Thioguanine-resistant primary clones were grown from single cell suspensions obtained from dog and human kidneys by enzymatic digestion. In medium containing a relatively high concentration (10μg/ ml) of thioguanine, thioguanine-resistant primary clones arose from each source at frequencies ranging from 10−4 to 10−5. A reduction in total hypoxanthine uptake was found in the thioguanine-resistant primary clones which had developed in thioguanine medium, consistent with a reduction in hypoxanthine phosphoribosyltransferase activity. When these thioguanine-resistant primary clones were subsequently grown in the absence of thioguanine and assayed for the thioguanine-resistant phenotype and hypoxanthine phosphoribosyltransferase activity, it was found that most were now thioguanine-sensitive and yielded cell free extracts with substantial amounts of hypoxanthine phosphoribosyltransferase activity. In contrast, thioguanine-resistant human clones grown continuously in the presence of thioguanine yielded cell free extracts with little or no detectable hypoxanthine phosphoribosyltransferase activity. Southern blot analysis demonstrated no structural alterations in the hypoxanthine phosphoribosyltransferase gene in thioguanine-resistant primary human kidney clones. These results suggest that a novel mechanism(s) for thioguanine resistance and the control of hypoxanth phosphoribosyltransferase expression may occur in dog and human kidney cells.
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Abbreviations
- AG:
-
8-azaguanine
- APRT:
-
adenine phosphoribosyltransferase
- DAPI:
-
4′-6 diamino-2-phenylindole
- DV:
-
Dulbecco-Vogt
- HAT:
-
hypoxanthine, aminopterin, thymidine
- HPRT:
-
hypoxanthine phosphoribosyltransferase
- PRPP:
-
5-phosphoribosyl 1-pyrophosphate
- TG:
-
6-thioguanine
- TGr :
-
thioguanine-resistant
- TGs :
-
thioguanine-sensitive
- TIP:
-
thymidine triphosphate
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Turker, M.S., Monnat, R.J., Fukuchi, KI. et al. A novel class of unstable 6-thioguanine-resistant cells from dog and human kidneys. Cell Biol Toxicol 4, 211–223 (1988). https://doi.org/10.1007/BF00119247
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DOI: https://doi.org/10.1007/BF00119247