Abnormal meiosis in bisporic mutants of white button mushroom Agaricus bisporus (Lange) Imbach

A formerly developed method of obtaining spread preparations of mushroom basidial nuclei was applied to study of meiotic prophase I in bisporic white button mushroom (Agaricus bisporus) strains. Meiotic recombination and assemblage of axial structures (axial elements and synaptonemal complexes) of chromosomes in meiotic prophase I are interrelated. It is known that the frequency of meiotic recombination is reduced in the bisporic A. bisporus variety. We showed that formation of axial structures of meiotic chromosomes in bisporic strains of this mushroom was disrupted. The phenotypes of disruptions in spread prophase nuclei are diverse. In leptotene and early zygotene, many nuclei contain abnormal, often short, and, as a rule, few chromosomal axial elements. The abnormalities in the formation of synaptonemal complexes at the zygotene-diplotene stage are of the same kind and even more pronounced. We discovered an important feature of meiosis in A. bisporus associated with fruit-body morphogenesis. Meiosis starting in basidia (meiocytes) of young closed fruit bodies is accompanied by disruption of chromatin condensation in prophase I and, probably, is arrested. After indusium breakage, the course of meiosis normalizes. Preparations with clearly observable chromosomal axial structures can be obtained only at this stage of fruit-body development.     

Abnormal meiosis in bisporic strains of white button mushroom Agaricus bisporus (Lange) imbach

A formerly developed method of microspreading of mushroom basidial nuclei was applied to study meiotic prophase I in bisporic white button mushroom (Agaricus bisporus) strains. Meiotic recombination and assemblage of axial structures (axial elements and synaptonemal complexes) of chromosomes in meiotic prophase I are interrelated. It is known that the frequency of meiotic recombination is reduced in the bisporic A. bisporus variety. We showed that formation of axial structures of meiotic chromosomes in bisporic strains of this mushroom was disrupted. The anomalous phenotypes in spread prophase nuclei are diverse. In leptotene and early zygotene, many nuclei contain abnormal, often short, and, as a rule, few chromosomal axial elements. The abnormalities in the formation of synaptonemal complexes at the zygotene-diplotene stage are of the same kind and even more pronounced. We discovered an important feature of meiosis in A. bisporus associated with fruit-body morphogenesis. Meiosis starting in basidia (meiocytes) of young closed fruit bodies is accompanied by disruption of chromatin condensation in prophase I and, probably, is arrested. After partial veil breakage, the course of meiosis normalizes. Preparations with clearly observable chromosomal axial structures can be obtained only at this stage of fruit-body development.     

Identification of the Meiotic Division of Malarial Parasites

Zygotes of Plasmodium berghei were cultured 15–25 h in vitro to yield mature infective ookinetes. Samples taken in the first 5 h of culture were examined by electron microscopy. Meiotic figures were detected in the nuclei of the zygotes. Threadlike leptotene chromatids (chromosomes) condensed from attachment plaques on the nuclear envelope; chromatid pairing followed (zygotene), with synaptonemal complexes subsequently appearing (pachytene). These complexes persisted into metaphase but dissociated when the chromatids rapidly decondensed during anaphase. At telophase of the first meiotic division the kinetochores were retracted toward two small spindle complexes, which were found at widely separated poles in the nuclear envelope. The observations are consistent with a haploid genome of 8–10 chromosomes.     

Microtubule-driven nuclear movements and linear elements as meiosis-specific characteristics of the fission yeasts Schizosaccharomyces versatilis and Schizosaccharomyces pombe

