Dependence of centriole formation on protein synthesis.

Abstract

Centriole formation was studied after inhibition of protein synthesis for various portions of the cell cycle. Synchronous populations of mitotic L929 (mouse) cells were plated into petri dishes and the course of procentriole formation was monitored by electron microscope analysis. The frequency with which procentrioles were seen in association with mature centrioles normally increased steadily in the interval from 4 to 12 h after mitosis. The formation of procentrioles was abruptly inhibited by the addition of cycloheximide at any time from mitosis until 12 h postmitosis (S phase). This suggested that the formation of procentrioles was dependent upon protein synthesis immediately before their appearance. Prophase-associated elongation of procentrioles appeared to occur normally in cells treated with cycloheximide for up to 4 h before prophase, though the mitotic index in treated cultures decreased somewhat. Thus, protein synthesis did not appear to be essential for procentriolar elongation to the mature length.The synthesis of centrioles is under tight control by the cell. Generally, a cell makes exactly two centrioles in the course of a cell cycle. Greater numbers of centrioles may be generated if needed as basal bodies when the cell is preparing to generate cilia (5, 10, 25). The time of centriole production has been worked out in proliferating cultured fibroblastic cells (15, 16; see reference 3 for review). It is found that each daughter cell of a division receives a pair of orthogonally arranged, full-length (about 0.55 /zm long) centrioles. The two centrioles separate from each other in G1 and, near the onset of S phase, one daughter procentriole appears adjacent and perpendicular to each mature centriole. The procentrioles are about half the length of a mature centriole and remain that length until prophase, when they elongate to the mature length (15,17). Thus, the pair of centrioles at the pole of the metaphase spindle again consists of two full-length centrioles. The mechanism by which the cell maintains such tight control on the time of centriole duplication and the quantity and size of procentrioles is not known.In an earlier study (15), we used an inhibitor of DNA synthesis to dissociate the event of nuclear DNA synthesis from procentriole production. It appeared that procentriole formation occurred independently of whether or not DNA synthesis was allowed to proceed. On the other hand, the process of elongation of procentrioles to their mature length appeared to be strictly correlated with the onset of prophase events. In this paper, we present the results of experiments which test the requirements for protein synthesis in procentriole

RNA polymerase II transcription is required for human papillomavirus type 16 E7- and hydroxyurea-induced centriole overduplication

“…We found that simultaneous treatment of U2OS/centrin-GFP cells with 1 mM HU and 5 mg/ml CHX for 72 h efficiently reduced the proportion of cells with centriole overduplication from 45 to 4.7% (9.6-fold; Pp0.0005; Figure 3a). In contrast to a previous report (Phillips and Rattner, 1976), we found an increase of cells with duplicated centrioles in asynchronously growing populations that were treated with CHX alone. Such increase was not seen in cells treated with a-amanitin alone or transfected with CBP siRNAs and may be related to the fact that CHX can induce a G2 cell cycle arrest leading to an accumulation of cells with duplicated centrioles.…”

Protein synthesis and the cell cycle: centrosome reproduction in sea urchin eggs is not under translational control.

“…The report that protein synthesis in late G~ is required for daughter centriole formation in cultured cells (20), may raise questions about the applicability of our results to somatic cells. In this regard, the important finding of our study is that the synthesis of new proteins, such as structural components or regulatory enzymes, at each cell cycle is not a strategy used by all cells to control centrosome reproduction.…”

“…For cultured L929 (mouse) cells, at least 4 h of protein synthesis in late G, is required for procentriole formation (20). However, this study is subject to the criticism that inhibition of protein synthesis arrests the cell cycle before the point when procentriole assembly is scheduled to occur.…”

The arithmetic of centrosome biogenesis

“…Because multiple duplication cycles proceed in X. laevis or sea urchin embryos in which protein synthesis is prevented (Gard et al, 1990;Sluder et al, 1990), centrosome duplication does not necessarily require a given protein to be synthesized at each cell cycle, even though protein synthesis is required for duplication in somatic cells (Phillips and Rattner, 1976). Since centrosome duplication is coupled to DNA replication, S phase might be permissive for duplication and, by extension, centrosome duplication might take place once per cell cycle merely because there is a single S phase.…”