The murine SCP3 gene is required for synaptonemal complex assembly, chromosome synapsis, and male fertility

During meiosis, the homologous chromosomes pair and recombine. An evolutionarily conserved protein structure, the synaptonemal complex (SC), is located along the paired meiotic chromosomes. We have studied the function of a structural component in the axial/lateral element of the SC, the synaptonemal complex protein 3 (SCP3). A null mutation in the SCP3 gene was generated, and we noted that homozygous mutant males were sterile due to massive apoptotic cell death during meiotic prophase. The SCP3-deficient male mice failed to form axial/lateral elements and SCs, and the chromosomes in the mutant spermatocytes did not synapse. While the absence of SCP3 affected the nuclear distribution of DNA repair and recombination proteins (Rad51 and RPA), as well as synaptonemal complex protein 1 (SCP1), a residual chromatin organization remained in the mutant meiotic cells.     

Formation of cytoplasmic synaptonemal-like polycomplexes at leptotene and normal synaptonemal complexes at zygotene in Ascaris suum male meiosis

A thread-like (more than 70 cm long) testis of Ascaris suum, when examined under the light and electron microscope, reveals the linear succession of meiotic stages. Beginning from, at least, late leptotene, the spermatocytes are synchronous in their development. Thus within each transverse section of the testis all the spermatocytes are in the same stage. The spermatocytes at each stage of prophase I occupies several (4 to 10) cm of the whole testis length. — At leptotene, synaptonemal-like polycomplexes of lateral and central stacked elements are formed in the cytoplasm of spermatocytes. At late leptotene, the polycomplexes are attached to the external nuclear membrane. The polycomplexes disappear at zygotene. Slightly discernable axial cores are observed in the late leptotene chromosomes. The synaptonemal complexes (SCs) are formed at the zygotene stage, their structure being characteristically tripartite. The SCs disappear from the nuclei at the diffuse stage of prophase I. In other organisms completely developed polycomplexes of stacked lateral and central elements were never found during the presynaptic period of meiosis, although single or two parallel layers of aggregated central regions of SC were found in Neottiella meiocytes at the stage prior to chromosome pairing (Westergaard and von Wettstein, 1970, 1972). — First appearance of the polycomplexes in the cytoplasm insetead of the nucleus is also a novel fact. It is concluded that the polycomplexes at leptotene are formed by a self-assembly of the SC molecular material precociously synthesized in the cytoplasm. Two hypotheses regarding possible function and the further fate for leptotene polycomplexes are discussed.     

Satellite DNA I in chromatin loops of rat pachytene chromosomes and in spermatids

Biotinylated rat satellite DNA I probe p93-50 was used to visualize the chromatin of surface-spread rat pachytene chromosomes. Fluorescein isothiocyanate (FITC)-conjugated avidin produces a beaded fluorescence pattern along the chromatin loops that insert in the centromeric region of the synaptonemal complex (SC), the paired cores of homologous chromosomes. The number of fluorescent beads ranges from zero for centromeres without satellite DNA I homologous to probe p 93-50, to several hundred for satellite-rich centromeric regions. For the chromosomes that can be identified, the relative amount of satellite DNA is chromosome specific. No satellite DNA I was detected at the non-centromeric ends of the chromosomes or interstitially. DNase-digested nuclei or isolated SCs did not have detectable amounts of satellite DNA in the centromeric regions of the chromosomes or in the residual SCs. The fate of the satellite DNA was followed during spermiogenesis. In the round spermatid the centromeric regions, which appear to be attached to the nuclear envelope, are still distinct and have converging loops of fluorescent chromatin. At later stages there are fewer but still bright fluorescent patches. Satellite DNA I is still detectable in the mature sperm head. These results demonstrate the organization of satellite DNA I in the chromatin loops at the centromeric regions, and they forecast the analysis of chromosome organization in unprecedented detail with a variety of probes in surface spreads of meiotic prophase chromosomes.     