Meiotic prophase in Schizosaccharomyces pombe is characterized by striking nuclear movements and the formation of linear elements along chromosomes instead of tripartite synaptonemal complexes. We analysed the organization of nuclei and microtubules in cells of fission yeasts undergoing sexual differentiation. S. japonicus var. versatilis and S. pombe cells were studied in parallel, taking advantage of the better cytology in S. versatilis. During conjugation, microtubules were directed towards the mating projection. These microtubules seem to lead the haploid nuclei together in the zygote by interaction with the spindle pole bodies at the nuclear periphery. After karyogamy, arrays of microtubules emanating from the spindle pole body of the diploid nucleus extended to both cell poles. The same differentiated microtubule configuration was elaborated upon induction of azygotic meiosis in S. pombe. The cyclic movements of the elongated nuclei between the cell poles is reflected by a dynamic and coordinated shortening and lengthening of the two microtubule arrays. When the nucleus was at a cell end, one array was short while the other bridged the whole cell length. Experiments with inhibitors showed that microtubules are required for karyogamy and for the elongated shape and movement of nuclei during meiotic prophase. In both fission yeasts the SPBs and nucleoli are at the leading ends of the moving nuclei. Astral and cytoplasmic microtubules were also prominent during meiotic divisions and sporulation. We further show that in S. versatilis the linear elements formed during meiotic prophase are similar to those in S. pombe. Tripartite synaptonemal complexes were never detected. Taken together, these findings suggest that S. pombe and S. versatilis share basic characteristics in the organization of microtubules and the structure and behaviour of nuclei during their meiotic cell cycle. The prominent differentiations of microtubules and nuclei may be involved in the pairing, recombination, and segregation of meiotic chromosomes.     

Meiotic recombination and synaptonemal complexes in Saccharomyces cerevisiae.

Summary The course of meiotic recombination, gene conversion and crossing-over, was investigated in Saccharomyces cerevisiae. Gene conversion was used as the selected event by removing cells from a medium inducing and promoting meiosis to a vegetative growth medium selective for convertants. Gene conversion started to increase at the same time as DNA synthesis, and nuclei entered a phase where the chromatin appeared as thread-like structures. Crossing-over of linked and unlinked markers also started early but remained at a low level until synaptonemal complexes were formed. However, gene conversion and a limited amount of crossing-over could be completed without synaptonemal complexes. It was concluded that meiotic recombination in yeast can occur as early as during DNA synthesis and does not require the function of synaptonemal complexes. Moreover, the low incidence of crossing-over early in meiosis is attributed to a low frequency of strand isomerization.     

Meiotic homologue alignment and its quality surveillance are controlled by mouse HORMAD1

Meiotic crossover formation between homologous chromosomes (homologues) entails DNA double-strand break (DSB) formation, homology search using DSB ends, and synaptonemal-complex formation coupled with DSB repair. Meiotic progression must be prevented until DSB repair and homologue alignment are completed, to avoid the formation of aneuploid gametes. Here we show that mouse HORMAD1 ensures that sufficient numbers of processed DSBs are available for successful homology search. HORMAD1 is needed for normal synaptonemal-complex formation and for the efficient recruitment of ATR checkpoint kinase activity to unsynapsed chromatin. The latter phenomenon was proposed to be important in meiotic prophase checkpoints in both sexes. Consistent with this hypothesis, HORMAD1 is essential for the elimination of synaptonemal-complex-defective oocytes. Synaptonemal-complex formation results in HORMAD1 depletion from chromosome axes. Thus, we propose that the synaptonemal complex and HORMAD1 are key components of a negative feedback loop that coordinates meiotic progression with homologue alignment: HORMAD1 promotes homologue alignment and synaptonemal-complex formation, and synaptonemal complexes downregulate HORMAD1 function, thereby permitting progression past meiotic prophase checkpoints.     

Coelomomyces psorophorae vartasmaniensis Couch+Laird (1988) (Coelomomycetaeceae: Blastocladiales), a fungal pathogen of the mosquitoAedes australis

The germination of sporangia inCoelomomyces psorophorae vartasmaniensis (C. p. tas.) is uncoordinated and thus there is a mixture of developmental stages in any given population. Continuous urografin gradients separated out the critical stages of germinating sporangia giving four bands, each band representing a consecutive stage of germination. These stages were investigated for changes in the sporangial wall using Transmission Electron Microscopy (TEM). The sporangia have a typical two-layered wall, an electron dense outer layer which can be divided into three distinct sub-layers D1, D2, and D3 and an inner electron transparent secondary wall. Stage 3 sporangia have an intact D1 layer on their outer wall. In the subsequent stages (4 & 4b) there is a progressive breakdown of this layer.     

