Cytogenetic observations on XX/XY chimaeras and a reassessement of the evidence for germ cell chimaerism in heterosexual twin cattle and marmosets.

Testicular preparations were obtained from 7 bulls, twins of freemartins, and 1 male marmoset, all proved XX/XY chimaeras. X and Y sex chromosomes were confidently identified in nearly all the 87 spermatogonia at mitotic metaphase and 1052 primary spermatocytes at diakinesis-metaphase examined: no cell was identified as containing two X chromosomes. The germ cell chimaerism previously reported in these species is therefore not confirmed. Cultures grown from presumptive somatic these species is therefore not confirmed. Cultures grown from presumptive somatic cells in the testes of two of the bulls yielded 248 identifiable mitotic spreads, all XY-type; cultures from the gonads of their freemartin twins yielded 442 mitotic spreads, all XX-type. Direct preparations from one freemartin gonad, however, yielded 3 XY mitotic spreads out of 18 examined. The conflicting evidence concerning germ cell chimaerism in cattle and marmosets is discussed, particularly in relation to reports of XX/XY bulls that have sired a great excess of daughters. The possibility that XX germ cells contributed to the functional spermatozoa of these bulls is not favoured by present information, but is not excluded.     

黑斑蛙的减数分裂研究

本文研究了黑斑蛙的减数分裂,发现其性染色体所形成的性二价体主要呈末端与末端联接,浓缩期占79.6％,中期Ⅰ占75％,这进一步证明黑斑蛙确实存在XY型性别决定机制,这种XY型性染色体虽形态相同,但已发生了质的分化,可能是同型异质。黑斑蛙的性染色体并不形成性泡,少数二价体有中间交叉。     

Freemartins in beef cattle twins induced by embryo transfer

The incidence of freemartinism in heterosexual twins (male-female) resulting from embryo transfer was studied by determining sex chromosome chimerism in lymphocytes and masculinization of female reproductive tracts at slaughter. In one group of calves, ten of 11 heifers born co-twin to full sib, paternal half sib, or unrelated bull calves exhibited sex chromosome chimerism, a proportion in close agreement with that observed in naturally occurring twins. The ten calves with sex chromosome chimerism also had masculinized tracts whereas the other had an apparently normal female tract. Bull calves had a percentage of XY cells similar to their female co-twins, except for the twin set from which the “normal” female was obtained. The bull calf from this set had 5.6% XX cells although no XY cells were observed in the heifer in 66 metaphase spreads. No association was observed between the degree of sex chromosome chimerism and abnormalities of the female tract. Reproductive tracts from all female-female twin sets were normal. In another group of calves, all 20 heifers from heterosexual twin sets had masculinized reproductive tracts. It is concluded that the induction of twins by embryo transfer results in normal expression of freemartinism even though calves may be unrelated and are known to develop in separate uterine horns.     

Cell-autonomous and inductive signals can determine the sex of the germ line of drosophila by regulating the gene Sxl

To investigate the mechanism of sex determination in the germ line, we analyzed the fate of XY germ cells in ovaries, and the fate of XX germ cells in testes. In ovaries, germ cells developed according to their X:A ratio, i.e., XX cells underwent oogenesis, XY cells formed spermatocytes. In testes, however, XY and XX germ cells entered the spermatogenic pathway. Thus, to determine their sex, the germ cells of Drosophila have cell-autonomous genetic information, and XX cells respond to inductive signals of the soma. Results obtained with amorphic and constitutive mutations of Sxl show that both the genetic and the somatic signals act through Sxl to achieve sex determination in germ cells.     

Sex reversal of genetic females (XX) induced by the transplantation of XY somatic cells in the medaka,Oryzias latipes

In order to investigate the function of gonadal somatic cells in the sex differentiation of germ cells, we produced chimera fish containing both male (XY) and female (XX) cells by means of cell transplantation between blastula embryos in the medaka, Oryzias latipes. Sexually mature chimera fish were obtained from all combinations of recipient and donor genotypes. Most chimeras developed according to the genetic sex of the recipients, whose cells are thought to be dominant in the gonads of chimeras. However, among XX/XY (recipient/donor) chimeras, we obtained three males that differentiated into the donor's sex. Genotyping of their progeny and of strain-specific DNA fragments in their testes showed that, although two of them produced progeny from only XX spermatogenic cells, their testes all contained XY cells. That is, in the two XX/XY chimeras, germ cells consisted of XX cells but testicular somatic cells contained both XX and XY cells, suggesting that the XY somatic cells induced sex reversal of the XX germ cells and the XX somatic cells. The histological examination of developing gonads of XX/XY chimera fry showed that XY donor cells affect the early sex differentiation of germ cells. These results suggest that XY somatic cells start to differentiate into male cells depending on their sex chromosome composition, and that, in the environment produced by XY somatic cells in the medaka, germ cells differentiate into male cells regardless of their sex chromosome composition.     