Unusual nuclear structures in meiotic prophase of fission yeast: a cytological analysis

Earlier results from sectioned nuclei indicating that Schizosaccharomyces pombe does not develop a classical tripartite synaptonemal complex (SC) during meiotic prophase are confirmed by spreading of whole nuclei. The linear elements appearing during prophase I resemble the axial cores (SC precursors) of other organisms. The number of linear elements in haploid, diploid, and tetraploid strains is always higher than the chromosome number, implying that they are not formed continuously along the chromosomes. Time course experiments reveal that the elements appear after DNA replication and form networks and bundles. Later they separate and approximately 24 individual elements with a total length of 34 microns are observed before degradation and meiotic divisions. Parallel staining of DNA reveals changes in nuclear shape during meiotic prophase. Strains with a mei4 mutation are blocked at a late prophase stage. In serial sections we additionally observed a constant arrangement of the spindle pole body, the nucleolus, and the presumptive centromere cluster. Thus, S. pombe manages to recombine and segregate its chromosomes without SC. This might correlate with the absence of crossover interference. We propose a mechanism for chromosome pairing with initial recognition of the homologs at the centromeres and suggest functions of the linear elements in preparation of the chromosomes for meiosis I disjunction. With the spreading technique combined genetic, molecular, and cytological approaches become feasible in S. pombe. This provides an opportunity to study essential meiotic functions in the absence of SCs which may help to clarify the significance of the SC and its components for meiotic chromosome structure and function.     

Differential distribution and association of repeat DNA sequences in the lateral element of the synaptonemal complex in rat spermatocytes

The synaptonemal complex (SC) is an evolutionarily conserved structure that mediates synapsis of homologous chromosomes during meiotic prophase I. Previous studies have established that the chromatin of homologous chromosomes is organized in loops that are attached to the lateral elements (LEs) of the SC. The characterization of the genomic sequences associated with LEs of the SC represents an important step toward understanding meiotic chromosome organization and function. To isolate these genomic sequences, we performed chromatin immunoprecipitation assays in rat spermatocytes using an antibody against SYCP3, a major structural component of the LEs of the SC. Our results demonstrated the reproducible and exclusive isolation of repeat deoxyribonucleic acid (DNA) sequences, in particular long interspersed elements, short interspersed elements, long terminal direct repeats, satellite, and simple repeats. The association of these repeat sequences to the LEs of the SC was confirmed by in situ hybridization of meiotic nuclei shown by both light and electron microscopy. Signals were also detected over the chromatin surrounding SCs and in small loops protruding from the lateral elements into the SC central region. We propose that genomic repeat DNA sequences play a key role in anchoring the chromosome to the protein scaffold of the SC. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Architecture of the nuclear periphery of rat pachytene spermatocytes: distribution of nuclear envelope proteins in relation to synaptonemal complex attachment sites

The nucleus of spermatocytes provides during the first meiotic prophase an interesting model for investigating relationships of the nuclear envelope (NE) with components of the nuclear interior. During the pachytene stage, meiotic chromosomes are synapsed via synaptonemal complexes (SCs) and attached through both ends to the nuclear periphery. This association is dynamic because chromosomes move during the process of synapsis and desynapsis that takes place during meiotic prophase. The NE of spermatocytes possesses some peculiarities (e.g., lower stability than in somatic cells, expression of short meiosis-specific lamin isoforms called C2 and B3) that could be critically involved in this process. For better understanding of the association of chromosomes with the nuclear periphery, in the present study we have investigated the distribution of NE proteins in relation to SC attachment sites. A major outcome was the finding that lamin C2 is distributed in the form of discontinuous domains at the NE of spermatocytes and that SC attachment sites are embedded in these domains. Lamin C2 appears to form part of larger structures as suggested by cell fractionation experiments. According to these results, we propose that the C2-containing domains represent local reinforcements of the NE that are involved in the proper attachment of SCs.     

