Abstract
Functional expression cloning strategies are highly suitable for the analysis of the molecular control of apoptosis. This approach has two critical advantages. Firstly, it eliminates prior assumptions about the properties of the proteins involved, and, secondly, it selectively targets proteins that are causally involved in apoptosis control and which affect the crucial cellular decision between survival and death. The application of this strategy to the isolation of cDNAs conferring resistance to dexamethasone and γ-irradiation resulted in the isolation of a partial cDNA for the catalytic subunit of protein phosphatase 4 (PP4). Cells transfected with this partial cDNA in an expression vector downregulated PP4 and were resistant to both dexamethasone and UV radiation, as demonstrated by both membrane integrity and colony-forming assays. These observations suggest that PP4 plays an important proapoptotic role in T lymphocytes.
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Abbreviations
- PBS:
-
phosphate-buffered saline
- UV:
-
ultraviolet
- PMSF:
-
phenylmethylsulphonylfluoride
- PVDF:
-
polyvinylidenedifluoride
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Acknowledgements
We thank the Dead Cell Laboratory, WEHI, for warm hospitality, help and advice, Dr. Janet Meredith for initial subcloning of 4n10, Dr. Robert Farrell for X-irradiation of cells, and the Wellcome Trust, the Leukaemia Research Fund and Medical Research Council (UK) for financial support.
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Mourtada-Maarabouni, M., Kirkham, L., Jenkins, B. et al. Functional expression cloning reveals proapoptotic role for protein phosphatase 4. Cell Death Differ 10, 1016–1024 (2003). https://doi.org/10.1038/sj.cdd.4401274
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DOI: https://doi.org/10.1038/sj.cdd.4401274
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