Spermatozoa Production by Triploid Males in the New Zealand Freshwater Snail .

Asexual lineages derived from dioecious taxa are typically assumed to be all female. Even so, asexual females from a variety of animal taxa occasionally produce males. The existence of these males sets the stage for potential gene flow across asexual lineages as well as between sexual and asexual lineages.

A mutation in the FHA domain of Coprinus cinereus Nbs1 Leads to Spo11-independent meiotic recombination and chromosome segregation.

Nbs1, a core component of the Mre11-Rad50-Nbs1 complex, plays an essential role in the cellular response to DNA double-strand breaks (DSBs) and poorly understood roles in meiosis. We used the basidiomycete Coprinus cinereus to examine the meiotic roles of Nbs1. We identified the C.

5'-Serial Analysis of Gene Expression studies reveal a transcriptomic switch during fruiting body development in Coprinopsis cinerea.

Background: The transition from the vegetative mycelium to the primordium during fruiting body development is the most complex and critical developmental event in the life cycle of many basidiomycete fungi. Understanding the molecular mechanisms underlying this process has long been a goal of research on basidiomycetes. Large scale assessment of the expressed transcriptomes of these developmental stages will facilitate the generation of a more comprehensive picture of the mushroom fruiting process.

Global gene expression in Coprinopsis cinerea meiotic mutants reflects checkpoint arrest.

The basidiomycete Coprinopsis cinerea is well-suited to studies of meiosis because meiosis progresses synchronously in 10 million cells within each mushroom cap. Approximately 20% of C. cinerea genes exhibit changing expression during meiosis, but meiosis and mushroom development happen concurrently and therefore differentially expressed genes might not be directly involved in meiotic processes.

The spread of a transposon insertion in Rec8 is associated with obligate asexuality in Daphnia.

Although transitions from sexual to asexual reproduction are thought to have important evolutionary consequences, little is known about the mechanistic underpinnings of these changes. The cyclical parthenogen Daphnia pulex is a powerful model in which to address these issues because female-limited meiosis suppression can be transmitted to sexual individuals via males, providing the opportunity for genetic dissection of the trait. A previous study identified genomic regions differentiating obligately asexual females from their sexual counterparts, and a candidate gene within one such region, encoding the meiotic cohesin Rec8, is the subject of this investigation.

Analysis of the Basidiomycete Coprinopsis cinerea reveals conservation of the core meiotic expression program over half a billion years of evolution.

Coprinopsis cinerea (also known as Coprinus cinereus) is a multicellular basidiomycete mushroom particularly suited to the study of meiosis due to its synchronous meiotic development and prolonged prophase. We examined the 15-hour meiotic transcriptional program of C. cinerea, encompassing time points prior to haploid nuclear fusion though tetrad formation, using a 70-mer oligonucleotide microarray.

Insights into evolution of multicellular fungi from the assembled chromosomes of the mushroom Coprinopsis cinerea (Coprinus cinereus).

The mushroom Coprinopsis cinerea is a classic experimental model for multicellular development in fungi because it grows on defined media, completes its life cycle in 2 weeks, produces some 10(8) synchronized meiocytes, and can be manipulated at all stages in development by mutation and transformation. The 37-megabase genome of C. cinerea was sequenced and assembled into 13 chromosomes.

Methods for meiotic chromosome preparation, immunofluorescence, and fluorescence in situ hybridization in Daphnia pulex.

The genus Daphnia has an intriguing reproductive mode of cyclical parthenogenesis. This reproductive mode has been studied for centuries, but cytogenetic information is lacking due to technical limitations of classical methods. We have developed methods for the preparation and examination of meiotic chromosomes of Daphnia pulex from oocytes and spermatocytes.

Meiotic cytogenetics in Coprinus cinereus.

The basidiomycete fungus Coprinus cinereus has naturally synchronous meiosis and is amenable to analysis using an array of well-developed genetic and molecular tools. In this chapter, we explain in detail the two methods most commonly employed for C. cinereus, staining of intact gill segments and chromosome spreads, with an example of the application of each.

Meiotic localization of Mre11 and Rad50 in wild type, spo11-1, and MRN complex mutants of Coprinus cinereus.

The Mre11-Rad50-Nbs1 (MRN) complex is required for numerous cellular processes that involve interactions with DNA double-strand breaks. For the majority of these processes, the MRN complex is thought to act as a unit, with each protein aiding the activity of the others. We have examined the relationship between Mre11 and Rad50 during meiosis in the basidiomycete Coprinus cinereus (Coprinopsis cinerea), investigating to what extent activities of Mre11 and Rad50 are interdependent.

Coprinus cinereus rad50 mutants reveal an essential structural role for Rad50 in axial element and synaptonemal complex formation, homolog pairing and meiotic recombination.

The Mre11/Rad50/Nbs1 (MRN) complex is required for eukaryotic DNA double-strand break (DSB) repair and meiotic recombination. We cloned the Coprinus cinereus rad50 gene and showed that it corresponds to the complementation group previously named rad12, identified mutations in 15 rad50 alleles, and mapped two of the mutations onto molecular models of Rad50 structure. We found that C.

The Coprinus cinereus adherin Rad9 functions in Mre11-dependent DNA repair, meiotic sister-chromatid cohesion, and meiotic homolog pairing.

