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The XX/XY system is the rule among mammals. However, many exceptions from this general pattern have been discovered since the last decades. One of these non-conventional sex chromosome mechanisms is the multiple sex chromosome system, which is evolutionary fixed among many bat species of the family Phyllostomidae, and has arisen by a translocation between one original gonosome (X or Y chromosome), and an autosome, giving rise to a "neo-XY body." The aim of this work is to study the synaptic behavior and the chromatin remodeling of multiple sex chromosomes in different species of phyllostomid bats using electron microscopy and molecular markers. Testicular tissues from adult males of the species Artibeus lituratus, Artibeus planirostris, Uroderma bilobatum, and Vampyrodes caraccioli from the eastern Amazonia were analyzed by optical/electron microscopy and immunofluorescence of meiotic proteins involved in synapsis (SYCP3 and SYCE3), sister-chromatid cohesion (SMC3), and chromatin silencing (BRCA1, γ-H2AX, and RNApol 2). The presence of asynaptic axes-labeled by BRCA1 and γ-H2AX-at meiotic prophase in testes that have a normal development of spermatogenesis, suggests that the basic mechanism that arrests spreading of transcriptional silencing (meiotic sex chromosome inactivation (MSCI)) to the autosomal segments may be per se the formation of a functional synaptonemal complex between homologous or non-homologous regions, and thus, this SC barrier might be probably related to the preservation of fertility in these systems.

Summary The basic molecular mechanisms by which chromosomal rearrangements in heterozygous state produce spermatogenic disturbances are poorly understood. Testicular biopsies from five patients ‐ one carrier of a Robertsonian translocation rob t(13;14), two carriers of two different Y-autosome translocations, a t(Y;6) and a t(Y;11), one carrier of a reciprocal translocation t(3;13) and one carrier of a heterochromatin duplication in chromosome 9 ‐ were processed for histopathological analysis, electron microscopy and fluorescent immunolocalization of meiotic proteins. In all the patients, the asynaptic regions during pachytene are labelled by BRCA1 and retained RAD51 foci. The variant histone c-H2AX is located on the chromatin domains of the asynaptic regions and the XY body. In contrast, these meiotic proteins are absent in those chromosomal segments that are non-homologously synapsed. The present observations on five new cases and a review of recent studies show that the common features shared by all these cases are the abnormal location of some meiotic proteins and the presence of transcriptionally silenced chromatin domains on asynaptic regions. The frequent association of these silenced regions with the XY body and the rescue of spermatocyte viability through non-homologous synapsis are also shared by all these carriers. A passive, random mechanism of clustering of asynaptic regions with the XY body is suggested.

BACKGROUND: The mechanisms of meiotic arrest in human spermatogenesis are poorly known. METHODS AND RESULTS: A testicular biopsy from an azoospermic male showed complete spermatogenesis arrest at the spermatocyte stage, asynapsis, lack of formation of the XY body, partial reversion to a mitotic-like division and cell degeneration both at the prophase and at the abnormal cell divisions. Synaptonemal complex analysis showed minor segments of synapsis and mainly single axes. Fluorescent immunolocalization of meiotic proteins showed normal SYCP3, scarcity of SYCP1, null MLH1 foci, about 10 patches of -H2AX, abnormal presence of BRCA1 among autosomal axes, absence of RAD51 in early and advanced spermatocytes and permanence of -H2AX labelling up to the abnormal spermatocyte divisions that are the most advanced stage reached. There are at least six dominions of evenly packed chromatin resembling that of the normal XY body, but no true XY body. CONCLUSIONS: The protein phenotype and the fine structure of the nuclei are compatible with a deficiency of the processing of double-strand DNA breaks in the zygotene-like spermatocytes, but the features of this defect do not agree with Spo11, Sycp1, Atm and Dmc1 null mutations, which give absence of XY body, synapsis disturbances and spermatocyte apoptosis in mice. Fil: Sciurano, Roberta Beatriz. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Biología Celular e Histología. Centro de Investigación en Reproducción; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina Fil: Rahn, Irene Monica. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Biología Celular e Histología. Centro de Investigación en Reproducción; Argentina Fil: Pigozzi, Maria Ines. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Biología Celular e Histología. Centro de Investigación en Reproducción; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina Fil: Brugo Olmedo, S.. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Biología Celular e Histología. Centro de Investigación en Reproducción; Argentina Fil: Solari, Alberto Juan. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Biología Celular e Histología. Centro de Investigación en Reproducción; Argentina

