Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
2067900 | Cell Biology International | 2006 | 9 Pages |
Abstract
The activity of the PCMV IE promoter was studied during the differentiation of ES cells into neurons. In order to do this, stable embryonic stem (ES) cell lines that express enhanced green fluorescent protein (EGFP) under the control of PCMV IE were created and these ES cells were differentiated by aggregation of cells in the presence of retinoic acid (RA). Based on our observations that the activity of PCMV IE was highest in undifferentiated cells, and that cell-cell interaction and addition of RA that lead to enhanced cell proliferation also increased expression from PCMV IE, we hypothesized that the activity of PCMV IE was positively regulated in cycling cells. However, when analysis was done at the single cell level it was found that BrdU label and EGFP expression were not correlated. EGFP expression was found to be down-regulated in many cells that were BrdU positive and conversely there were significant numbers of BrdU negative cells that were EGFP positive. Further, PCMV IE activity was not observed in cells that were nestin positive or in differentiated neurons, but PCMV IE was active in cells with a fibroblast-like morphology. Finally, several proteins present in undifferentiated ES cells were found to bind to regulatory regions of PCMV IE. These were absent when cells were aggregated in the presence of RA. The above results have implications for expression of transgenes in ES cells as well as providing new insight into the mechanism of lineage restriction.
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Authors
Bandita Bagchi, Manoj Kumar, Shyamala Mani,