Altered polyamine metabolism in chinese hamster cells growing in a defined medium

✍ Scribed by Gary J. Sertich; James R. Glass; David J. M. Fuller; Eugene W. Gerner

• Publisher: John Wiley and Sons
• Year: 1986
• Tongue: English
• Weight: 799 KB
• Volume: 127
• Category: Article
• ISSN: 0021-9541
• DOI: 10.1002/jcp.1041270115

✦ Synopsis

Chinese hamster cells (line CHO) maintained in McCoy's 5A medium (modified) supplemented with insulin (10 pg/ml), transferrin (5 pglml), and ferrous sulfate (1.1 pg/ml) proliferate at rates similar to cultures growing jn the Mc-Coy's medium supplemented with 10% fetal bovine serum. Colony-forming ability is similar in cultures supplemented with either serum or the combination of growth factors. By 6 hours after replacement of serum with growth factors, ornithine decarboxylase (ODCase) activity increases, reaching a max- imum value by 24 hours after serum replacement. This maximum is cell density dependent and can exceed a 30-fold increase over enzyme activity in cultures supplemented with serum. T h e increased enzyme activity is due to a decrease in t h e turnover rate of t h e enzyme, based on protein synthesis inhibition studies, and an accumulation of active enzyme molecules rather than an activation of existing molecules, since the catalytic activity of ODCase, determined using the radiolabeled form of a-difluoromethylornithine (an enzyme-activated, irreversible inhibitor of ODCase) in concert with immunochemical techniques, is unchanged by the different growth medium supplements. lntracellular putrescine and sperrnidine levels are substantially decreased when cultures are maintained in medium supplemented with insulin, transferrin, and ferrous sulfate, rather than serum, which is the sole source of exogenous ornithine. Titration of cultures growing in the defined medium with ornithine leads to a decrease in ODCase activity and an increase in intracellular putrescine and spermidine levels. Putrescineand spermidinedependent S-adenosyl-L-methionine decarboxylase activities are similar in cultures maintained in either medium. These data demonstrate that some, but not all, aspects of polyamine biosynthesis are affected by the availability of ornithine, the first substrate in the pathway.