H-Y Antigen Negative Germ Cells in Gonadal Sex Organization in vitro

Dissociation-reorganization experiments were done with gonadal cells of newborn rats. Rotation cultures consisted of mixtures of somatic and germ cells of opposite sex. Somatic cells, ovarian or testicular, determined a female or male type respectively, of gonadal histomorphic organization. Germ cells did not affect the type of organization of somatic cells. Accordingly, suspensions containing somatic cells of one sex together with germ cells of both sexes, reorganized in rotation culture, into either a) follicles containing XX or XY germ cells, or b) tubules containing XX or XY or both types of germ cells. These results give morphological evidence for heterosexual germ-somatic cells interactions. Based on morphological and H-Y antigen studies, failure of germ cells to bind and express H-Y antigen is considered as a possible factor for this failure of germ cells to affect gonadal sex.     

Sex determination in the Drosophila germline is dictated by the sexual identity of the surrounding soma

It has been suggested that sexual identity in the germline depends upon the combination of a nonautonomous somatic signaling pathway and an autonomous X chromosome counting system. In the studies reported here, we have examined the role of the sexual differentiation genes transformer (tra) and doublesex (dsx) in regulating the activity of the somatic signaling pathway. We asked whether ectopic somatic expression of the female products of the tra and dsx genes could feminize the germline of XY animals. We find that Tra(F) is sufficient to feminize XY germ cells, shutting off the expression of male-specific markers and activating the expression of female-specific markers. Feminization of the germline depends upon the constitutively expressed transformer-2 (tra-2) gene, but does not seem to require a functional dsx gene. However, feminization of XY germ cells by Tra(F) can be blocked by the male form of the Dsx protein (Dsx(M)). Expression of the female form of dsx, Dsx(F), in XY animals also induced germline expression of female markers. Taken together with a previous analysis of the effects of mutations in tra, tra-2, and dsx on the feminization of XX germ cells in XX animals, our findings indicate that the somatic signaling pathway is redundant at the level tra and dsx. Finally, our studies call into question the idea that a cell-autonomous X chromosome counting system plays a central role in germline sex determination.     

Testicular Development Induced by a Recessive Mutation during Gonadal Differentiation of Female Common Carp (Cyprinus carpio, L.)

The phenotypic effects of a new recessive mutation mas ⁻¹, which in homozygous condition induces testicular development in XX animals of common carp ( Cyprinus carpio L.), are described. Sexual differentiation of XX; mas ⁻⁺/ mas ⁻¹ and XX; mas ⁻¹/ mas ⁻¹ animals was compared with the gonad development of XX wild type females and XY males. In XX females gonadal differentiation starts with the formation of an ovarian cavity and entry into meiosis of germ cells at around 80 days post hatching (ph). Male gonads remain quiescent until 120 days ph during which period they develop a network of loose connective tissue. Spermatogenesis starts with tubule formation and the differentiation of germ cells into spermatogonia type B. Heterozygous XX; mas ⁻⁺/ mas ⁻¹ animals developed as normal females, but in homozygous XX; mas ⁻¹/ mas ⁻¹ animals two types of gonad development were observed. In the first type, germ cells did not enter meiosis until 100 days ph when they differentiated as spermatogonia. An ovarian cavity was not formed but male specific connective tissue developed instead. These gonad developed as normal testes. In the second type, germ cells differentiated at 80 days ph as either oocytes or spermatocytes, which resulted in the gonads developing as ovotestes. The formation of an ovarian cavity was in most cases incomplete. The phenotypic effects of mas ⁻¹ are interpreted as a timing mismatch between mas activation and female sex differentiation.     

陕西野生啤酒花染色体形态的研究

作者观察了陕西野生啤酒花的染色体数目和形态,分析了这种野生啤酒花及栽培啤酒花的核型,为植物分类及啤酒花育种工作提供了资料。     

Cytogenetic analysis of experimental interspecies goat-sheep chimera

Chromosomal analysis was carried out on blood lymphocytes, skin fibroblasts, and germinal cells of an interspecies goat-sheep chimera. This chimera was produced by aggregation of blastomeres of goat and sheep embryos. A cell chimerism 54,XX/60,XY was found in blood lymphocytes and skin fibroblasts. At birth the percentage of lymphocytes with karyotype 54,XX (sheep) amounted to 80% and with karyotype 60,XY (goat) to 20%. With age the percentage of lymphocytes with chromosome complement 54,XX increased, so that at 18 months it was 94% sheep and 6% goat. At the same age, in skin fibroblasts the percentage of cells with goat karyotype reached 25%. Analysis of germinal cells showed in spermatogonia the presence of only karyotype 60,XY and in primary spermatocytes of 29 autosomal bivalents and the sex bivalent XY.     