Spermatogenesis in Bombyx mori. II. The ultrastructure of synapsed bivalents

In Bombyx mori the male is the homogametic sex, crossing over occurs only in males, and chiasmata are observed in spermatocytes, but not in oocyte nuclei. If the assembly of synaptonemal complexes is an essential prerequisite for genetic crossing over and chiasmata formation, then the nuclei of Bombyx spermatocytes should contain synaptonemal complexes. Synaptonemal complexes were found in spermatocytes from young four instar larvae. The structure of meiotic bivalents is described using micrographs taken with 100 and 1000 KV electron microscopes. These data together with that from the literature are used to construct a three-dimensional model of the synaptonemal complex and to suggest its method of origin and its function during crossing over.     

Synaptonemal polycomplexes in spermatocytes of the gooseneck barnacle,Pollicipes polymerus Sowerby (Crustacea: Cirripedia)

Polycomplexes are described for the first time in spermatocytes of a cirripede crustacean, Pollicipes polymerus Sowerby. Synaptonemal complexes of regular tripartite construction are seen from zygotene to mid-pachytene. Although some of the synaptonemal complexes are disrupted at late pachytene and may degenerate at this stage, some persist and by diplotene may form polycomplexes by the bending and self-fusion of their lateral elements. These polycomplexes are still encompassed by chromosomes and consist of four dense plates and intercalated central elements and transverse fibers. Other polycomplexes with five or six dense plates, all of which are considerably wider than lateral elements of mid-pachytene synaptonemal complexes, are also seen in diplotene nuclei. These may be attached to a chromosome at only one end or may be in the nucleoplasm, free of chromosomal involvement except for fine fibrous connectives. No polycomplexes are seen in meiotic cells after diplotene and their fate is unknown. It is suggested that poly-complexes serve as sequestra for synaptonemal material which could prevent normal chromosomal disjunction.     

COORDINATNE NUCLEAR AND CHLOROPLAST DIVISION IN UNILOCULAR SPORANGIA OF LAMINARIA ANGUSTATA (LAMINARIALES,PHAEOPHYCEAE)1

Changes in the number of nuclei and chloroplasts were examined during the process of unispore formation in unilocular sporangia of Laminaria angustata. Just before meiosis, eight chloroplasts were always present in unilocular sporangial mother cells. The number of chloroplasts remained constant through meiosis. After the resulting four nuclei divided again (third nuclear division), a close association between a nucleus and a chloroplast developed among each of the eight nuclei and eight chloroplasts. The eight chloroplasts divided ahost synchronously before the synchronous division of the eight nuclei. Following the 16 nucleate stage with 16 chloroplasts and the final 32 nucleate stage with 32 chloroplasts, 32 unispores, each with a nucleus and a chloroplast, were fomd in unilocular sporangza of L. angustata. Immunofluorescence microscopy using an anti-centrin antibody showed that two anticentrin-stained structures (as future mitotic poles) occurred adjacent to each of the premitotic four nuclei, and each spot was located near a chloroplast. Therefore, after the third division, each of the eight nuclei established close contact with a chloroplast presumably mediated by the centrosomes.     

Sequential analysis of synaptonemal complexes in the repopulating spermatocytes of Rattus norvegicus after restricting the germ cell population to spermatogonia by gossypol treatment

Synaptonemal complexes of the repopulating spermatocytes of male rats were analyzed day by day using silver-stained surface spread nuclei between 8 and 25 days after restricting the germ cell population to spermatogonia by treatment of gossypol acetic acid at 30 mg/kg body weight/day for 70 days. The method allowed sequential analysis of male meiotic prophase on successive days after the last day of treatment. The leptotene cells appeared on day 11 and were characterized by a network of lateral elements and large nucleolar bodies in a diffuse mass. On day 13 the unpaired lateral elements and short stretches of synaptonemal complexes characteristic for zygotene could be seen. Pachytene nuclei showing 20 autosomal synaptonemal complexes and XY axes appeared on day 15. The diplotene cells were defined on day 22 by the loss of a complete synaptonemal complex set and by the appearance of disjoined lateral elements and persistent segments of synaptonemal complexes.     