Atm Inactivation Results in Aberrant Telomere Clustering during Meiotic Prophase

A-T (ataxia telangiectasia) individuals frequently display gonadal atrophy, and Atm-/- mice show spermatogenic failure due to arrest at prophase of meiosis I. Chromosomal movements take place during meiotic prophase, with telomeres congregating on the nuclear envelope to transiently form a cluster during the leptotene/zygotene transition (bouquet arrangement). Since the ATM protein has been implicated in telomere metabolism of somatic cells, we have set out to investigate the effects of Atm inactivation on meiotic telomere behavior. Fluorescent in situ hybridization and synaptonemal complex (SC) immunostaining of structurally preserved spermatocytes I revealed that telomere clustering occurs aberrantly in Atm-/- mice. Numerous spermatocytes of Atm-/- mice displayed locally accumulated telomeres with stretches of SC near the clustered chromosome ends. This contrasted with spermatogenesis of normal mice, where only a few leptotene/zygotene spermatocytes I with clustered telomeres were detected. Pachytene nuclei, which were much more abundant in normal mice, displayed telomeres scattered over the nuclear periphery. It appears that the timing and occurrence of chromosome polarization is altered in Atm-/- mice. When we examined telomere-nuclear matrix interactions in spermatocytes I, a significant difference was observed in the ratio of soluble versus matrix-associated telomeric DNA sequences between meiocytes of Atm-/- and control mice. We propose that the severe disruption of spermatogenesis during early prophase I in the absence of functional Atm may be partly due to altered interactions of telomeres with the nuclear matrix and distorted meiotic telomere clustering.     

A method for preparation of synaptonemal complexes of meiotic chromosomes from basidial protoplasts of the common mushroom Agaricus bisporus (Lange) Imbach

For the first time, preparations of synaptonemal complexes (SCs) were made from meiotic chromosomes of white button mushroom (Agaricus bisporus) basidia. It is the first experience of obtaining SC preparations of filamentous fungi from isolated meiosporangium protoplasts. Previously, only yeast SC preparations were obtained following this approach. The method includes four major stages: isolation of basidium protoplasts by treatment of basidia with lytic enzymes, spreading of protoplast nuclei on a filmy support by osmotic shock, staining the preparations with silver nitrate, and examination under light and electron microscopes. The structures of spread premeiotic nuclei, axial elements of chromosomes, SCs, chromatin, and nucleoli were studied at the leptotene-diplotene stage of meiotic prophase I.     

A Method for Preparation of Synaptonemal Complexes of Meiotic Chromosomes from Basidial Protoplasts of the White Button Mushroom Agaricus bisporus (Lange) Imbach.

For the first time, preparations of synaptonemal complexes (SCs) were made from meiotic chromosomes of white button mushroom (Agaricus bisporus) basidia. It is the first experience of obtaining SC preparations of filamentous fungi from isolated meiosporangium protoplasts. Previously, only yeast SC preparations were obtained following this approach. The method includes four major stages: isolation of basidium protoplasts by treatment of basidia with lytic enzymes, spreading of protoplast nuclei on a filmy support by osmotic shock, staining the preparations with silver nitrate, and examination under light and electron microscopes. The structures of spread premeiotic nuclei, axial elements of chromosomes, SCs, chromatin, and nucleoli were studied at the leptotene–diplotene stage of meiotic prophase I.     

The relationship between nuclear envelope-derived annulate lamellae and the asymmetric division of primary spermatocytes in aphids

The structure of dividing primary spermatocytes of Amphorophora tuberculata (Aphididae, Hemiptera) as determined by electron microscopy and serial sectioning is described. The developmental stages examined extend from late prophase I to late telophase I. We looked for any asymmetric organization that could be causally linked to the differences in chromatin behaviour between the two daughter nuclei towards the end of meiosis I of this species. In late prophase I, evaginations of the nuclear envelope in the vicinity of two neigh-bouring centrosomes develop into closed cytoplasmic compartments with a dense content. The compartments open in prometaphase I and come to lie together with fragments of the nuclear envelope within the spindle area. Since nuclear pores are preserved in the membranes, intraspindle annulate lamellae have formed. These and material of presumed nuclear origin associated with them are asymmetrically distributed within the cell. Although dispersed at stages beyond prometaphase I, the material may be largely incorporated into one of the two daughter cells and thus be decisive for further development. Some annulate lamellae form a cap at the chromosome surface opposite to the neighbouring centrosomes in prometaphase I. These membranes may prevent interaction between spindle microtubules and chromosomes until a bipolar spindle forms in metaphase I. At this stage, both the banana-shaped autosomal bivalent and the X univalent occupy the equatorial plane. This is strange, because the X univalent has microtubular connections with one spindle pole and would be expected to migrate towards that pole. Possibly, the kinetochore of the X chromosome is inactive, and remains so in anaphase I, when the X univalent remains located between the two autosomal half-bivalents.M.F. Trendelenburg     

