The synaptonemal complexes of Caenorhabditis elegans

Normal synaptonemal complexes (SCs), consisting of two lateral elements and a central element, are present in wild-type, him-4 and him-8 mutant strains in both hermaphrodites and males of Caenorhabditis elegans. Thus, the increase in rate of nondisjunction in the him mutants is not related to aberrant SC morphology. The wild-type hermaphrodite has six SCs, as determined from 3-D reconstruction analysis of serial sections from electron microscopy. Thus, n = 6 and this confirms early reports based on cytological studies with the light microscope. Only one end of the SC is attached to the nuclear envelope while the other end is free in the nucleoplasm and there is no apparent bouquet formation. Either end of the SC can attach to the nuclear envelope. The pairing behavior of the XX bivalent is normal and occurs synchronously with the autosomes. Electron dense bodies, or knobs, are associated with the SC via the central element and displace the chromatin for a distance of 200 nm. Each pachytene nucleus of the wild-type hermaphrodite has six such structures that are randomly dispersed along the bivalents such that some SCs have one or two knobs while others have none. Their function is unknown.     

Spreading synaptonemal complexes from Zea mays

Four different inversion heterozygotes of maize were examined for the occurrence of synaptic adjustment. Three substages of pachytene were identified in synaptonemal complex (SC) spreads using side-by-side comparisons of chromosome squashes with two-dimensional spreads of SCs. In SC spreads, inversion loop frequency did not change substantially from early through late pachytene for any of the four inversion heterozygotes examined. In addition, the position and size of the inversion loops remained essentially constant throughout pachytene. These results indicate that synaptic adjustment of inversion loops does not occur during pachytene in Zea mays.     

Surface spreading of synaptonemal complexes in locusts

Details are given of techniques for preparing surface spreads of locust spermatocytes for light and electron microscopy. The pachytene synaptonemal complex (SC) karyotypes of Locusta migratoria and Schistocerca gregaria are analysed and compared. Up to six different SCs can be identified in Locusta migratoria based on lengths, centromere positions, and possession of nucleolar organiser regions, but only two SCs are identifiable in Schistocerca gregaria. The total SC length is significantly greater in Schistocerca gregaria than in Locusta migratoria, and this difference is almost exactly proportional to the difference in the genomic DNA contents of the two species.     

Isolation of synaptonemal complexes from hamster spermatocytes

Nuclei from Chinese hamster testicular cells in suspension were prepared in a sucrose gradient. Following the basic procedure of Blobel and co-workers for separating a fibrous lamina-nuclear pore complex, synaptonemal complexes (SCs) from spermatocytes were isolated free of other nuclear structures, except for fibrillar tufts at the attachment plaques in which annuli were observed. All the major morphological components of the SC appeared to be intact, showing that the structure could survive the procedure and was not dispersed by the removal of DNA with DNase and solubilization of membranes and some proteins with Triton X-100. Isolated sex bodies were also well preserved, as were various structures from other cell types in the mixed cell suspension, such as spermatid manchettes, acrosomal ‘ghosts’, axonemes, etc. While no nuclear matrix was found associated with autosomal SCs, a residual material was present in the sex body, in which the X and Y axes were embedded. The results indicate the feasibility of isolating and fractionating SCs from testicular cell suspensions enriched for pachytene spermatocytes. The association between SC attachment plaques and annuli that is seen in spreads of whole nuclei persists through the isolation procedure and implies an integrated structural relationship.     

Pachytene synaptonemal complexes and meiotic achiasmatic chromosomes

Microsporocytes sometimes undergo an achiasmatic meiosis when placed into culture early in the season at a time after premeiotic S but prior to leptonema. Trillium meiocytes were examined by light and electron microscopy to analyze the frequency of cells in various stages of meiotic prophase and the occurrence of the synaptonemal complex at different times of culture. On the basis of the results, a hypothesis is proposed that suggests there is a tripartite sensitive period that occurs between S phase and leptonema. Where the cells are in this sensitive period at the time of transplantation into culture determines whether the cells do not enter meiotic prophase, enter but produce achiasmatic division figures, or enter and develop normally.This work was supported in part by grants from the National Science Foundation (GB 5173X and GB 6476) and the National Institute of Health (GM 16882)     

Electron microscopy of spread maize pachytene synaptonemal complexes

The Counce-Meyer microspreading technique for animal synaptonemal complexes (SCs) has been adapted to allow spreading of the SCs of maize pachytene microsporocytes for examination in the electron microscope (EM). The spread nuclei were well dispersed and flattened, and unstained SCs could be seen with light microscope (LM) phase optics. After PTA or ammoniacal silver staining, the SCs and kinetochores were readily recognized in the EM. Variable degrees of asynapsis, stretching of the SCs, and nonhomologous synapsis of lateral elements were noted, and cases of interlocking of lateral elements or SCs were not uncommon. Distinct lens-shaped thickenings of one or both lateral elements were observed at numerous sites along the SC in most nuclei. — The yield of well spread, complete nuclei, although not high, was sufficient to allow karyotypes to be prepared, based on relative SC lengths and arm ratios. The karyotypes agreed well with published EM and LM determinations, establishing the accuracy of the spreading technique for maize. However, considerable variation in absolute lengths of the SCs was noted. To evaluate the utility of the technique for cytogenetic investigations, two paracentric inversions, and two reciprocal translocations were spread and examined in the EM. The breakpoints estimated from measurements of spread SCs were in agreement with LM determinations.     

