Cell cycle-associated expression of M2-type isozyme of pyruvate kinase in proliferating rat thymocytes.
During a complete cell cycle of rat thymocytes stimulated by concanavalin A and interleukin 2, the activity and mRNA level of pyruvate kinase reached a maximum (8-12-fold increase) 48 h after stimulation coinciding with the S-phase of the cell cycle. Increases of cellular enzyme activity, pyruvate kinase protein, and mRNA levels are correlated up to 48 h of culture. Afterwards pyruvate kinase activity and mRNA levels decrease, whereas the pyruvate kinase protein continues to increase throughout mitosis. This change of specific pyruvate kinase activity points to a posttranslational modification of the enzyme besides its transcriptional regulation. The presence of the M2-type isozyme was determined by the following methods: (a) native cellulose acetate electrophoresis and activity staining, (b) Northern blot hybridization with M1- and M2-specific cDNA probes, and (c) determination of kinetic parameters. The isozyme pattern did not change during the cell cycle progression. The induction of pyruvate kinase is completely abolished by 2-difluoromethylornithine-mediated polyamine depletion. However, the proportion of hybridizable pyruvate kinase mRNA was not affected. These data suggest the requirement of polyamines for efficient translation rather than transcription during cell growth.
Netzker
R
R
Institute of Biochemistry, Medical Faculty, University of Erlangen-Nuremberg, Federal Republic of Germany.
Greiner
E
E
Eigenbrodt
E
E
Noguchi
T
T
Tanaka
T
T
Brand
K
K
D000818:Animals; D002453:Cell Cycle / Q000502:physiology*; D002455:Cell Division; D002478:Cells, Cultured; D003208:Concanavalin A / Q000494:pharmacology; D000518:Eflornithine / Q000494:pharmacology; D005260:Female; D015151:Immunoblotting; D007527:Isoenzymes / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism*; D007700:Kinetics; D008213:Lymphocyte Activation; D011770:Pyruvate Kinase / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism*; D012333:RNA, Messenger / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D013601:T-Lymphocytes / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology*; D013950:Thymus Gland / Q000201:enzymology*
D007527:Isoenzymes; D012333:RNA, Messenger; D003208:Concanavalin A; D011770:Pyruvate Kinase; D000518:Eflornithine