Multiple complexes in human spermatocytes

Multiple complexes develop during metaphase I in normal human spermatocytes. Usually they form two separate bodies about 1 m in diameter, composed of tripartite units and a denser matrix. The tripartite units are structurally identical to the components of the central space of synaptonemal complexes (SCs). Formation of the multiple complexes occurs by shedding of SC fragments from a few chromosomal regions at prometaphase I. The combined total length of central elements in each multiple complex is 1 to 3 m. Multiple complexes remain as cytoplasmic, perinuclear bodies during telophase I and interphase of spermatocytes II, but they were not observed during or after the second meiotic division. Although multiple complexes are initially located in the spindle, they do not show microtubular attachments and seem to be passively moved towards the periphery.     

Nuclear architecture of human pachytene spermatocytes: quantitative analysis of associations between nucleolar and XY bivalents

Summary Nucleolar association and heterochromatin coalescence have both been invoked as mechanisms involved in the origin of chromosomal associations between nucleolar bivalents themselves, as well as between these bivalents and the XY pair, during meiotic prophase in human spermatocytes. However, these mechanisms do not satisfactorily explain how associating bivalents meet each other within the nuclear space. To elucidate this problem, we have characterized different types of nucleolar-nucleolar and nucleolar-XY bivalent associations, and their frequencies, in light and electron microscope serial sections of spermatocyte nuclei. In the pachytene nucleus, nucleolar bivalent associations were found to involve only one nucleolar sphere of RNP granules connected through a fibrillar center to a chromatin mass composed of two, or more, nucleolar-bivalent short arms. Structural relationships between these elements were examined using 3D computer models of various nucleolar associations. XY and nucleolar bivalents were usually located towards the nuclear periphery associated with the inner face of the nuclear envelope. Some nucleolar bivalents, whether single or associated appeared beside or over XY chromatin. When nucleolar-bivalent short arms (BK) were found over nucleolar or over XY chromatin, their telomeres were unattached to the nuclear envelope and the corresponding synaptonemal complexes were not observed. Ninety nucleoli were found in sixty pachytene nuclei. Thirty six percent of these nucleoli were bound to associated BKs and the remaining 64% to single BKs. Over 40% of individual spermatocytes showed at least one cluster of associated BKs and about 20% presented single or multiple BKs associated with the XY pair. The frequencies of random BK associations, over the total or restricted areas of the nuclear envelope, were calculated according to a probabilistic nuclear model. A correspondence was found in comparing the observed frequencies of associated BKs with those calculated on the basis of bouquet formation. Such an analysis strongly suggests that the occurrence of associations between nucleolar bivalents may arise at random within the bouquet. Thus, the architecture of the meiocyte nucleus, particularly the organization of the bouquet, may be the primary mechanism by which nucleolar bivalents meet each other and, consequently, become associated either through common nucleolus formation or by heterochromatin coalescence.     

Synaptonemal complexes in insects

The synaptonemal complex (SC) is the key nuclear element formed in meiotic prophase I to join 2 homologous chromosomes at the pachytene bivalent. It is a highly conserved structure that is universally present in eukaryotes. The SC is presented as a tripartite protein structure, which consists of 2 lateral elements and a central region. In insects, the central region is particularly distinct and highly ordered. This made it possible to describe the fine structure of the central region and propose a model of its architecture. Chromatid DNA is arranged in chromatin loops extending radially from the SC. The loops appear to consist of a basic chromatin fiber with a diameter of 20–30 nm. In many insect species, synaptonemal polycomplexes occur in postpachytene cells. They represent one of the possible ways of SC degradation. Another process, which occurs beyond pachytene, is the formation of proteinaceous chromatid axis, the silver-stained chromatid core. Based on results in insect models, the chromatid cores have been related to the structure and formation of the SC. Research on insect models significantly contributed to understanding individual steps of the SC formation and temporal sequence of chromosome pairing. These include the formation of lateral elements of the SC, pairing initiation, interlocking of chromosomes, and synapsis of homologous chromosomes. Attention is also given to non-homologous pairing, including synaptic adjustment, correction of pairing, and pairing of sex chromosomes. In the next section, chiasmatic and achiasmatic modes of meiosis are compared with respect to the SC formation. In the chiasmatic mode, the SCs display recombination nodules that are believed to mediate the process of recombination. These nodules were discovered in insects, and indirect evidence for their role comes from insects. Two different examples of achiasmatic meiosis, occurring in the heterogametic sex of several insect orders, are given: one involves the SC formation, whereas in the other, SCs are absent. Finally, the potential of SC karyotyping for analysis of the insect genome is discussed.     

