During embryonic development,normal cell turnover, and other physiological and pathological processes, cells die by the activation of a mechanism of self-destruction, loosely termed apoptosis or programmed cell death. This process appears to be under genetic and biochemical control and often involves the activation of genes and the synthesis of proteins probably involved in the process. Radiation and chemotherapeutic agents also generally elicit cell death by apoptosis. Little is known about the genes of biochemical mechanisms involved in apoptosis. For this reason, we have begun a study to identify genes involved in apoptosis in radiation-sensitive fetal rat brain neuroblasts following radiation treatment. It is anticipated that among the genes activated by irradiation will be genes required for neuroblast apoptosis, either for the triggering or for the execution of this process. Fetal E17 rats were irradiated (or sham-irradiated) with sublethal neutron/gamma irradiation in utero, and RNA isolated from brains 5 and 24 hours later was used to construct a subtractive CDNA library enriched in sequences of transcripts increased by irradiation. Clones from this library were screened and analyzed by differential colony and Northern blot hybridization. At least 76 out of 682 analyzed clones were found to represent transcripts increased in abundance by irradiation, generally by a factor of 2-3 but much greater for at least one transcript. Sequencing of the inserts of these clones is in progress. Some clones were found to have previously unreported insert DNA sequences, while other represent ubiquitin, ferritin light chain, 12.3 protein (related by Gbeta proteins) and subunits of mitochondrial NADH dehydrogenase. These results indicate that a variety of transcripts are increased moderately during radiation-induced neuroblast cell death. Additional more strongly activated genes, which are expected in the triggering of cell death, are currently being sought. Further analyses in cultured cell systems and with antisense and other molecular genetic strategies are needed to establish a role for each of the activated genes in the mechanism of apoptosis.
