Journal
MOLECULAR & CELLULAR PROTEOMICS
Volume 8, Issue 6, Pages 1401-1412Publisher
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
DOI: 10.1074/mcp.M800478-MCP200
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- Medical Research Council and Engineering
- Physical Sciences Research Council [GR/S84347/01, EP/E036252/1]
- EPSRC [EP/E036252/1] Funding Source: UKRI
- Engineering and Physical Sciences Research Council [EP/E036252/1, GR/S84347/01] Funding Source: researchfish
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The functional reprogramming of a differentiated cell to a pluripotent state presents potential beneficial applications in regenerative medicine. We report here the proteomic profile of 293T epithelial cells reprogrammed to a pluripotent state using undifferentiated embryonal carcinoma (NCCIT) cellular extracts. 293T cells were reversibly permeabilized with streptolysin O, incubated in an extract of NCCIT cells or a control extract of 293T cells for 1 h, resealed with CaCl2, and cultured. OCT4 and SOX2 gene expression were up-regulated in NCCIT extract-treated cells relative to control cells, whereas there was no alteration in DNMT3B gene expression. Thirty percent of NCCIT extract-treated cells were positive for SSEA-4, and karyotyping confirmed their 293T origin, excluding the possibility of contamination from NCCIT cells. Two-dimensional PAGE revealed similar to 400 protein spots for each cell type studied. At least 10 protein spots in the proteome of NCCIT extract-treated cells had an expression profile similar to that of NCCIT and remained unaltered in control cells. Using tandem mass spectrometry, we identified these proteins, which include 78-kDa glucose-regulated protein precursor and tropomyosin alpha - 3 chain. This investigation provides the first evidence that proteins are altered in a specific manner in NCCIT extract-treated cells. This is the first report on the proteomic characterization of the nuclear reprogramming process. Molecular & Cellular Proteomics 8: 1401-1412, 2009.
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