Centriole cycle in Chinese hamster ovary cells as determined by whole-mount electron microscopy

Abstract

In interphase Chinese hamster ovary (CHO) cells, the centrosome is attached to the nucleus very firmly. This nuclear-centrosome complex is isolated as a coherent structure by lysis and extraction of cells with Triton X-100 in a low ionic strength medium. Under these conditions, the ultrastructure of the centrioles attached to the nucleus can be discerned by electron microscopy of whole-mount preparations. The structural changes of the centrioles as a function of the cell cycle were monitored by this technique. Specifically, centriolar profiles were placed into six categories according to their orientation and the length ratio of daughter and parent centrioles. The proportion of centrioles in each category was plotted as a frequency histogram. The morphological changes in the centriole cycle were characterized by three distinguishable events: nucleation, elongation, and disorientation. The progress of centrioles through these stages was determined in synchronous populations of cells starting from S or M phase, in cells inhibited in DNA synthesis by addition of thymidine, and in cytoplasts. The results provide a quantitative description of the events of the centriole cycle. They also show that, in complete cells, nucleation, elongation, and disorientation are not dependent upon DNA synthesis. However, in cytoplasts, although elongation and disorientation occur as in normal cells, nucleation is blocked. Procentriole formation appeared to be inhibited by the removal of the nucleus. We suggest that coordination of centriole replication and nuclear replication may depend upon a signal arising from the nucleus.