Co-regulated expression of dbl and poly(ADP-ribose) polymerase in Ewing's sarcoma cells and dbl-transformed NIH3T3...

Poly(ADP-ribose) polymerase (PADPRP) is a ubiquitous enzyme constitutively expressed at low levels in the majority of eukaryotic cells, including most mammalian tumors and tumor-derived cell lines. Overexpression of PADPRP following the introduction of cDNA recombinant constructs into various cell types results in cell death. An exception to this effect are Ewing's sarcoma (ES) cells, which have been shown to contain elevated steady-state levels of PADPRP mRNA and high constitutive levels of protein and polymerase activity. In fact, this excess of PADPRP has been suggested to participate in the intrinsic radiosensitivity of Ewing's sarcomas, a highly malignant childhood bone tumor frequently curable with radiotherapy. It appears that ES cells might possess a hitherto unknown mechanism(s) by which PADPRP overexpression is controlled and made compatible with cell survival and proliferation. In order to investigate the contribution of other genetic alterations to PADPRP regulation in ES cells, we analysed the expression levels of PADPRP and of other genes, such as oncogenes and tumor suppressor genes, which may enhance the proliferative potential of ES cells. We have detected a positive correlation between the expression levels of the DNA-repair enzyme poly(ADP-ribose) polymerase and the dbl proto-oncogene in Ewing's sarcoma cells. The co-regulated expression of these genes has been established in NIH3T3 cells transformed by the human dbl oncogene or by overexpression of the dbl proto-oncogene. In both instances, the increase in dbl expression resulted in elevated levels of PADPRP mRNA and polymerase activity. The dbl oncogene was more efficient than the proto-oncogene in upregulating PADPRP expression. The inability of other oncogenes to upregulate PADPRP upon transformation of NIH3T3 cells demonstrated the specificity of the dbl in the process. Transfection of dbl-transformed NIH3T3 cells with retroviral PADPRP vectors resulted in the establishment of clones with PADPRP levels higher than those detectable in untransformed NIH3T3 cells transfected with the same retroviral constructs. These results suggest that dbl plays a role in the mechanism by which mammalian cells autoregulate their endogenous levels of PADPRP. Post-translational modification of the dbl or proto-dbl proteins by cytoplasmic PADPRP does not participate in the mechanism(s) underlying the observed PADPRP/dbl co-regulation.