Mitotic sister-chromatid cohesion (SCC) is known to depend in part on conserved proteins called adherins, which although necessary for SCC are not themselves localized between sister chromatids. We have examined mitotic DNA-repair and meiotic chromosome behavior in the Coprinus cinereus adherin mutant rad9-1. Genetic pathway analysis established that Rad9 functions in an Mre11-dependent pathway of DNA repair.

The art and design of genetic screens: filamentous fungi.

In the 1940s, screens for metabolic mutants of the filamentous fungus Neurospora crassa established the fundamental, one-to-one relationship between a gene and a specific protein, and also established fungi as important genetic organisms. Today, a wide range of filamentous species, which represents a billion years of evolutionary divergence, is used for experimental studies. The developmental complexity of these fungi sets them apart from unicellular yeasts, and allows the development of new screens that enable us to address biological questions that are relevant to all eukaryotes.

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.

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.

An mre11 mutant of Coprinus cinereus has defects in meiotic chromosome pairing, condensation and synapsis.

The rad11 gene of the basidiomycete Coprinus cinereus is required for the completion of meiosis and for survival after gamma irradiation. We have cloned the rad11 gene and shown that it is a homolog of MRE11, a gene required for meiosis and DNA repair in numerous organisms. The expression of C.

Insertional mutagenesis in Coprinus cinereus: use of a dominant selectable marker to generate tagged, sporulation-defective mutants.

We have constructed a dominant selectable marker, PHT1, for transformation of the basidiomycete Coprinus cinereus. PHT1 consists of a bacterial hygromycin B resistance gene fused to the promoter and terminator regions of the C. cinereus beta-tubulin gene.

Homolog pairing and meiotic progression in Coprinus cinereus.

We have used fluorescence in situ hybridization to examine homolog pairing during the synchronous meiosis of the basidiomycete Coprinus cinereus. Using spread preparations of meiotic nuclei, we confirmed previous studies that showed that at 6 h post-karyogamy essentially all meiotic nuclei are in pachytene. We found that homolog pairing occurs rapidly after karyogamy, that a 1 Mb chromosome does not associate more quickly than a 2.

Molecular analysis of pcc1, a gene that leads to A-regulated sexual morphogenesis in Coprinus cinereus.

A homokaryotic strain (5337) in our culture stock of Coprinus cinereus produced fertile fruit bodies after prolonged culture. Microscopic examination revealed that hyphae dedifferentiated from the tissues of one of the fruit bodies, as well as all basidiospore derivatives from the fruit body, exhibited pseudoclamps, whereas vegetative hyphae of 5337, from which the fruit body developed, had no clamp connections. Genetic analysis showed that the formation of pseudoclamps results from a recessive mutation in a gene designated pcc1 (pseudoclamp connection formation), which is distinct from the A and B mating type genes.

Functions of DNA repair genes during meiosis.

One of the most basic functions in any organism is DNA repair. In addition, programmed DNA "damage," in the form of DNA double-strand breaks (DSBs), is a regular part of the physiology of most organisms. There are three main types of DSB repair: homologous recombination; single-strand annealing; and nonhomologous end joining.

Isolation and characterization of rad51 orthologs from Coprinus cinereus and Lycopersicon esculentum, and phylogenetic analysis of eukaryotic recA homologs.

In eubacteria, the recA gene has long been recognized as essential for homologous recombination and DNA repair. Recent work has identified recA homologs in archaebacteria and eukaryotes, thus emphasizing the universal role this gene plays in DNA metabolism. We have isolated and characterized two new recA homologs, one from the basidiomycete Coprinus cinereus and the other from the angiosperm Lycopersicon esculentum.

Macrorestriction analysis of Caenorhabditis elegans genomic DNA.

The usefulness of genomic physical maps is greatly enhanced by linkage of the physical map with the genetic map. We describe a "macrorestriction mapping" procedure for Caenorhabditis elegans that we have applied to this endeavor. High molecular weight, genomic DNA is digested with infrequently cutting restriction enzymes and size-fractionated by pulsed field gel electrophoresis.

The rad9 gene of Coprinus cinereus encodes a proline-rich protein required for meiotic chromosome condensation and synapsis.

The rad9 gene of Coprinus cinereus is essential for the normal completion of meiosis. We examined surface-spread preparations of wild-type and rad9-1 nuclei from the meiotic stages of karyogamy through metaphase I, and we determined the primary sequence, structure, and meiotic expression of the rad9 gene. In wild-type C.

Chromosome-length polymorphism in fungi.

The examination of fungal chromosomes by pulsed-field gel electrophoresis has revealed that length polymorphism is widespread in both sexual and asexual species. This review summarizes characteristics of fungal chromosome-length polymorphism and possible mitotic and meiotic mechanisms of chromosome length change. Most fungal chromosome-length polymorphisms are currently uncharacterized with respect to content and origin.

Chromosome dynamics in rad12 mutants of Coprinus cinereus.

We have characterized the phenotypes of three rad12 mutants of the basidiomycete Coprinus cinereus, which were isolated on the basis of sensitivity to ionizing radiation. Electron microscopic studies of meiotic nuclear spreads showed that all three rad12 mutants are defective in chromosomal synapsis. For rad12-1 and rad12-4, very limited assembly of the synaptonemal complex occurs.