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Hoffmann, R. A wiki for the life sciences where authorship matters. Nature Genetics (2008)

Subcellular fate of protein antibiotic neocarzinostatin in culture of a lymphoid cell line from Burkitt's lymphoma.

14C-LABELED PROTEIN ANTIBIOTIC NEOCARZINOSTATIN (NCS) was prepared efficiently by chemical modification. With the use of lymphoma-derived cell line P3HR-1, the subcellular behavior of this antitumor antibiotic was studied by the uptake a. nd autoradiography of isolated nuclei of radioactive NCS. The antibiotic was taken up by the cells, reaching the maximum value at 1.5 hr and decreasing in value at 4.0 hr to the level at 0.5 hr. The silver grains in the autoradiograms were also found in the isolated nuclei. The grain count in the nuclei showed a tendency similar to the uptake of NCS by the whole cells, i.e., a gradual increase at 0.5 hr, reaching the maximum value at 1.5 hr, and then decreasing after 4.0 hr to the level at 0.5 hr. These facts indicated that NCS reached not only to cytosol but also into the nucleus, and/or at least to the nuclear membrane of the lymphoid cell. The number of NCS molecules incorporated into the cells at 1.5 hr was calculated to be about 1 x 10-6/cells at a concentration of 3 mug NCS per ml of medium, which can be extrapolated to 1 x 10-4 molecules per cell at the minimum inhibitory concentration. The number of molecules should be even less within the nucleus. In cell-free systems, the interaction of DNA and NCS, which is an inhibitor of DNA synthesis, was investigated with use of a Sephadex G-100 column, with negative results. In the cell culture system, NCS molecules were degraded into smaller polypeptides of certain sizes by proteolysis either by serum component(s) or by cells themselves. An inactive isomer, pre-NCS, which is an antagonist of NCS and a partially denatured homologous molecule, behaved similarly to NCS in all of these experiments. Because the chemically modified NCS used in this study retained biological activity essentially similar to that of parental NCS, the results obtained here could be interpreted as similar to those of parental NCS in vitro.[1]


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