The Activity of the Pyrimidine Biosynthetic Pathway in MGH-U1 Transitional Carcinoma Cells Grown in Tissue Culture

Abstract

MGH-U1, a new cell line of transitional carcinoma grown in tissue culture, is described. Growth characteristics, growth kinetics and activity of the pyrimidine biosynthetic pathway are outlined. Pathway activity was measured in intact MGH-U1 cells and 4 of the biosynthetic enzymes were assayed in cellular extracts. The level of activity of the pathway remains unaltered in cultures that range in density from 104-105 cells/cm2 and the cells actively incorporate precursors into macromolecules throughout the entire range of densities investigated. These results differ from those reported previously, in which the synthesis of RNA, DNA and protein is reduced more than 90% in stationary suspension cultures of Novikoff rat hepatoma cells and confluent monolayers of human fibroblasts that exhibit contact inhibition. The levels of 3 of the 4 enzymes assayed in MGH-U1 cells are reduced 50-60% when the cells undergo the transition from rapid exponential growth to confluence but are still present in levels that are adequate to sustain a high level of pyrimidine biosynthesis. The compound, N-(phosphonacetyl)-L-aspartate inhibited aspartate transcarbamylase, the 2nd enzyme of the pathway in cellular extracts of MGH-U1 cells and retarded the growth of these cells when added to the culture medium. The latter effect could be circumvented by the simultaneous administration of uridine, which is used via salvage pathways for the continuing requirement of the cells for pyrimidine nucleotides.