THE RAS SUPPRESSOR RSU-1 LOCALIZES TO 10P13 AND ITS EXPRESSION IN THEU251 GLIOBLASTOMA CELL-LINE CORRELATES WITH A DECREASE IN GROWTH-RATEAND TUMORIGENIC POTENTIAL

Rsu-1, which was isolated based on its ability to suppress transformat ion by v-Ras, is a highly conserved gene which shares homology with ye ast adenylyl cyclase in the region required for activation by Ras, Gen omic DNA clones of human RSU-1 have been isolated and used as a probe for fluorescence in situ hybridization (FISH) to assign RSU-1 to 10p13 , confirming the previous results of somatic cell hybrid mapping local izing RSU-1 to chromosome 10. Screening of more than 20 human tumor ce ll lines for RSU-1 expression revealed that most cell lines contained abundant RSU-1 RNA and protein. However, the p33 RSU-1 protein was und etectable in the U251 glioblastoma cell line and transfection of a rsu -1 expression vector into U251 cells yielded a cell line in which rsu- 1 was under the control of a regulatable metallothionein promoter. Add ition of Cd2+ to the U251-Rsu-1 transfectant resulted in transcription of rsu-1 RNA and the accumulation of p33 Rsu-1 protein. Appearance of the Rsu-1 protein correlated with a reduction in growth rate of the U 251-Rsu-1 transfectant. In addition, reduction in anchorage independen t growth and phenotypic alteration in U251-Rsu-1 transfectant agar col onies was observed. Two U251-Rsu-1 transfectant cell lines were non tu morigenic when injected subcutaneously into athymic nude mice. These r esults, in conjunction with the frequent deletions observed in chromos ome 10 in glioblastomas, suggest that RSU-1 loss of function may play a role in the progression of this disease.