COPPER-DEPENDENT METABOLISM OF CU,ZN-SUPEROXIDE DISMUTASE IN HUMAN K562 CELLS - LACK OF SPECIFIC TRANSCRIPTIONAL ACTIVATION AND ACCUMULATION OF A PARTIALLY INACTIVATED ENZYME

The regulation of Cu,Zn-superoxide dismutase by copper was investigate d in human K562 cells. Copper ions caused a dose- and time-dependent i ncrease, up to 3-fold, of the steady-state level of Cu,Zu-superoxide d ismutase mRNA. A comparable increase was also observed for actin and r ibosomal protein L32 mRNAs, but not for metallothionein mRNA which was augmented more than 50-fold and showed a different induction pattern. The copper-induced mRNAs were actively translated as judged from thei r enhanced loading on polysomes, the concomitantly increased cellular protein levels and an augmented incorporation of [H-3]lysine into acid -precipitable material. Cu,Zn-superoxide dismutase protein followed th is general trend, as demonstrated by dose- and time-dependent increase s in immunoreactive and enzymically active protein. However, a specifi c accumulation of Cu,Zn-superoxide dismutase was noticed in cells grow n in the presence of copper, that was not detectable for other protein s. Purification of the enzyme demonstrated that Cu,Zn-superoxide dismu tase was present as a reconstitutable, copper-deficient protein with h igh specific activity (k(cat)/Cu = 0.89 x 10(9) M(-1).s(-1)) in untrea ted K562 cells and as a fully metallated protein with low specific act ivity (k(cat)/Cu = 0.54 x 10(9) M(-1).s(-1)) in copper-treated cells. Pulse-chase experiments using [H-3]lysine indicated that turnover rate s of Cu,Zn-superoxide dismutase in K562 cells were not affected by gro wth in copper-enriched medium, whereas turnover of total protein was s ignificantly enhanced as a function of metal supplementation. From the se results we conclude that: (i) unlike in yeast [Carri, Galiazzo, Cir iolo and Rotilio (1991) FEBS Lett. 278, 263-266] Cu,Zn-superoxide dism utase is not specifically regulated by copper at the transcriptional l evel in human K562 cells, suggesting that this type of regulation has not been conserved during the evolution of higher eukaryotes; (ii) cop per ions cause an inactivation of the enzyme in intact K562 cells; and (iii) the metabolic stability of Cu,Zn-superoxide dismutase results i n its relative accumulation under conditions that lead to increased pr otein turnover.