Two-dimensional spreads of synaptonemal complexes from solanaceous plants

By using serial sectioning and a new hypotonic bursting technique on primary microsporocytes of tomato (Lycopersicon esculentum), relatively large numbers of recombination nodules (RNs) are observed on the synaptonemal complexes forming during zygonema. In pachynema most, but not all, of these RNs are lost. If RNs represent sites of potential crossing over during zygonema and sites of actual crossing over during late pachynema, the observed temporal and spatial distribution of RNs may provide answers for some classic cytogenetic questions such as: how is at least one crossover per bivalent assured? How are crossovers localized? What is the basis for positive chiasma interference?     

Histone electrophoretic pattern in the characterization of synaptonemal complexes

The presence of a stoechiometric electrophoresis pattern of histones in carefully isolated synaptonemal complexes is reported. The use of this pattern is suggested as an internal standard of synaptonemal complex purification, in addition to the more generally used electron micrographs. This is especially useful in experiments leading to the characterization of the protein components of SCs. The use of mice of an age at which pachytenes predominate (90%) in the prophase-cell population is also advantageous to improve the final yield in synaptonemal complexes.     

Deformed lateral elements in synaptonemal complexes of Phaedranassa viridiflora

An electron microscopical study was made of an intermittent structural deformity in the lateral elements of synaptonemal complexes in pollen mother cells during zygotene of diploid and triploid forms of Phaedranassa viridiflora. The deformed regions, which extended linearly for distances up to 0.6 m in otherwise normal elements, are interpreted as hollow-centred bulges in a structure comprised of protein fibres apparently arranged in the form of a tightly wound helix. It is suggested that the bulging may arise if contraction of the lateral element is out of step with that in the chromosome, during prophase contraction at zygotene/ pachytene, as may perhaps follow under certain environmental conditions in organisms where there is some degree of genetical unbalance. There was evidence which suggested that local precocious duplication may sometimes have occurred in the chromosome axes proximal to the deformities in the lateral elements.     

A monoclonal antibody to lateral element proteins in synaptonemal complexes of Lilium longiflorum

To identify synaptonemal complex (SC) proteins in Lilium longiflorum (lily), monoclonal antibodies were generated using mice immunized with isolated pachytene nuclei. While most of the resulting monoclonal antibodies recognized nucleolar or chromatin proteins, one monoclonal antibody (anti-LE) was found that binds to lateral elements. Anti-LE bound more to lateral elements of SCs digested with DNase than to lateral elements that had not been digested with DNase. The opposite pattern of labeling was observed using monoclonal antibodies to lily chromatin and nucleolar proteins. These results indicate that anti-LE is specifically recognizing lateral element proteins and not chromatin or nucleolar proteins surrounding the lateral elements. On immunoblots, anti-LE binds to three pachytene nuclear proteins (Mr 60000, 66000 and 70000), two tetrad (early microspore) nuclear proteins (Mr 60000 and 70000), and two root tip nuclear proteins (Mr 52000 and 60000). However, anti-LE does not bind to proteins from leaf nuclei. Of these four tissues, leaf is the only one that does not have actively dividing cells. This observation suggests that at least some SC proteins are related to nuclear proteins from mitotically active cells.     

Multiple synaptonemal complexes (polycomplexes): origin, structure and function

Multiple synaptonemal complexes (polycomplexes) (PC) are similar in structure to synaptonemal complexes (SC) and are also highly conserved through evolution. They have been described in over 70 organisms throughout all life forms. The appearance of PCs are restricted to meiotic and germ-line derived tissues and are most commonly present after SC formation. However, in a number of animals and plants, both extra- and intranuclear PCs are present during premeiotic and pre-pachytene stages. The structure and biochemical composition of PCs is similar to SCs that the basic unit is tripartite, consisting of two lateral elements and a central region (in which transverse elements are located), and the dimensions of such structures are equivalent. Stacking of SC subunits, while still maintaining equivalent SC dimensions, creates a problem since the lateral elements (LE) would then be twice as thick in the PC as compared to the SC. Recently, it has been shown that the LE of the SC is actually multistranded, thus the LE of each subunit of the PC is half as thick as its counterpart in the SC.     

S. pombe linear elements: the modest cousins of synaptonemal complexes

Synaptonemal complexes (SCs) are not formed during meiotic prophase in the fission yeast, Schizosaccharomyces pombe. Instead, so-called linear elements (LinEs) are formed at the corresponding stages. LinEs are remarkable in that their number does not correspond to the number of chromosomes or bivalents and that the changes in their organisation during prophase do not evidently reflect the pairing of chromosomes. Yet, LinEs are necessary for full meiotic pairing levels and for meiotic recombination. In this review, the composition of LinEs, their evolutionary relationship to SCs and their possible functions are discussed.     