Three xenarthrans species Chaetophractus villosus, Chaetophractus vellerosus, and Zaedyus pichiy have been used for the analysis of the structure, behavior, and immunochemical features of the XY body during pachytene. In all these species, the sex chromosomes form an XY body easily identifiable in thin sections by the special and regular packing of the chromatin fibers of the internal region of the XY body ("differential" regions) and those of the peripheral region (synaptic region). Spermatocyte spreads show a complete synapsis between the X- and the Y-axis, which lasts up to the end of pachytene. From the early pachytene substages to the late ones, the X-axis develops prominent branches, which in late pachytene span the synaptic region. Synapsis is regular as shown by SYCP1 labeling. Axial development is followed by SYCP3 labeling and in the asynaptic region of the X-axis by BRCA1. Gamma-H2AX labels exclusively the differential (asynaptic) region of the X chromosome. A single focus is labeled by MLH1 in the synaptic region. The location of this MLH1 focus spans from 0.3 to 1.6 μm from the telomere in the analyzed xenarthrans, covering approximately half of the Y-axis length. It is concluded that xenarthrans, as basal placental mammals, harbor the largest pseudoautosomal regions of presently analyzed mammals, and shows the typical features of meiotic sex chromosome inactivation (MSCI).

The basic molecular mechanisms by which chromosomal rearrangements in heterozygous state produce spermatogenic disturbances are poorly understood. Testicular biopsies from five patients - one carrier of a Robertsonian translocation rob t(13;14), two carriers of two different Y-autosome translocations, a t(Y;6) and a t(Y;11), one carrier of a reciprocal translocation t(3;13) and one carrier of a heterochromatin duplication in chromosome 9 - were processed for histopathological analysis, electron microscopy and fluorescent immunolocalization of meiotic proteins. In all the patients, the asynaptic regions during pachytene are labelled by BRCA1 and retained RAD51 foci. The variant histone γ-H2AX is located on the chromatin domains of the asynaptic regions and the XY body. In contrast, these meiotic proteins are absent in those chromosomal segments that are non-homologously synapsed. The present observations on five new cases and a review of recent studies show that the common features shared by all these cases are the abnormal location of some meiotic proteins and the presence of transcriptionally silenced chromatin domains on asynaptic regions. The frequent association of these silenced regions with the XY body and the rescue of spermatocyte viability through non-homologous synapsis are also shared by all these carriers. A passive, random mechanism of clustering of asynaptic regions with the XY body is suggested.

background: Klinefelter syndrome is the most frequent chromosome abnormality in human males. This paper aims to investigate the ploidy of meiotic and pre-meiotic germ cells found in spermatogenic foci, and furthermore, the sex chromosome constitution of Sertoli cells which surround these germ cells in non-mosaic Klinefelter patients.Klinefelter syndrome is the most frequent chromosome abnormality in human males. This paper aims to investigate the ploidy of meiotic and pre-meiotic germ cells found in spermatogenic foci, and furthermore, the sex chromosome constitution of Sertoli cells which surround these germ cells in non-mosaic Klinefelter patients. methods and results: A survey of 11 adult patients diagnosed with classical, non-mosaic Klinefelter syndrome who underwent testicular biopsies, showed that six of them had spermatogenesis foci. The topographical study of the biopsies showed that tubuli with germ cells are a minor fraction (8–24%) of all tubuli, although the overwhelming majority is devoid of germ cells. Using fluorescenceA survey of 11 adult patients diagnosed with classical, non-mosaic Klinefelter syndrome who underwent testicular biopsies, showed that six of them had spermatogenesis foci. The topographical study of the biopsies showed that tubuli with germ cells are a minor fraction (8–24%) of all tubuli, although the overwhelming majority is devoid of germ cells. Using fluorescence in situ hybridization (FISH) with probes for the X-centromere and immunolocalization of meiotic proteins, the present work shows that all the 92 meiotic spermatocytes analyzed with FISH were euploid, 46,XY, and thus can form normal, haploid gametes. On the other hand, Sertoli cells show two marks for the X chromosome, meaning that they are 47,XXY.hybridization (FISH) with probes for the X-centromere and immunolocalization of meiotic proteins, the present work shows that all the 92 meiotic spermatocytes analyzed with FISH were euploid, 46,XY, and thus can form normal, haploid gametes. On the other hand, Sertoli cells show two marks for the X chromosome, meaning that they are 47,XXY. conclusions: These results provide a rationale for the high rate of success in the testicular sperm extraction plus ICSI procedures when applied to Klinefelter patients. It is also in agreement with previous studies in the XXY-mouse model. These spermatogenic foci most probably originate from clones of spermatogonia that have randomly lost one of the X chromosomes, probably during periods of life when high spermatogonial mitotic activity occurs.These results provide a rationale for the high rate of success in the testicular sperm extraction plus ICSI procedures when applied to Klinefelter patients. It is also in agreement with previous studies in the XXY-mouse model. These spermatogenic foci most probably originate from clones of spermatogonia that have randomly lost one of the X chromosomes, probably during periods of life when high spermatogonial mitotic activity occurs. Fil: Sciurano, Roberta Beatriz. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Biología Celular e Histología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina Fil: Luna Hisano, C.V.. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Biología Celular e Histología; Argentina Fil: Rahn, Irene Monica. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; Argentina Fil: Olmedo, S. Brugo. No especifíca; Fil: Rey Valzacchi, Gaston Javier. No especifíca; Fil: Coco, Roberto. Fecunditas Instituto de Medicina Reproductiva; Argentina Fil: Solari, Alberto Juan. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Biología Celular e Histología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina

