Centriole Number of Spindle Poles and the Reproductive Capacity

The reproduction of spindle poles is a key event in the cell's preparation for mitosis. To gain further insight into how this process is controlled, we systematically characterized the ultrastructure of spindle poles whose reproductive capacity had been experimentally altered. In particular, we wanted to determine if the ability of a pole to reproduce before the next division is related to the number of centrioles it contains. We used mercaptoethanol to indirectly induce the formation of monopolar spindles in sea urchin eggs. We followed individually treated eggs in vivo with a polarizing microscope during the induction and development of monopolar spindles. We then fixed each egg at one of three predetermined key stages and serially semithick sectioned it for observation in a high-voltage electron microscope. We thus know the history of each egg before fixation and, from earlier studies, what that cell would have done had it not been fixed. We found that spindle poles that would have given rise to monopolar spindles at the next mitosis have only one centriole whereas spindle poles that would have formed bipolar spindles at the next division have two centrioles. By serially sectioning each egg, we were able to count all centrioles present. In the twelve cells examined, we found no cases of acentriolar spindle poles or centriole reduplication. Thus, the reproductive capacity of a spindle pole is linked to the number of centrioles it contains. Our experimental results also show, contrary to existing reports, that the daughter centriole of a centrosome can acquire pericentriolar material without first becoming a parent. Furthermore, our results demonstrate that the splitting apart of mother and daughter centrioles is an event that is distinct from, and not dependent on, centriole duplication. At the end of mitosis, each new daughter cell receives a single spindle pole from the parent cell and by the start of the next mitosis, each of these cells has two, and only two, spindle poles. This reproduction of spindle poles must be tightly controlled by the cell since the wrong number of poles would inevitably lead to aneuploidy and a consequent loss of viability for the progeny of that cell. The mechanisms that are involved in the precise doubling of the spindle poles before each mitosis are poorly understood. A number of studies have shown that the reproduction of spindle poles can be experimentally manipulated in sea urchin eggs (15, 28, 29a). When the first mitosis of the fertilized egg is prolonged by any of several techniques (mercaptoethanol, colcemid, or micromanipulation), the two existing spindle poles are observed to split and separate to form a tetrapolar spindle (29a). Although these four poles appear normal, the subsequent development of the daughter cells shows that these poles have only half the reproductive capacity of normal poles. Each of the four daughters that result from the tetrapolar division form only a monopolar spindle at the next mitosis. These monopolar spindles are truly halfofa spindle because two of them can come together to give a bipolar spindle of normal appearance and function (15, 29). Subsequent divisions of the progeny of such cells are normal. Thus, monopolar spindles have one pole with full reproductive capacity. When a monopolar spindle is formed individually in its own cell, that cell often spends significantly more time in mitosis than normal (29). In such cases, the single pole is observed to split to form two poles that separate and transform the monopolar spindle into a bipolar spindle of normal appearance and function (28, 29a). Although the two new poles appear normal, they have only half the normal reproductive THE JOURNAL OF CELL BIOLOGY . VOLUME 100 MARCH 1985 887-896 © The Rockefeller University Press • 0021-9525[85/03[0887/10 $1.00 8 8 7 on Jne 1, 2017 D ow nladed fom Published March 1, 1985