Germination of the resting sporangia ofPhysoderma maydis,the causal agent ofPhysoderma disease of maize

Summary The structural and developmental characteristics of the resting sporangium in uniflagellate phycomycetes, together with the type of zoospore, are of high taxonomic value. Among these fungi, however, only a few electron microscopic investigations have been published on this topic, mainly due to technical problems. In the present study ofPhysoderma maydis (Blastocladiales) these problems were overcome as the resting sporangia in this species are formed synchronously, in large numbers, the germination is readily induced and the impermeability of the resting sporangium wall can be circumvented by shaking the prefixed sporangia with glass beads.The germination of the resting sporangia ofP. maydis is described by correlative light and electron microscopic studies and discussed in relation to related investigations on sporogenesis: The germination process starts by a breakdown of large electron-dense accretions found in the resting stage. Simultaneously, the peripheral location of the lipid bodies is lost. The large operculum is pushed open by a protrusion of the inner sporangial wall; an additional wall layer is formed during this process. Synaptonemal complexes are found in the nuclei at this stage, as are nuclear division figures which suggests anEuallomyces type of life cycle for this fungus. Cleavage vesicles, formed from dictyosomes or endoplasmic reticulum, ultimately separate the sporangial content into meiospores. The sequential assembly of organelles into the side body complex is described. Sequestering of the ribosomes into a nuclear cap is interpreted as taking place immediately prior to zoospore discharge.     

Loss of HR6B ubiquitin-conjugating activity results in damaged synaptonemal complex structure and increased crossing-over frequency during the male meiotic prophase

The ubiquitin-conjugating enzymes HR6A and HR6B are the two mammalian homologs of Saccharomyces cerevisiae RAD6. In yeast, RAD6 plays an important role in postreplication DNA repair and in sporulation. HR6B knockout mice are viable, but spermatogenesis is markedly affected during postmeiotic steps, leading to male infertility. In the present study, increased apoptosis of HR6B knockout primary spermatocytes was detected during the first wave of spermatogenesis, indicating that HR6B performs a primary role during the meiotic prophase. Detailed analysis of HR6B knockout pachytene nuclei showed major changes in the synaptonemal complexes. These complexes were found to be longer. In addition, we often found depletion of synaptonemal complex proteins from near telomeric regions in the HR6B knockout pachytene nuclei. Finally, we detected an increased number of foci containing the mismatch DNA repair protein MLH1 in these nuclei, reflecting a remarkable and consistent increase (20 to 25%) in crossing-over frequency. The present findings reveal a specific requirement for the ubiquitin-conjugating activity of HR6B in relation to dynamic aspects of the synaptonemal complex and meiotic recombination in spermatocytes.     

Meiosis in the aquatic fungusCatenaria allomycis

Summary Catenaria allomycis Couch (Blastocladiales) is an endobiotic fungal parasite primarily of species of the genusAllomyces. The life cycle ofC. allomycis contains both sexual and asexual phases. Synaptonemal complexes have been found in young developing resistant sporangia (RS) suggesting that meiosis occurs within the thick walled RS prior to syngamy. Ultrastructural evidence suggests that meiosis proceeds through pachytene in the developing RS and is arrested in diplotene of prophase I until the sporangia are induced to germinate at which time the meiotic process is completed. Quantitative nuclear counts in developing RS support the ultrastructural observations. Meiotic nuclei are characterized by polar fenestrae in the nuclear envelope and intranuclear plaque-like microtubule organizing centers (MTOC).Portion of a Ph.D. dissertation submitted by the senior author to the Graduate School, University of Georgia.     