Peculiarities of chromosome organization in meiosis

Meiotic and mitotic chromosomes have a complex of differences. (1) At the early prophase I of meiosis, chromosomes acquire protein axial elements (AEs) that were absent in mitosis; in addition to somatic cohesins, AEs contain the meiosis-specific cohesins REC8, SMC1β, and STAG3. (2) At the middle prophase I, protein lateral elements (LEs) of synaptonemal complexes (SCs) are formed on the basis of AEs. The LE proteins are not conserved, but in Saccharomyces cerevisiae and Arabidopsis thaliana they contain functional domains with conserved secondary structures. Among the almost 679 thousand proteins of primitive eukaryotes that we studied by bioinformatics methods, in green and brown algae, some lower fungi, and Coelenterata, we revealed proteins or functional domains similar to SC proteins. (3) During the pachytene and diplotene stages of meiosis, chromosomes of spermatocytes and mother pollen cells acquire a general structure resembling the structure of amphibian and avian lampbrush chromosomes in miniature. Lateral chromatin loops with sizes of 90, 160, and even over 480 Kb were observed in human spermatocytes during the diplotene stage. In combination, all these observations confirm the considerable conservation of the scheme of molecular and ultrastructural organization of meiotic chromosomes in a large variety of eukaryotic organisms.     

Rapid telomere movement in meiotic prophase is promoted by NDJ1, MPS3, and CSM4 and is modulated by recombination

Haploidization of the genome in meiosis requires that chromosomes be sorted exclusively into pairs stabilized by synaptonemal complexes (SCs) and crossovers. This sorting and pairing is accompanied by active chromosome positioning in meiotic prophase in which telomeres cluster near the spindle pole to form the bouquet before dispersing around the nuclear envelope. We now describe telomere-led rapid prophase movements (RPMs) that frequently exceed 1 microm/s and persist throughout meiotic prophase. Bouquet formation and RPMs depend on NDJ1, MPS3, and a new member of this pathway, CSM4, which encodes a meiosis-specific nuclear envelope protein required specifically for telomere mobility. RPMs initiate independently of recombination but differ quantitatively in mutants that fail to complete recombination, suggesting that RPMs respond to recombination status. Together with recombination defects described for ndj1, our observations suggest that RPMs and SCs balance the disruption and stabilization of recombinational interactions, respectively, to regulate crossing over.     

Sheathed nuclear division in primary spermatocytes of Orgyia antiqua (Lepidoptera, Insecta)

The restructuring of spermatocytes during the first meiotic division is examined in the moth Orgyia antiqua (Lymantriidae, Lepidoptera) using transmission electron microscopy. Particular emphasis was placed on the behaviour of the perispindle membrane system. These membranes develop from layers of smooth endoplasmic reticulum wrapped around the prophase I nucleus and are retained until early telophase I. The original nuclear envelope is dissolved in metaphase I. Polar fenestrae in the perispindle membrane stacks are filled with numerous irregular membrane elements. The formation of new nuclear envelopes around the daughter nuclei takes place inside the perispindle membrane system. Finally, the membrane stacks rupture concomitantly with spindle elongation in late telophase I. Thus, division of primary spermatocytes in Orgyia antiqua has a surprising degree of similarity with the so-called closed mitosis. This mode of division is typical for many protozoa, algae and fungi. In the pertinent cells, the original nuclear envelope persists around the spindle area during nuclear division. In order to distinguish the closed mitosis from the situation in Orgyia antiqua spermatocytes, the term 'sheathed nuclear division' is suggested for the latter.     