Analysis of sex differences in EGC imprinting

Sex-specific differences are apparent in the methylation patterns of H19 and Igf2 imprinted genes in embryonic germ cells (EGCs) derived from 11.5 or 12.5 days post coitum (dpc) primordial germ cells (PGCs). Here we studied whether these differences are associated either with the sex chromosome constitution of the EGCs or with the sex of the genital ridge (testis versus ovary) from which the PGCs were isolated. For this purpose we derived pluripotent EGC lines from sex-reversed embryos, either XY embryos deleted for Sry (XY(Tdym1)) or XX embryos carrying an Sry transgene. Southern blotting of the EGC DNA was used to analyze the differentially methylated regions of Igf2 and H19. The analysis revealed that both genes were more methylated in EGCs with an XY sex chromosome constitution than in those with an XX sex chromosome constitution, irrespective of the phenotypic sex of the genital ridge from which the EGCs had been derived. We conclude that the sex-specific methylation is intrinsic and cell-autonomous, and is not due to any influence of the genital ridge somatic cells upon the PGCs.     

Mammalian X Chromosome Dosage Compensation: Perspectives From the Germ Line

Sex chromosomes are advantageous to mammals, allowing them to adopt a genetic rather than environmental sex determination system. However, sex chromosome evolution also carries a burden, because it results in an imbalance in gene dosage between females (XX) and males (XY). This imbalance is resolved by X dosage compensation, which comprises both X chromosome inactivation and X chromosome upregulation. X dosage compensation has been well characterized in the soma, but not in the germ line. Germ cells face a special challenge, because genome wide reprogramming erases epigenetic marks responsible for maintaining the X dosage compensated state. Here we explain how evolution has influenced the gene content and germ line specialization of the mammalian sex chromosomes. We discuss new research uncovering unusual X dosage compensation states in germ cells, which we postulate influence sexual dimorphisms in germ line development and cause infertility in individuals with sex chromosome aneuploidy.     

Germ cell development in the XXY mouse: evidence that X chromosome reactivation is independent of sexual differentiation

Prior to entry into meiosis, XX germ cells in the fetal ovary undergo X chromosome reactivation. The signal for reactivation is thought to emanate from the genital ridge, but it is unclear whether it is specific to the developing ovary. To determine whether the signals are present in the developing testis as well as the ovary, we examined the expression of X-linked genes in germ cells from XXY male mice. To facilitate this analysis, we generated XXY and XX fetuses carrying X chromosomes that were differentially marked and subject to nonrandom inactivation. This pattern of nonrandom inactivation was maintained in somatic cells but, in XX as well as XXY fetuses, both parental alleles were expressed in germ cell-enriched cell populations. Because testis differentiation is temporally and morphologically normal in the XXY testis and because all germ cells embark upon a male pathway of development, these results provide compelling evidence that X chromosome reactivation in fetal germ cells is independent of the somatic events of sexual differentiation. Proper X chromosome dosage is essential for the normal fertility of male mammals, and abnormalities in germ cell development are apparent in the XXY testis within several days of X reactivation. Studies of exceptional germ cells that survive in the postnatal XXY testis demonstrated that surviving germ cells are exclusively XY and result from rare nondisjunctional events that give rise to clones of XY cells.     

60,XY/60,XX chimerism in the germ cell line of mature bulls born in heterosexual twinning

According to present knowledge there is a germ cell chimerism (XY/XX) in young bulls born in heterosexual twinning due to exchange of primordial germ cells in embryonic life. These germ cells were believed to have been eliminated in the young bull. Two-color fluorescence in situ hybridization (FISH) identification of the sex chromosomes by biotinylated and digoxygenin labeled probes have been used. The material consisted of three bulls born in heterosexual twinning. The results obtained indicated that even mature bulls (more than two years old) demonstrate spermatogonial chimerism. Several authors state that the bulls with blood cell chimerism, originating from dizygous twinning, are characterized by decreased fertility. Changes of the sex ratio of offspring due to proliferation of the female cells have also been proposed. The present observations should give a renewed interest in checking the possibility of survival and differentiation of germ cells from the female partner in the germ cell lines.     

Detection of leucochimaerism in bovine twins by DNA fingerprinting

Karyotyping and hypervariable genetic markers indicate extensive leucochimaerism between pairs of dizygotic twins in cattle, a result of placental vascular anastomosis. The extent of this chimaerism includes both kind and number of cells exchanged. All heterosexual twin pairs harboured two types of leucocytes, having either XX or XY chromosome pairs, and 30 of 31 pairs of twins shared identical DNA fingerprints. Although chromosome results from skin fibroblasts indicate that some chimaerism occurs in the skin, the low level allows for differentiation of genotypes between twins. The results warrant against the common practice of using blood samples for DNA typing if twinning is not properly documented.     

Further studies of chimerism in heterosexual cattle twins

This study was designed to determine whether chimerism in heterosexual twin cattle could be detected in spleen and bone marrow and whether chimeric germ cells could survive into maturity and undergo meiosis. Differential erythrocyte typing and cytogenetic technics were employed. Somatic cell chimerism was usually equal in the various tissues examined. