| Budget Amount *help |
¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1992: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 1991: ¥1,200,000 (Direct Cost: ¥1,200,000)
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| Research Abstract |
Cadmium-resistant Chinese hamster V79 cells were isolated and the mechanism for the resistance was investigated as follows. Intracellular level of glutathione and inducibility of metallothionein, both of which were crucial factors determining the sensitivity of cells to cadmium toxi-city, were not different between Cd-resistant and Cd-sensitive cells. However, accumulation of Cd in Cd-resistant cells decreased markedly as compared to that in sensitive cells. The en-hancement of Cd efflux was considered as a mechanism for the decreased accumulation of Cd. But, the radioactive Cd once incorporated into cells did not release outside the cells in both Cd-resistant and Cd-sensitive cells, indicating that the active extrusion of Cd does not participate in the resistant phenotype.
SH proteins in plasma membrane that have high affinity to Cd were investigated by the combination of SDS-PAGE and SH-blotting. A protein band of the molecular weight 100K was found in plasma membrane proteins of Cd-resistant cells but not in Cd-sensitive cells. This protein is being purified by the method of two dimensional gel electrophoresis for the subsequent formation of the antibody. Likewise, several cadmium binding membrane proteins were detected by the technique of SDS-PAGE and ^<109>Cd-blotting of the membrane proteins. However, at present no difference in the binding of Cd with these proteins is observed between Cd-resistant and Cd-sensitive cells.
The hybrid cells between Cd-resistant and Cd-sensitive cells were isolated and their sensitivity to Cd and accumulation of ^<109>Cd were compared with the parental cells. Of the hybrids most were resistant to Cd similarly to the Cd-resistant parental cells. However, for ^<109>Cd accumulation these hybrids showed an intermediate level of the accumulation by Cd-resistant and Cd-sensitive parental cells, suggesting expression of both phenotypes for Cd uptake in hybrid cells.
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