The pair of L5178Y sublines (LY-R and LY-S) is exceptional among mammalian cell lines because of their unique inverse cross-sensitivity to ionizing radiation and hydrogen peroxide. The high sensitivity of LY-S cells to ionizing radiation is reasonably explained by the impairment of DNA double strand breaks rejoining. Although the enzymatic defect of LY-S cells is not yet identified, the more pronounced effect of DNA-dependent protein kinase (DNA-PK) inhibitor (OK-1035) on DNA damage repair after 8 Gy x-irradiation in LY-R cells than in LY-S cells, suggests that LY-S cells may be defective in DNA-PK activity or in the other enzymatic activities downstream from DNA-PK. An additional feature is a higher protection of DNA against ionizing radiation by nuclear proteins in LY-R cells. These data support the concept that nuclear matrix organization may contribute to the cellular susceptibility to DNA damaging agents. Ionizing radiation-sensitive LY-S cells suffer also more DNA base damage than ionizing radiation-resistant LY-R cells. However, the repair rates of the g-ray-induced DNA base damage in LY sublines are related neither to the initial amounts of the damaged bases nor to the lethal or mutagenic effects of ionizing radiation.
In contrast, hydrogen peroxide (H₂O₂) sensitive LY-R cells suffer more DNA base damage after H₂O₂ treatment. This may be due to the lower activity of catalase and/or the lower level of glutathione and other monobromobimane-reactive thiols in LY-R cells than in LY-S cells. However, the main cause of LY-R cells’ sensitivity to H₂O₂ seems to be a higher iron ion content in these cells as compared to LY-S cells. A higher content of iron ions and a higher iron:copper ratio is found in isolated nuclei of LY-R cells than in those of LY-S cells. This is further confirmed by a higher "labile iron" pool in LY-R cells than in LY-S cells. Further evidence of different ions content in LY cells and its influence on nuclear matrix organization comes from a different effect of chelators of iron (desferrioxamine, DFO) and copper (neocupreine, NEO) on DNA properties in LY cells. The effect of NEO is more marked in LY-R than in LY-S cells and in both sublines it is expressed as enhanced tail moment (the measure of DNA damage in the comet assay) and increased micronuclei frequency. On the contrary, the effect of DFO on the tail moment is less pronounced in LY-R than in LY-S cells. With increasing DFO concentrations there is a gradual decrease in the tail moment values below the control level in LY-S cells. In LY-R cells the tail moment values initially increase, then gradually decreases, eventually falling below the control level. This points to a dramatic conformational change that masks the effect of DNA discontinuities. The presence of the latter is indicated by an increase in micronuclei frequency. These results support the postulated differential role of iron and copper ions in maintaining the higher order DNA structure in LY sublines.
The high iron ions content in LY-R is associated with a markedly greater content of ferritin H-subunits (H-Ft) in LY-S cells. The different expression of H-Ft in LY cells is a consequence of an up-regulation of H-Ft mRNA in LY-S cell line. In contrast, post-transcriptional control occurring at the level of iron responsive element (IRE)-iron regulatory protein (IRP) interaction is similar in both cell lines, as visualized by the identical IRE-binding activity of both IRP1 and IRP2. Furthermore, the data stress the importance of mutual dependence between the amount of IRP1 and the LIP level for final trans-regulatory activity of IRP1 and, in turn, for the overall control of intracellular iron balance.