Sporulation-deficient mutants were isolated from a homothallic strain of Saccharomyces cerevisiae. Sporulation was induced in these mutants by procedures to sporulate the products of protoplast fusion between mutants and wild-type strains. Spores formed in this way were crossed to wild-type strains in order to analyze them genetically. Twenty-three genes essential to sporulation were identified by tetrad analysis and complementation tests. Gene symbols spoT1 to spoT23 were tentatively assigned to them. These mutants fell into four classes by examination of premeiotic DNA synthesis and meiotic nuclear division: (i) Premeiotic DNA synthesis did not occur (spoT1 - spoT11); (ii) premeiotic DNA synthesis occurred but meiosis I did not occur (spoT12 - spoT15); (iii) meiosis II did not occur (spoT16 - spoT18); (iv) meiosis II occurred but mature spores were not formed (spoT19 - spoT23). Genes spoT4, spoT8, spoT20, and spoT23 were mapped on chromosomes IV, II, XVI and XI, respectively. SpoT18-1 was a UAG nonsense mutation.

Of the genes involved in galactose metabolism, GAL7, GAL10, and GAL1 are tightly linked in this order on chromosome II in Saccharomyces cerevisiae. While several species of the order Saccharomycetales have similar gene organization, Kazachstania naganishii is unique, in which GAL7 and GAL1 are close to each other whereas GAL10 is substantially apart from them on chromosome XI. In this study, we inserted the recognition sequence of I-SceI homing-endonuclease into GAL10 and also into the intervening segment of GAL7-GAL1. By cleaving chromosome DNA of the gene-manipulated strain with I-SceI, we obtained evidence that chromosome XI (610 kbp) was replaced with three fragments (305, 265, and 40 kbp). Using appropriate probes, we further found that GAL10 was about 40 kbp apart from the GAL7-GAL1 cluster and that orientation of GAL10 was reversed comparing to the S. cerevisiae counter part. We, therefore, contend that comparison of the organization of the GAL cluster among Saccharomycetales is of importance to elucidate evolution of chromosomes and that the experimental scheme developed in this study is useful for this line of investigation.

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The turnover of nucleic acids and changes in ribonuclease activity during sporulation of Saccharomyces cerevisiae were studied. In the sporulating strains, 37-58% of vegatatively synthesized RNA were degraded during the sporulation process. The degree of degradation of vegetative RNA was proportional to the sporulation ability. In the non-sporulating strains, the degradation of vegetative RNA was less than 28% in the sporulation medium. Accompanied by the degradation of vegetative RNA, a ribonuclease activity increased several times during sporulation. We have found a close relation among the sporulation rate, the degree of the degradation of vegetative RNA and the increase in ribonuclease activity in the sporulation medium, using cells of which sporulation ability was repressed by changing the age or carbon source in various degrees.

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