Induction of apoptosis by overexpression of the DNA-binding and DNA-PK-activating protein C1D
| Autor: | Rothbarth, K., Spiess, E., Juodka, B., Yavuzer, U., Nehls, P., Stammer, H., Werner, D. |
|---|---|
| Rok vydání: | 1999 |
| Předmět: |
Recombinant Fusion Proteins
Gene Expression DNA-binding protein Apoptosis DNA-Activated Protein Kinase Protein Serine-Threonine Kinases DNA dependent protein kinase Transfection DNA-PK Bystander effect Mice Tumor Cells Cultured Vector-dependent overexpression Animals Humans Carcinoma Ehrlich Tumor Enhanced green fluorescent protein C1D gene Nuclear Proteins Cell Biology DNA binding protein DNA-Binding Proteins Enzyme Activation Repressor Proteins Tumor Suppressor Protein p53 DNA nucleotidylexotransferase Co-Repressor Proteins HeLa Cells Plasmids |
| Zdroj: | Journal of Cell Science |
| ISSN: | 1477-9137 0021-9533 |
| DOI: | 10.1242/jcs.112.13.2223 |
| Popis: | Apoptosis is induced in various tumor cell lines by vector-dependent overexpression of the conserved gene C1D that encodes a DNA-binding and DNA-PK-activating protein. C1D is physiologically expressed in 50 human tissues tested, which points to its basic cellular function. The expression of this gene must be tightly regulated because elevated levels of C1D protein, e.g. those induced by transient vector-dependent expression, result in apoptotic cell death. Cells transfected with C1D-expressing constructs show terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling of DNA ends. Transfections with constructs in which C1D is expressed in fusion with the (enhanced) green fluorescent protein from A. victoria (EGFP) allow the transfected cells to be identified and the morphological changes induced to be traced. Starting from intense nuclear spots, green fluorescence reflecting C1D expression increases dramatically at 12–24 hours post-transfection. Expression of C1D-EGFP protein is accompanied by morphological changes typical of apoptotic cell death, e.g. cytoplasmic vacuolation, membrane blebbing and nuclear disintegration. Cell shrinkage and detachment from extracellular matrix are observed in monolayer cultures while suspension cells become progressively flattened. The facility to differentiate between transfected and non-transfected cells reveals that non-transfected cells co-cultured with transfected cells also show the morphological changes of apoptosis, which points to a bystander effect. C1D-dependent apoptosis is not induced in cells with non-functional p53. Accordingly, C1D-induced apoptosis is discussed in relation to its potential to activate DNA-PK, which has been considered to act as an upstream activator of p53. |
| Databáze: | OpenAIRE |
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