Transcriptomics

Dataset Information

3

Determination of the protein interactome of the transcription factor Sox2 in embryonic stem cells engineered for inducible expression of four reprogramming factors


ABSTRACT: Coordinate expression of the somatic cell reprogramming factors Oct4, Sox2, Klf4 and c-Myc within embryonic stem cells preserves the self-renewal of these cells, while allowing for the expression epitope tagged Sox2. Taking advantage of this observation, we engineered embryonic stem cells (i-OSKM-ESC) to inducibly express Oct4, Klf4, c-Myc and an epitope tagged form of Sox2 from a polycistronic element, in the presence of doxycycline. We isolated Sox2 and its associated protein complexes by co-immunoprecipitation. Subsequently, we identified the Sox2-protein interactome in self-renewing embryonic stem cells using an unbiased proteomic screen (Multidimensional Protein Identification Technology [MudPIT]). Affymetrix microarrays were used to characterize the gene expression profile of i-OSKM-ESC in the absence and presence of doxycycline. Mouse embryonic stem cells (KH2) were engineered to express Oct4, epitope tagged Sox2, Klf4 and c-Myc in the presence of doxycyline, to produce i-OSKM-ESC. The i-OSKM-ESC were cultured in the absence or presence of doxycyline (4 µg/mL) for 24 hours. RNA was extracted from each condition, and used for microarray analysis.

ORGANISM(S): Mus musculus  

SUBMITTER: Sunil K Mallanna   Michael P Washburn  Joshua M Gilmore  Angie Rizzino  Jesse L Cox  Zhiguang Gao  Angie Rizzinio  Briana D Ormsbee 

PROVIDER: E-GEOD-34801 | ArrayExpress | 2012-02-20

SECONDARY ACCESSION(S): GSE34801PRJNA151129

REPOSITORIES: GEO, ArrayExpress

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Publications

Determination of protein interactome of transcription factor Sox2 in embryonic stem cells engineered for inducible expression of four reprogramming factors.

Gao Zhiguang Z   Cox Jesse L JL   Gilmore Joshua M JM   Ormsbee Briana D BD   Mallanna Sunil K SK   Washburn Michael P MP   Rizzino Angie A  

The Journal of biological chemistry 20120209 14


Unbiased proteomic screens provide a powerful tool for defining protein-protein interaction networks. Previous studies employed multidimensional protein identification technology to identify the Sox2-interactome in embryonic stem cells (ESC) undergoing differentiation in response to a small increase in the expression of epitope-tagged Sox2. Thus far the Sox2-interactome in ESC has not been determined. To identify the Sox2-interactome in ESC, we engineered ESC for inducible expression of differen  ...[more]

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