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Inhibition of master transcription factors in pluripotent cells induces early stage differentiation.

ABSTRACT: The potential for pluripotent cells to differentiate into diverse specialized cell types has given much hope to the field of regenerative medicine. Nevertheless, the low efficiency of cell commitment has been a major bottleneck in this field. Here we provide a strategy to enhance the efficiency of early differentiation of pluripotent cells. We hypothesized that the initial phase of differentiation can be enhanced if the transcriptional activity of master regulators of stemness is suppressed, blocking the formation of functional transcriptomes. However, an obstacle is the lack of an efficient strategy to block protein-protein interactions. In this work, we take advantage of the biochemical property of seventeen kilodalton protein (Skp), a bacterial molecular chaperone that binds directly to sex determining region Y-box 2 (Sox2). The small angle X-ray scattering analyses provided a low resolution model of the complex and suggested that the transactivation domain of Sox2 is probably wrapped in a cleft on Skp trimer. Upon the transduction of Skp into pluripotent cells, the transcriptional activity of Sox2 was inhibited and the expression of Sox2 and octamer-binding transcription factor 4 was reduced, which resulted in the expression of early differentiation markers and appearance of early neuronal and cardiac progenitors. These results suggest that the initial stage of differentiation can be accelerated by inhibiting master transcription factors of stemness. This strategy can possibly be applied to increase the efficiency of stem cell differentiation into various cell types and also provides a clue to understanding the mechanism of early differentiation.

SUBMITTER: De D

PROVIDER: S-EPMC3918783 | biostudies-literature | 2014 Feb

REPOSITORIES: biostudies-literature

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Inhibition of master transcription factors in pluripotent cells induces early stage differentiation.

De Debojyoti D Jeong Myong-Ho MH Leem Young-Eun YE Svergun Dmitri I DI Wemmer David E DE Kang Jong-Sun JS Kim Kyeong Kyu KK Kim Sung-Hou SH

Proceedings of the National Academy of Sciences of the United States of America 20140116 5

The potential for pluripotent cells to differentiate into diverse specialized cell types has given much hope to the field of regenerative medicine. Nevertheless, the low efficiency of cell commitment has been a major bottleneck in this field. Here we provide a strategy to enhance the efficiency of early differentiation of pluripotent cells. We hypothesized that the initial phase of differentiation can be enhanced if the transcriptional activity of master regulators of stemness is suppressed, blo ...[more]

PMID: 24434556

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Project description:Dry eye disease is the most prevalent pathological condition in aging eyes. One potential therapeutic strategy is the transplantation of lacrimal glands, generated in vitro from pluripotent stem cells such as human embryonic stem cells, into patients. One of the preceding requirements is a method to differentiate human embryonic stem cells into lacrimal gland epithelium cells. As the first step for this approach, this study aims to identify a set of transcription factors whose overexpression can promote the differentiation of human embryonic stem cells into lacrimal gland epithelium-like cells. We performed microarray analyses of lacrimal glands and lacrimal glands-related organs obtained from mouse embryos and adults, and identified transcription factors enriched in lacrimal gland epithelium cells. We then transfected synthetic messenger RNAs encoding human orthologues of these transcription factors into human embryonic stem cells and examined whether the human embryonic stem cells differentiate into lacrimal gland epithelium-like cells by assessing cell morphology and marker gene expression. The microarray analysis of lacrimal glands tissues identified 16 transcription factors that were enriched in lacrimal gland epithelium cells. We focused on three of the transcription factors, because they are expressed in other glands such as salivary glands and are also known to be involved in the development of lacrimal glands. We tested the overexpression of various combinations of the three transcription factors and PAX6, which is an indispensable gene for lacrimal glands development, in human embryonic stem cells. Combining PAX6, SIX1, and FOXC1 caused significant changes in morphology, i.e., elongated cell shape and increased expression (both RNAs and proteins) of epithelial markers such as cytokeratin15, branching morphogenesis markers such as BARX2, and lacrimal glands markers such as aquaporin5 and lactoferrin. We identified a set of transcription factors enriched in lacrimal gland epithelium cells and demonstrated that the simultaneous overexpression of these transcription factors can differentiate human embryonic stem cells into lacrimal gland epithelium-like cells. This study suggests the possibility of lacrimal glands regeneration from human pluripotent stem cells.

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Inhibition of master transcription factors in pluripotent cells induces early stage differentiation.

Publications

Inhibition of master transcription factors in pluripotent cells induces early stage differentiation.

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