Replication-dependent early meiotic requirement for Spo11 and Rad50.

Spo11 and the Rad50-Mre11 complex have been indirectly implicated in processes associated with DNA replication. These proteins also have been shown to have early meiotic roles essential for the formation of a programmed DNA double-strand break known in Saccharomyces cerevisiae to initiate meiotic recombination. In both S. cerevisiae and the basidiomycete Coprinus cinereus, spo11 and rad50 mutants are defective in chromosome synapsis during meiosis. Here we demonstrate that a partial restoration of synapsis occurs in C. cinereus spo11 and rad50 mutants if premeiotic DNA replication is prevented. Double mutants were constructed with spo11-1 or rad50-4 and another mutant, spo22-1, which does not undergo premeiotic DNA replication. In both cases, we observed an increase in the percentage of nuclei containing synaptonemal complex (SC) structures, with concomitant decreases in the percentage of nuclei containing axial elements (AE) only or no structures. Both types of double mutants demonstrated significant increases in the average numbers of AE and SC, although SC-containing nuclei did not on average contain more AE than did nuclei showing no synapsis. Our results show that Spo11-induced recombination is not absolutely required for synapsis in C. cinereus, and that the early meiotic role of both Spo11 and Rad50 in SC formation partially depends on premeiotic S phase. This dependency likely reflects either a requirement for these proteins imposed by the premeiotic replication process itself or a requirement for these proteins in synapsis when a sister chromatid (the outcome of DNA replication) is present.

Multiple roles of Spo11 in meiotic chromosome behavior.

Spo11, a type II topoisomerase, is likely to be required universally for initiation of meiotic recombination. However, a dichotomy exists between budding yeast and the animals Caenorhabditis elegans and Drosophila melanogaster with respect to additional roles of Spo11 in meiosis. In Saccharomyces cerevisiae, Spo11 is required for homolog pairing, as well as axial element (AE) and synaptonemal complex (SC) formation. All of these functions are Spo11 independent in C.elegans and D.melanogaster. We examined Spo11 function in a multicellular fungus, Coprinus cinereus. The C.cinereus spo11-1 mutant shows high levels of homolog pairing and occasionally forms full-length AEs, but no SC. In C.cinereus, Spo11 is also required for maintenance of meiotic chromosome condensation and proper spindle formation. Meiotic progression in spo11-1 is aberrant; late in meiosis basidia undergo programmed cell death (PCD). To our knowledge, this is the first example of meiotic PCD outside the animal kingdom. Ionizing radiation can partially rescue spo11-1 for both AE and SC formation and viable spore production, suggesting that the double-strand break function of Spo11 is conserved and is required for these functions.

A putative rhamnogalacturonase required for sexual development of Neurospora crassa.

In previous work, the asd-I (ascus development) gene of the filamentous fingus Neurospora crassa was identified as a gene expressed preferentially during the sexual cycle and shown to be essential for normal sexual development. The asd-I gene has been sequenced and further characterized. It contains two introns, the first of which is in-frame and inefficiently or differentially spliced. The predicted ASD-I protein has extensive homology with rhamnogalacturonase B of Aspergillus aculeatus, which cleaves the backbone within the ramified hairy regions of pectin. In homozygous asd-I crosses, sexual development is initiated and large numbers of normal-sized asci are formed. Ascospore delineation does not occur, however, and no sexual progeny are produced. As most asd-I asci contain eight nuclei, the two meiotic divisions and subsequent mitotic division typical of normal crosses seem to occur, but the haploid nuclei are not partitioned into ascospores. In wild-type crosses, the ASD-I protein is present in large amounts in croziers and young asci, but it is only faintly detectable in more mature asci containing developing ascospores. Models to explain the possible role of a rhamnogalacturonase in sexual development are presented.

Pseudohomothallism and evolution of the mating-type chromosome in Neurospora tetrasperma.

Ascospores of Neurospora tetrasperma normally contain nuclei of both mating-type idiomorphs (a and A), resulting in self-fertile heterokaryons (a type of sexual reproduction termed pseudohomothallism). Occasional homokaryotic self-sterile strains (either a or A) behave as heterothallics and, in principle, provide N. tetrasperma with a means for facultative outcrossing. This study was conceived as an investigation of the population biology of N. tetrasperma to assess levels of intrastrain heterokaryosis (heterozygosity). The unexpected result was that the mating-type chromosome and autosomes exhibited very different patterns of evolution, apparently because of suppressed recombination between mating-type chromosomes. Analysis of sequences on the mating-type chromosomes of wild-collected self-fertile strains revealed high levels of genetic variability between sibling A and a nuclei. In contrast, sequences on autosomes of sibling A and a nuclei exhibited nearly complete homogeneity. Conservation of distinct haplotype combinations on A and a mating-type chromosomes in strains from diverse locations further suggested an absence of recombination over substantial periods of evolutionary time. The suppression of recombination on the N. tetrasperma mating-type chromosome, expected to ensure a high frequency of self fertility, presents an interesting parallel with, and possible model for studying aspects of, the evolution of mammalian sex chromosomes.