Background The close apposition of multivalents with the XY body has been repeatedly described in heterozygous carriers of chromosomal rearrangements. Because in many of these carriers spermatogenesis is deeply disturbed at the spermatocyte level, the association of autosomal chromatin with the XY body may impair the spermatocyte life. Methods Testicular biopsies from three men carriers of three different chromosomal rearrangements have been analysed by electron microscopy (EM) and immunolocalization of meiotic proteins. Results There is an ordered transition from isolated multivalents at early pachytene to XY body association in late pachytene, as shown in a carrier of a rob t(13;14) translocation by EM and in a reciprocal translocation t(9;14) carrier by immunofluorescence. The non-synapsed ends of the quadrivalent show BRCA1 located on the axes and the variant histone gamma-H2AX located on the chromatin. The area covered by gamma-H2AX increases with the association of the asynaptic ends with the XY body in the t(9;14) carrier, and the area covered with gamma-H2AX in the t(Y;15) carrier is larger than that of the XY body of controls. Conclusions The affinity between the inactive XY body and asynaptic regions of multivalents is given a material basis, and transcriptional inactivation is probably shared by these two chromatin types.

As with most platyrrhines, the systematics of Ateles is under discussion. In order to help clarify its systematic, we employed chromosomic and molecular characters to analyze the phylogenetic relationship among some species of the genus Ateles. Chromosomic studies were conducted on 14 atelid specimens: eight Ateles from A. paniscus, A. chamek, A. belzebuth and A. geoffroyi, and six Alouatta caraya. Ateles paniscus showed 2N=32, whereas A. chamek, A. belzebuth and A. geoffroyi presented 2N=34, XX/XY (with a submetacentric X and a variable Y) corroborated by male meiosis. Nucleotide sequence variation at the mitochondrial cytochrome c oxidase subunit II gene (COII) was analyzed in ten New World monkey specimens. Parsimony trees showed consistent phylogenetic relationships using both chromosomic forms and mitochondrial COII gene sequences as characters. Particularly, chromosomic phylogenies showed A. hybridus as a divergent taxon from the remaining group, whereas A. chamek, A. belzebuth and A. marginatus form an unresolved clade with A. geoffroyi as sister group.

The meiotic cytology and fine structure of the sex multiples in males from two species of the genus Alouatta are presented and compared with descriptions from other species of this genus. As shown in pachytene by synaptonemal complex analysis and in metaphase I by spreading, there is a quadrivalent in male meiosis in A. caraya, which is formed by an X(1)X(2)Y(1)Y(2) complex, while in A. palliata there is a trivalent formed by an X(1)X(2)Y(1) complex. Chromosome painting with human probes shows that A. caraya sex multiples share the same components as those of A. seniculus sara and A. seniculus arctoidea. However, as shown here for A. palliata and by others in A. fusca, there are differences among the multiples of some species. It is shown that in this genus there are several varieties of sex multiples that share some features, and that the origin of these multiples is most probably a primitive development in the genus Alouatta.

