AMBER SUPPRESSION IN MAMMALIAN-CELLS DEPENDENT UPON EXPRESSION OF AN ESCHERICHIA-COLI AMINOACYL-TRANSFER-RNA SYNTHETASE GENE

As an approach to inducible suppression of nonsense mutations in mamma lian and in higher eukaryotic cells, we have analyzed the expression o f an Escherichia coli glutamine-inserting amber suppressor tRNA gene i n COS-1 and CV-1 monkey kidney cells. The tRNA gene used has the suppr essor tRNA coding sequence flanked by sequences derived from a human i nitiator methionine tRNA gene and has two changes in the coding sequen ce. This tRNA gene is transcribed, and the transcript is processed to yield the mature tRNA in COS-1 and CV-1 cells. We show that the tRNA i s not aminoacylated in COS-1 cells by any of the endogenous aminoacyl- tRNA synthetases and is therefore not functional as a suppressor. Conc omitant expression of the E. coli glutaminyl-tRNA synthetase gene resu lts in aminoacylation of the suppressor tRNA and its functioning as a suppressor. These results open up the possibility of attempts at regul ated suppression of nonsense codons in mammalian cells by regulating e xpression of the P. coli glutaminyl-tRNA synthetase gene in an inducib le, cell-type-specific, or developmentally regulated manner.