Immunocytogenetics. II. Human autoantibodies to synaptonemal complexes

Synaptonemal complex (SC) autoantibodies are spontaneously produced by patients with various autoimmune diseases. Immunofluorescence staining of pachytene cells localized the antigen to the central element or transverse filaments of the SC but not to the lateral elements. Specific antibody labeling was confined to the SC at synapsis. Cytochemical tests showed that the SC autoantigen is a basic protein possibly bound to DNA. An unusual characteristic of the SC autoantigen is its species specificity. Patients were found whose sera selectively labeled the SCs of other humans, mice, or newts. The combining of anti SC and anti-kinetochore antibodies provides a new immunocytochemical method for the analysis of SC karyotypes. The optimum conditions for preparation of pachytene cells for visualization by indirect immunofluorescence were determined. The nature and functions of the SC antigen, as well as possible applications of SC-specific autoantibodies in cytogenetics and cell biology, are discussed.     

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.     

The synaptonemal complexes of Meloidogyne: relationship of structure and evolution of parthenogenesis

The synaptonemal complexes of three amphimictic (meiotic) strains of Meloidogyne are examined in this study. M. microtyla (n = 19) has a tripartite synaptonemal complex (SC) comprised of two lateral elements and one central region with a distinct central element. The central region of the SC in both M. carolinensis (n = 18) and M. megatyla (n = 18) lack a distinct central element. The evolutionary history is different in the strains since M. microtyla has arisen by a mechanism involving an increase in chromosome number (from an ancestral stock of n = 18) while both M. carolinensis and M. megatyla have maintained the number of chromosomes of the ancestral stock. The structure of the SCs of the latter two strains are identical to the structure of the SC of the meiotic parthenogenetic M. hapla. Thus, the pachytene karyotype of M. carolinensis was reconstructed to establish the pairing pattern and identify any changes that may be related to the different morphology of the SC in an amphimictic stock. Although recombination nodules (RN) have been observed in the parthenogenetic M. hapla, none of the three amphimictic strains had any SC associated structures that resembled a RN.     

Characterization of all human male synaptonemal complexes by subtelomere multiplex-FISH

During meiotic prophase I, homologous chromosomes synapse and recombine. Both events are of vital importance for the success of meiosis. When homologous chromosomes synapse, a proteinaceous structure called synaptonemal complex (SC) appears along the pairing axis and meiotic recombination takes place. The existence of immunolabeling techniques for SC proteins (SCP1, SCP2 and SCP3) and for DNA mismatch repair proteins present in late recombination nodules (MLH1) allow analyses of both synapsis and meiotic recombination in the gametocyte I. In situ hybridization methods can be applied afterwards because chromatin is preserved during cell fixation for immunoanalysis. The combination of both methodologies allows the analysis of synapsis and the creation of recombination maps for each bivalent. In this work we apply the seven-fluorochrome subtelomere-specific multiplex FISH assay (stM-FISH) to human male meiotic cells previously labeled by immunofluorescence (SCP1, SCP3, MLH1, CENP) to assess its utility for human SC karyotyping. This FISH method consists of microdissected subtelomeric probes labeled combinatorially with seven different fluorochromes. Results prove its usefulness for the identification of all human SCs. Furthermore, by labeling subtelomeric regions this one-single-step method enables the characterization of interstitial and terminal SC fragments and SC delineation even if superposition is present in pachytene spreads.     

Alteration of meiotic chromosomal pairing and synaptonemal complexes by cycloheximide

When cells are exposed to cycloheximide during the synaptic period of meiotic prophase, the structure of the synaptonemal complex is markedly altered. The bulk of the lateral component is removed. When lily zygotene microsporocytes are subsequently transferred into a culture medium free from cycloheximide, normal synaptonemal complexes are again seen. Modification of the structure of the synaptonemal complex by treatment with cycloheximide for 4 days has little permanent effect on meiosis except at late zygonema or early pachynema. Treatment at this time produces meiocytes in which no synaptonemal complexes reform. When these cells proceed into diplotene and diakinesis they are devoid of chiasmatic chromosomes. The data suggest that the synaptonemal complex is essential if chiasmata are to be formed, and that a unique period exists when the formation can be interrupted.This work was supported by grants from the National Science Foundation (GB 5173X and GB 6476) and the National Institutes of Health (GM 16882).     

A method for the sequential study of synaptonemal complexes by light and electron microscopy

Summary A method is described for the sequential study of synaptonemal complexes by light and electron microscopy. The method is easy, permits one to determine the geometry of chromosome pairing, and should become a routine procedure in the diagnosis of human male subfertility. It should also be useful to establish the risk of recurrence of chromosome aberrations in the progeny of carriers of chromosome rearrangements.This paper is dedicated to the memory of Prof. Gerónimo Forteza Bover, the pioneer of human genetics in Spain, an excellant teacher and a good friend     

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.