Protein turnover, growth and proliferation in CHO cells. Variation within and between mutant classes for salvage pathway enzymes.
We have examined the clonal variation in rates of amino acid transport, protein synthesis, protein degradation, growth and proliferation for CHO cells with mutations in the purine and pyrimidine salvage pathways. First we compared three clonal cell lines, each with a different mutation, with the heterozygous parental line AT3-2. Overall, the correlation between rates of protein turnover, growth and proliferation was excellent. The slower growth and proliferation of one mutant, AB3 (TK-, APRT-), is explained by a low intrinsic rate of protein synthesis coupled with a smaller response in rates of amino acid transport, protein synthesis and protein degradation to insulin, serum and dexamethasone. Secondly, we compared seven aza-adenine-resistant and 14 thioguanine-resistant mutants of AT3-2 and found significant differences in control and insulin-stimulated rates of protein turnover both within and between mutant populations. A significant difference between the populations was unexpected because each individual cell line was cloned from a spontaneous pre-existing mutation in AT3-2, and each population should have the same average rate. Remarkably, all 24 mutants had lower rates of protein synthesis than AT3-2. We cannot explain the data solely in terms of mutations in the salvage pathways. Rather, we propose that the mutant survivors have randomly down-regulated the intrinsically fixed growth factor-regulated pathways of protein turnover, resulting in a broad spectrum of lower metabolic rates.
Gunn
J M
JM
Department of Biochemistry and Biophysics, Texas A & M University, College Station 77843-2128.
Brancheau
M R
MR
D000228:Adenine Phosphoribosyltransferase / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D016466:CHO Cells; D002455:Cell Division; D002999:Clone Cells; D006224:Cricetinae; D003470:Culture Media; D003907:Dexamethasone / Q000494:pharmacology; D014644:Genetic Variation; D007328:Insulin / Q000494:pharmacology; D007700:Kinetics; D007930:Leucine / Q000378:metabolism; D014176:Protein Biosynthesis; D011506:Proteins / Q000378:metabolism*; D011865:Radioisotope Dilution Technique; D013937:Thymidine Kinase / Q000235:genetics / Q000378:metabolism*; D014316:Tritium
D003470:Culture Media; D007328:Insulin; D011506:Proteins; D014316:Tritium; D003907:Dexamethasone; D000228:Adenine Phosphoribosyltransferase; D013937:Thymidine Kinase; D007930:Leucine