Tissue distribution of two major components of synaptonemal complexes of the rat

In this paper we describe an analysis of the tissue distribution of two recently identified components of synaptonemal complexes (SCs), an Mr 125000 and an Mr 190000 protein, in the male rat by immunoblot analysis and immunocytochemical techniques. We compared the tissue distribution of these antigens with that of two earlier identified SC components, an Mr 30000 and an Mr 33000 polypeptide. For this purpose we used monoclonal antibodies (Mabs) that react exlusively with SCs in lysed spermatocytes, and that recognize the above mentioned antigens specifically in immunoblots of SC proteins or of nuclear proteins from spermatocytes: these were Mab IX9D5 (anti-190000), Mab IX5B2 (anti-125000), Mab II52F10 (anti-30000+33000), and Mab IX8G9 (anti-30000+33000). In the immunoblot experiments, we could detect the Mr 190000 and 125000 antigens exclusively in blots of SC proteins or nuclear proteins from spermatocytes; these antigens were not detectable in blots of nuclear proteins from liver, brain, spermatogonia or spermatids or in blots of proteins from mitotic chromosomes or nuclear laminae. With the anti-30000+33000 Mabs we obtained essentially the same result, except that Mab IX8G9, but not II52F10, recognizes a small amount of Mr 30000 antigen in blots of nuclear proteins from spermatids and spermatogonia. Although this might be ascribed to contamination of the isolated spermatids and spermatogonia, we cannot exclude that a small amount of Mr 30000 antigen is present in these cells. In the immunofluorescence analysis, the testis was the only tissue that reacted detectably with the above antibodies. Within the testis, spermatocytes and some early spermatids were the only cell types that contained detectable amounts of antigen. The Mr 125000 antigen was exclusively observed in nuclei of spermatocytes, from zygotene up to and including diplotene, in paired segments of SCs. The Mr 30000+33000 and 190000 antigens were present in paired as well as unpaired segments of SCs in nuclei of permatocytes, from zygotene up to and including diplotene and in the nuclei of some early spermatids in presumed remnants of SCs. We conclude that SCs consist largely of meiosisspecific proteins.by U. Scheer     

Immunofluorescent synaptonemal complex analysis in azoospermic men

The molecular cause of germ cell meiotic defects in azoospermic men is rarely known. During meiotic prophase I, a proteinaceous structure called the synaptonemal complex (SC) appears along the pairing axis of homologous chromosomes and meiotic recombination takes place. Newly-developed immunofluorescence techniques for SC proteins (SCP1 and SCP3) and for a DNA mismatch repair protein (MLH1) present in late recombination nodules allow simultaneous analysis of synapsis, and of meiotic recombination, during the first meiotic prophase in spermatocytes. This immunofluorescent SC analysis enables accurate meiotic prophase substaging and the identification of asynaptic pachytene spermatocytes. Spermatogenic defects were examined in azoospermic men using immunofluorescent SC and MLH1 analysis. Five males with obstructive azoospermia, 18 males with nonobstructive azoospermia and 11 control males with normal spermatogenesis were recruited for the study. In males with obstructive azoospermia, the fidelity of chromosome pairing (determined by the percentage of cells with gaps [discontinuities]/splits [unpaired chromosome regions] in the SCs, and nonexchange SCs [bivalents with 0 MLH1 foci]) was similar to those in normal males. The recombination frequencies (determined by the mean number of MLH1 foci per cell at the pachytene stage) were significantly reduced in obstructive azoospermia compared to that in controls. In men with nonobstructive azoospermia, a marked heterogeneity in spermatogenesis was found: 45% had a complete absence of meiotic cells; 5% had germ cells arrested at the zygotene stage of meiotic prophase; the rest had impaired fidelity of chromosome synapsis and significantly reduced recombination in pachytene. In addition, significantly more cells were in the leptotene and zygotene meiotic prophase stages in nonobstructive azoospermic patients, compared to controls. Defects in chromosome pairing and decreased recombination during meiotic prophase may have led to spermatogenesis arrest and contributed in part to this unexplained infertility.     

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.