Abstract
Identifying novel players of the pluripotency gene regulatory network centered on Oct4, Sox2, and Nanog as well as delineating the interactions within the complex network is key to understanding self-renewal and early cell fate commitment of embryonic stem cells (ESC). While overexpression of the transcriptional regulator Cited2 sustains ESC pluripotency, its role in ESC functions remains unclear. Here, we show that Cited2 is important for proliferation, survival, and self-renewal of mouse ESC. We position Cited2 within the pluripotency gene regulatory network by defining Nanog, Tbx3, and Klf4 as its direct targets. We also demonstrate that the defects caused by Cited2 depletion are, at least in part, rescued by Nanog constitutive expression. Finally, we demonstrate that Cited2 is required for and enhances reprogramming of mouse embryonic fibroblasts to induced pluripotent stem cells. Stem Cells 2015;33:699–712
Highlights
The maintenance of pluripotency of embryonic stem cells (ESC) is orchestrated by growth factors and signaling pathways that control the optimal expression of the transcription factors Oct4, Sox2, and Nanog, master regulators of pluripotency and self-renewal [1]
C2fl/fl ESC colonies maintained in culture with leukemia inhibitory factor (LIF) supplementation showed a typical ESC morphology, displayed alkaline phosphatase (AP) activity, www.StemCells.com and expressed pluripotency markers (Fig. 1A and Supporting Information Fig. S1A)
We transiently transfected C2fl/fl ESC with a Cre and green fluorescent protein (GFP) coexpressing plasmid, sorted highly GFP-positive ESC to enhance the chances of deleting both Cited2 alleles, and allowed the cells to grow in undifferentiating conditions for more than a month
Summary
The maintenance of pluripotency of embryonic stem cells (ESC) is orchestrated by growth factors and signaling pathways that control the optimal expression of the transcription factors Oct, Sox, and Nanog, master regulators of pluripotency and self-renewal [1]. A transcriptional regulator, strongly binds to p300 and CBP and is essential for mouse embryonic development [5,6,7,8,9] and the maintenance of fetal and adult hematopoietic stem cells [10, 11]. The mechanisms regulating Cited expression in mouse and human ESC are still poorly understood, but Cited transcriptional regulatory elements are bound by transcriptional factors critical for pluripotency such as Oct, Sox, Nanog, FoxP1, and Zfp206 [14,15,16]
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