Suppressor tRNAs in Mammalian Cells

With the objective of isolating animal viruses with nonsense mutations in their genes, we previously established mammalian cell lines carrying an inducible amber suppressor tRNA gene (with Phillip Sharp). We are developing more generally applicable systems for inducible expression of tRNA genes and/or tRNA function. One approach is to introduce genes for suppressor tRNAs which are not substrates for mammalian aminoacyl-tRNA synthetases, and express a cognate aminoacyl-tRNA synthetase gene in an inducible manner. We have shown that amber, ochre and opal suppression in mammalian cells can be made dependent upon expression of the E. coli glutaminyl-tRNA synthetase (GlnRS) gene. We have demonstrated tetracycline-regulated expression of GlnRS and, thereby, tetracycline-regulated suppression of amber codons in mammalian cells. The next step is to isolate mammalian cell lines carrying such tRNAs and aminoacyl-tRNA synthetase genes.