HERBERT WEINFELD , and AVERY

Using the cultured Chinese hamster cell line Don, G1 or S or a mixture of late-S/G2 cells were prepared by release from metaphase arrest. Metaphase (M) cells were also obtained by mitotic arrest of log-phase cultures with Coicemid and held in metaphase; such M cells remained untreated with any other compound and were termed standard M cells. When interphase (I) cells were fused at pH 8.0 and 37~ with standard M cells in the presence of Colcemid by means of UV-inactivated Sendai virus, binucleate interphase-metaphase (I-M) cells were obtained. In a given I-M cell there occurred within 30 min after fusion either prophasing of the I nucleus or formation of a nuclear envelope (NE) around the chromosomes. About 20% of early G1 cells, 35% of cells at the G I /S boundary, 50% of S cells, and 70% of late-S/G2 cells could induce NE formation. If, before fusion, cycloheximide (CHE), an inhibitor of protein synthesis, was present during release from M arrest, the ceils entered G~ but not S. About 20% of such early GI cells, like the untreated early Gx cells, had the capacity to induce NE formation during subsequent fusion. If the cells were blocked in S with 5 mM thymidine (TdR), at least 80% of these cells could induce NE formation during subsequent fusion, but in the presence of both TdR and CHE only 35% could do so. It appeared, therefore, that protein synthesis in interphase was required for NE formation. Experiments with actinomycin D indicated that RNA synthesis was also necessary for acquisition of NE-inducing capacity. About 35% of G1 cells from confluent monolayers had the NE-inducing capacity, but prolonged exposure to CHE reduced their number to 8% Removal of CHE restored the ability while the cells still remained in Gv This result indicated that continuing protein synthesis in the G~ cell was needed for NE formation subsequent to fusion. The fact that macromolecular synthesis must occur in the I cell before fusion if NE formation was to occur in the fused I-M cell lends further support to evidence adduced earlier that this phenomenon is a normal mitotic event. 378 THE JOURNAL OF CELL BIOLOGY . VOLUME 64, 1975 . pages 378 388 on Jne 6, 2017 D ow nladed fom Published February 1, 1975