Most primates studied have the usual XX/XY sex-chromosome system. However, exceptions to this rule among howler monkeys have been suggested by several authors. Recently a quadrivalent was discovered in male meiosis of Alouatta caraya and it was established that this species has an X1X2Y1Y2 sex chromosome system. On that basis, a cytogenetic analysis of 25 males of this species is described. showing the corrected karyotype of this species. Each chromosome involved in the particular sex-chromosome system of this species is identified on the basis of mitotic chromosome measurements. G and C-banding patterns as well as on the relative measurements of synaptonemal complexes. It is now established that A. caraya has a karyotype with 2n = 52 in both sexes, and that the male one shows a single autosome 7 (X2) besides the X (X1) and the two products of the reciprocal translocation between the second autosome #7 and the Y chromosome (Y1 and Y2), while females show a homomorphic pair #7 (X2) and a pair of X1. The evolutionary implications of the exceptional primate species having composite sex-chromosome systems are discussed. Fil:Mudry, M.D. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Gorostiaga, M. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.

A multiple sex chromosome system was found in three unrelated individuals of the primate Alouatta caraya. This mechanism is originated by a translocation between the Y chromosome and one of the autosomes (A7). Mitotic karyotypes show two small, acrocentric chromosomes (AY and YA), which are the translocation products. In metaphase I of male meiosis, there is a very long chain quadrivalent in which the order of the element is: X-YA-A7-AY. Segregation in the quadrivalent is alternate and gives balanced products. Synaptonemal complex karyotypes at pachytene show the structure of the quadrivalent made by the four axes. There is a slight difference in the relative length of AY and YA and the kinetochore of A7 aligns with that of AY. The synaptic pattern and changes in the quadrivalent during pachytene are described. Thin sections of the quadrivalent body show that the chromatin packing in the sex chromosome region is different from that of the autosomal region. This X1X2Y1Y2/X1X1XX2 sex chromosome system may be extended among other members of the genus Alouatta.

The meiotic behavior of the XY pair of the chinchilla (C. laniger), has been studied with C-banding and with the microspreading procedure for synaptonemal complex (SC) karyotyping. The large X chromosome of the chinchilla has a paracentromeric and two subterminal (stl and st2) C+ bands, while the minute Y chromosome has a C+ long arm. At metaphase-I the X chromosome is associated end-to-end with the short arm of the Y chromosome. The appearance of the X axis in microspread spermatocytes is delayed up to early pachytene. The X axis, as well as the Y axis appear as thick, separated threads. The thick X and Y axes are then co-aligned without forming a SC. The Y axis becomes thinner and then an asymmetrical SC is formed between the axes. This SC becomes symmetrical by a later thinning of the corresponding part of the X axis. During mid-pachytene an additional SC is formed at the other end of the XY pair in many cells. During late pachytene the SCs become resolved and finally the four termini are separated from each other although they tend to be located in the same region of the nucleus. These observations suggest that the SCs in the XY pair of the chinchilla are formed between non-homologous regions. The presence of non-homologous synapsis suggests that the XY pair ofC. laniger is joined by an achiasmatic mechanism, different from the chiasmatic joining usually found in eutherian mammals. The presence of subterminal C+ bands in the X chromosome may be a hindrance for the formation of a SC during early pachytene. It is concluded that the XY pair of the chinchilla shows an intermediate behavior when compared to that of the sand rat on one side, and that of most mammals having a euchromatic X chromosome, on the other side.

Chicken oocytes at pachytene were processed with the microspreading technique (Moses, 1977), and their synaptonemal complex (SC) complements were analyzed by electron microscopy. Ellipsoidal nodules, 140 X 120 nm in diameter, were associated with the central space of synaptonemal complexes. The average number of nodules per pachytene oocyte was 57.5. The number of nodules per bivalent showed a clear linear relationship with SC length, except for the microchromosomes, which showed a single obligatory nodule. The distribution of nodules along the 10 longest SCs was nonrandom, with low frequencies in the vicinity of kinetochores and high frequencies near the telomeres. The microchromosomes showed a single nodule whose average location was 1.21 micron from the kinetochore. In the ZW pair there was a single nodule whose average location was 0.31 micron from the paired telomeres and not more than 0.65 micron from them. The total number of nodules per cell and the number of nodules in each of the five major bivalents showed good agreement with the total number of chiasmata and the number of chiasmata of the major bivalents of roosters. Thus, these nodules share the characteristics of recombination nodules described in other organisms. The single, obligatory, strictly localized recombination nodule found in the pairing end of the ZW pair strongly suggests that recombination between the Z and W chromosomes in the female chicken is a regular process that may be similar to the obligatory recombination between the pairing ends of the human X and Y chromosomes that was recently described in studies using DNA probes.