Cell, Vol. 60, 1009-1017, March 23, 1990, Copyright 0 1990 by Cell Press Segregation of Recombined Chromosomes in Meiosis I Requires DNA Topoisomerase II David Rose, Winston Thomas, and Connie Holm Department of Cellular and Developmental Biology Harvard University Cambridge, Massachusetts 02138 Summary To understand better the similarities and differences between meiosis and mitosis, we examined the mei- otic role of DNA topoisomerase II, an enzyme that is required mltotlcally to disentangle sister chroma- tids at the time of chromosome segregation. In meio- sis, we found that topolsomerase II is required only at the time of nuclear division. When cold-sensitive fop2 mutants are induced to sporulate at the restrictive temperature, they undergo premeiotlc DNA synthesis and commitment to meiotic levels of recombination but fail to complete the first meiotic nuclear division. The introduction of a mutation blocking recombina- tion relieves the requirement for topoisomerare II in meiosis I. These results suggest that topoisomerase II is required at the time of chromosome segregation in meiosis I for the resolution of recombined chromo- somes. Introduction The processes of both meiosis and mitosis require that chromosomes be replicated faithfully and segregated pre- cisely in microtubule-mediated nuclear divisions. There are, however, important differences between the two processes. In contrast to the single mitotic nuclear divi- sion, meiosis comprises two chromosome segregation events. Premeiotic DNA synthesis also differs from premi- totic DNA synthesis, as manifested in part by its extended duration (Williamson et al., 1983). In addition, meiotic and mitotic recombination require different gene products (for review, see Haynes and Kunz, 1981) and occur at different levels. Study of the effects of single mutations on both mitosis and meiosis is frequently informative. For example, muta- tions in the structural gene for 8-tubulin confer similar mi- totic and meiotic phenotypes, blocking both mitosis and sporulation (Huffaker et al., 1988). Mutations in the RADSO gene, on the other hand, have strikingly different meiotic and mitotic phenotypes. tad50 mutants fail to un- dergo meiotic recombination (Game et al., 1980) but in mi- tosis are hyper-recombinant and deficient in DNA repair (Haynes and Kunz, 1981). These studies point out impor- tant similarities and differences between mitosis and mei- osis and also provide a more complete picture of the func- tion of the gene product examined. With these studies as a background, we compared the roles of DNA topoisomer- ase II in mitosis and meiosis. The role of DNA topoisomerase II in mitosis is well un- derstood. DNA topoisomerase II acts by making a double- strand break in a DNA molecule, passing another double- stranded segment through the break, and then resealing it (Brown and Cozzarelli, 1979; Liu et al., 1980). In this way, DNA topoisomerase II can disentangle intertwined DNA molecules and can relax both positively and negatively supercoiled DNA (Baldi et al., 1980; Hsieh, 1983; Hsieh and Brutlag, 1980; Kreuzer and Cozzarelli, 1980; Liu et al., 1981). In yeast, temperature-sensitive top2 mutants al- lowed to progress through mitosis at the restrictive tem- perature suffer a high level of inviability, which is accom- panied by elevated levels of chromosome breakage and nondisjunction (Holm et al., 1985, 1989; Uemura and Yanagida, 1988; Uemura et al., 1987). Inviability can be prevented by treatment with the microtubule-destabilizing drug nocodazole (Holm et al., 1985) indicating that the loss in viability is specifically associated with cells at- tempting to segregate their chromosomes in the absence of topoisomerase II. These observations have led to the hypothesis that topoisomerase II facilitates chromosome segregation in mitosis by resolving entangled sister chro- matids, which are intertwined to some extent following the completion of DNA synthesis (Sundin and Varshavsky, 1980, 1981; DiNardo et al., 1984; Weaver et al., 1985). Because topoisomerase II is required for the resolution of intertwined sister chromatids in mitosis, we anticipated that it might play an analogous role in meiosis. In particu- lar, the second meiotic nuclear division is very similar to a mitotic nuclear division in that each involves the segre- gation of sister chromatids. Meiosis I, on the other hand, differs from both meiosis II and mitosis in that it involves the segregation of homologous chromosomes. Because homologous chromosomes are not replicative sisters, topoisomerase Ii may not be required for their separation. However, topoisomerase II may be required for other processes in meiosis I, such as premeiotic DNA synthe- sis, initiation of meiotic recombination, or the resolution of recombined DNA molecules. We report here that DNA topoisomerase II is required for the meiotic nuclear divisions. Cold-sensitive top2 mu- tants induced to sporulate at the restrictive temperature undergo premeiotic DNA synthesis and commitment to meiotic levels of recombination but arrest before the first meiotic nuclear division. Our results suggest that DNA topoisomerase II is required for the resolution of recom- bined chromosomes at the time of chromosome segrega- tion in meiosis I. Results DNA Topoisomerase II Is Required for Sporulatlon We first determined that topoisomerase II is required for the completion of sporulation, which includes both the process of meiosis and the encapsulation of meiotic prod- ucts in spore walls. TCP2+/7?F2+ and top2cs/top2cs strains were induced to sporulate at both the permissive tempera- ture (25%) and the restrictive temperature (10%). Although TCR?+/Topr strains exhibited only a slight reduction in