Title

Inactivation Of Klf5 By Zinc Finger Nuclease Downregulates Expression Of Pluripotent Genes And Attenuates Colony Formation In Embryonic Stem Cells

Keywords

Differentiation; ES cells; Klf5; Self-renewal; Zinc finger nucleases

Abstract

Recent studies suggest that Klf5 is required to maintain embryonic stem (ES) cells in an undifferentiated state. However, whether Klf5 can be inactivated by novel fusion technology of zinc finger nucleases (ZFN) has never before been examined. Therefore, we used ZFN technology to target the Klf5 gene in mouse ES cells, and examined the effects of the Klf5 gene on the expression of pluripotency-related genes, Oct3/4, Nanog, and Sox2 and on the self-renewal of ES cells. In Klf5-ZFN-transfected cells, expression of the Klf5 mRNA was downregulated by ∼80 % compared to the control. Furthermore, expression of the Oct3/4 and Nanog mRNAs was significantly decreased in the Klf5-ZFN-targeted cells. RT-PCR analysis, however, showed no significant change in the level of Sox2 mRNA, but a decreased trend was evident in the Klf5-ZFN-targeted cells. Moreover, we observed the spontaneous differentiation of Klf5-ZFN-transfected cells and quantitative analysis revealed a significant decrease in colony formation in Klf5-ZFN-transfected cells. In conclusion, our data suggest that ZFN methodology is an effective approach to target the Klf5 gene and that Klf5 plays an important role in the maintenance of ES cell self-renewal. © 2013 Springer Science+Business Media New York.

Publication Date

10-1-2013

Publication Title

Molecular and Cellular Biochemistry

Volume

382

Issue

1-2

Number of Pages

113-119

Document Type

Article

Personal Identifier

scopus

DOI Link

https://doi.org/10.1007/s11010-013-1724-5

Socpus ID

84884675613 (Scopus)

Source API URL

https://api.elsevier.com/content/abstract/scopus_id/84884675613

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