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Protein and Ribonucleic Acid Metabolism in Single-Cell Suspensions from Morris Hepatoma 5123tc and from Normal Rat Liver¹

Biochemisches Institut [M. S., G. S.] and Institut Biologie II, Department of Cell Biology [J. K.], Universität Freiburg, Freiburg im Breisgau, Germany

The plasma membranes of suspended single cells from Morris hepatomas 5123tc and from liver were freely permeable for L-leucine and L-methionine. Thus, after incubation with L-leucine-l-¹⁴C, net rates of protein synthesis could be calculated from specific radioactivities of precursor and product. 5123tc cells synthesized 1.4 µg protein per hr per mg cellular protein compared with 0.54 µg obtained in liver cells. Incorporation of L-leucine into protein of the hepatoma cells was independent of the concentration of K⁺, Mg²⁺, and Ca²⁺ in the medium, whereas distinct maxima of incorporation rates were observed in liver cells at 65 mM K⁺, 3.5 mM Mg²⁺, and 2.5 mM Ca²⁺. Mitochondrial protein synthesis contributed only a minor portion to total protein synthesis in both cell types. Radioactive protein was transferred into the medium by hepatoma or liver cells incubated with L-leucine-l-¹⁴C with kinetics similar to those found for liver in the intact animal. This is in contrast to the lack of secretion of serum proteins by the hepatoma observed in vivo. The transfer was not inhibited when protein synthesis was interrupted by puromycin or cycloheximide or when the cells were incubated under nitrogen.

Uridine, uracil, and orotic acid were efficiently incorporated into the RNA of hepatoma cells, whereas in liver cells only uridine was incorporated effectively.

The ultrastructure of the cells was well preserved 1 hr after incubation. The conformation of the mitochondria changed during incubation from the condensed to the orthodox state. Hepatoma cells produced tumors in all cases (n = 20) when reimplanted into Buffalo rats.

¹ The experiments were supported by the Deutsche Forschungsgemeinschaft, Bad Godesberg (SFB 46, Freiburg).

² Present address: The Walter and Eliza Hall Institute of Medical Research, Melbourne, Australia.

³ Present address: The Russell Grimwade School of Biochemistry, University of Melbourne, Parkville, Victoria, 3052, Australia.

Received 12/10/73. Accepted 4/12/74.