Presence of abnormal synaptonemal complexes in heterothallic species of Neurospora

Synaptonemal complex abnormalities are frequent in reconstructed meiotic prophase nuclei of Neurospora crassa and Neurospora intermedia. Three kinds of synaptonemal complex anomalies were seen: lateral component splits, lateral component junctions, and multiple complexes. The anomalies apparently are formed during or after the pairing process, as they were not seen in the largely unpaired early zygotene chromosomes. Their presence at all the other substages from mid-zygotene to late pachytene indicates that they are not eliminated before the synaptonemal complex decomposes at diplotene. Abnormal synaptonemal complexes were seen in all 19 crosses of N. crassa and N. intermedia that were examined, including matings between standard laboratory strains, inversions, Spore killers, and strains collected from nature. The frequency of affected nuclei and degree of abnormality within a nucleus varied in different matings. No abnormalities were present in the homothallic species Neurospora africana and Neurospora terricola. Structural chromosome aberrations, introgression, and heterozygosity have been eliminated as causes for pairing disorder. The abnormal synaptonemal complexes seemingly do not interfere with normal ascus development and ascospore formation. The affected nuclei are not aborted during meiotic prophase, nor are they eliminated by abortion of mature asci. The abnormal meiocytes do not lead to aneuploidy, as judged by the low frequency of white ascospores in crosses between wild type strains that have many abnormalities. Thus, the abnormal synatonemal complexes do not appear to prevent chiasma formation between homologues.     

Morphology of mitochondrial nucleoids, mitochondria, and nuclei during meiosis and sporulation of the yeast Saccharomycodes ludwigii

The morphology of mitochondrial nucleoids (mt-nucleoids), mitochondria, and nuclei was investigated during meiosis and sporulation of the diploid cells of the ascosporogenic yeast Saccharomycodes ludwigii. The mt-nucleoids appeared as discrete dots uniformly distributed in stationary-phase cells as revealed by 4',6-diamidino-2-phenylindole (DAPI) staining. Throughout first and second meiotic divisions, the mt-nucleoids moved to be located close to the dividing nuclei with the appearance of dots. On the other hand, mitochondria, which had tubular or fragmented forms in stationary-phase cells, increasingly fused with each other to form elongated mitochondria during meiotic prophase as revealed by 3,3' -dihexyloxacarbocyanine iodide [DiOC(6)(3)] staining. Mitochondria assembled to be located close to dividing nuclei during first and second meiotic divisions, and were finally incorporated into spores. During the first meiotic division, nuclear division occurred in any direction parallel, diagonally, or perpendicular to the longitudinal axis of the cell. In contrast, the second meiotic division was exclusively parallel to the longitudinal axis of the cell. The behavior of dividing nuclei explains the formation of a pair of spores with opposite mating types at both ends of cells. In the course of this study, it was also found that ledges between two spores were specifically stained with DiOC(6)(3).     

Ultrastructure of Wall Swellings of Germinating Sporangia of Phytophthora palmivora (Butl.) Butl.

Germ tubes of directly germinating sporangia of P. palmivoraincubated in yeast extract solution at 30 ?C usually developedinto prominent swellings from which hyphae later emerged. Thegerm tubes arose as an extension of a new germination wall formedinternal to the sporangial wall prior to germination. The germtube swellings contained typical hyphal organelles. The germtube swelling possessed a thicker wall than both hyphae growingout of it and germ tubes that did not form swellings.     

Three-Dimensional Ultrastructural Karyotype Analysis from the Meiotic Parthenogenetic Nematode Heterodera betulae

Meiotic chromosome structure and function are described in the plant-parasitic nematode Heterodera betulae. Twelve synaptonemal complexes (SCs) were reconstructed from pachytene nuclei; therefore, n=12 is predicted for this species. Morphologically distinct sex chromosomes were not observed. Only one end of the SC is attached to the nuclear envelope, and there is no bouquet arrangement at pachytene. The structure of the SC in this meiotic parthenogenetic nematode was different than in other nematodes that reproduce via amphimixis; a striated central element with transverse filaments was not observed. Multiple SCs, or polycomplexes, were present in the nucleus. Recombination nodules were not observed. The centrioles were comprised of nine doublet microtubules connected by a ring, which is a distinct modification from the typical nine triplet microtubules without any interconnecting structure.