The isolation and characterization of a mutant murine
T-cell lymphoma (S49) with altered
purine metabolism is described. This mutant, AU-100, was isolated from a mutagenized population of S49 cells by virtue of its resistance to 0.1 mM
6-azauridine in semisolid
agarose. The AU-100 cells are resistant to
adenosine mediated cytotoxicity but are extraordinarily sensitive to killing by
guanosine. High performance liquid chromatography of AU-100
cell extracts has demonstrated that intracellular levels of
GTP,
IMP, and GMP are all elevated about 3-fold over those levels found in wild type cells. The AU-100 cells also contain an elevated intracellular level of pyrophosphoribosylphosphate (PPriboseP), which as in wild type cells is diminished by incubation of AU-100 cells with
adenosine. However AU-100 cells synthesize
purines de novo at a rate less than 35% of that found in wild type cells. In other growth rate experiments, the AU-100 cell line was shown to be resistant to
6-thioguanine and
6-mercaptopurine. Levels of
hypoxanthine-guanine phosphoribosyltransferase (HGPRTase) measured in AU-100
cell extracts, however, are 50-66% greater than those levels of HGPRTase found in wild type
cell extracts. Nevertheless this mutant S49 cell line cannot efficiently incorporate labeled
hypoxanthine into
nucleotides since the salvage
enzyme HGPRTase is inhibited in vivo. The AU-100 cell line was found to be 80% deficient in
adenylosuccinate synthetase, but these cells are not auxotrophic for
adenosine or other
purines. The significant alterations in the control of
purine de novo and salvage metabolism caused by the defect in
adenylosuccinate synthetase are mediated by the resulting increased levels of
guanosine nucleotides.