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Ets1 is required for p53 transcriptional activity in UV-induced apoptosis in embryonic stem cells
Dakang Xu1, Trevor J Wilson, David Chan
1Centre for Functional Genomics and Human Disease, Monash Institute of Reproduction and Development, Monash University, 246 Clayton, Clayton, Victoria 3168, Australia.
Abstract:
Embryonic stem (ES) cells contain a p53-dependent apoptosis mechanism to avoid the continued proliferation and differentiation of damaged cells. We show that mouse ES cells lacking Ets1 are deficient in their ability to undergo UV-induced apoptosis, similar to p53 null ES cells. In Ets1(-/-) ES cells, UV induction of the p53 regulated genes mdm2, perp, cyclin G and bax was decreased both at mRNA and protein levels. While p53 protein levels were unaltered in Ets1(-/-) cells, its ability to transactivate genes such as mdm2 and cyclin G was reduced. Furthermore, electrophoretic mobility shift assays and immunoprecipitations demonstrated that the presence of Ets1 was necessary for a CBP/p53 complex to be formed. Chromatin immunoprecipitations demonstrated that Ets1 was required for the formation of a stable p53-DNA complex under physiological conditions and activation of histone acetyltransferase activity. These data demonstrate that Ets1 is an essential component of a UV-responsive p53 transcriptional activation complex in ES cells and suggests that Ets1 may contribute to the specificity of p53-dependent gene transactivation in distinct cellular compartments.
Insights
Ets1 is crucial for UV-induced apoptosis in embryonic stem (ES) cells by enabling p53 to activate target genes. This finding highlights Ets1
Area of Science:
- Cell Biology
- Molecular Biology
- Stem Cell Research
Background:
- Embryonic stem (ES) cells possess a p53-dependent apoptosis pathway to eliminate damaged cells.
- UV radiation triggers apoptosis in ES cells, a process critical for genomic integrity.
Purpose of the Study:
- To investigate the role of Ets1 in UV-induced apoptosis and p53 transcriptional activity in mouse ES cells.
- To elucidate the molecular mechanism by which Ets1 influences p53 function.
Main Methods:
- Comparison of UV-induced apoptosis in wild-type, Ets1(-/-), and p53 null ES cells.
- Analysis of p53-regulated gene expression (mRNA and protein) via quantitative PCR and Western blotting.
- Electrophoretic mobility shift assays (EMSA), co-immunoprecipitation, and chromatin immunoprecipitation (ChIP) to assess protein-DNA interactions and complex formation.
Main Results:
- Ets1(-/-) ES cells exhibit impaired UV-induced apoptosis, similar to p53 null ES cells.
- UV induction of p53 target genes (mdm2, perp, cyclin G, bax) is significantly reduced in Ets1(-/-) ES cells at both mRNA and protein levels.
- Ets1 is essential for p53's transactivation of target genes, CBP/p53 complex formation, stable p53-DNA binding, and histone acetyltransferase activity.
Conclusions:
- Ets1 is a critical component of the UV-responsive p53 transcriptional activation complex in ES cells.
- Ets1 facilitates p53-dependent gene transactivation, suggesting a role in cellular compartment-specific p53 activity.