Green M H
J Virol. 1972 Jul;10(1):32-41. doi: 10.1128/JVI.10.1.32-41.1972.
Under normal growth conditions, all of the newly synthesized polyoma deoxyribonucleic acid (py DNA) that could be extracted from infected mouse cell cultures by the Triton procedure of Green, Miller, and Hendler was in the form of a 55S nucleoprotein complex. Inhibition of protein synthesis by cycloheximide reduced the sedimentation rate of the polyoma complex synthesized during the first hour after addition of the drug to 25 to 35S. Since the 55S and the 25 to 35S complexes each contain closed circular 20S py DNA, it is suggested that the slower complex contains less protein per DNA molecule and that there is normally a small or unstable pool of protein available for binding to newly replicated py DNA. In the presence of cycloheximide, the newly formed 25 to 35S complex was not derived from preexisting 55S complex. Thus, some py DNA which was not solubilized by the Triton method served as a template for replication. Further evidence for the existence of polyoma replication sites is provided by the demonstration that, during the inhibition of protein synthesis, a class of newly replicated py DNA can be solubilized by the sodium dodecyl sulfate procedure of Hirt, but not by the Triton method. It is postulated that continuous protein synthesis is required to release py DNA from replication sites in the form of a Triton-extractable nucleoprotein complex.
在正常生长条件下，通过格林、米勒和亨德勒的Triton方法从感染的小鼠细胞培养物中提取的所有新合成的多瘤脱氧核糖核酸（py DNA）均为55S核蛋白复合物形式。放线菌酮抑制蛋白质合成后，在加入该药物后的第一小时内合成的多瘤复合物的沉降速率降至25至35S。由于55S复合物以及25至35S复合物均包含闭环20S py DNA，因此表明沉降较慢的复合物每个DNA分子所含蛋白质较少，并且通常存在少量或不稳定的蛋白质池可用于与新复制的py DNA结合。在放线菌酮存在的情况下，新形成的25至35S复合物并非源自预先存在的55S复合物。因此，一些未被Triton方法溶解的py DNA充当了复制模板。蛋白质合成受抑制期间，一类新复制的py DNA可通过赫特的十二烷基硫酸钠方法溶解，但不能通过Triton方法溶解，这一现象证明了多瘤复制位点的存在。据推测，需要持续的蛋白质合成才能以Triton可提取的核蛋白复合物形式从复制位点释放py DNA。
