drive differentiation. When overexpressed in epiblast stem cells or human induced pluripotent stem cells, the L122A mutants enhanced reprogramming into ground-state pluripotency. These findings demonstrate that structural and biophysical information on key transcriptional factors provides insights into the manipulation of stem cell behaviors and a framework for rational protein engineering.

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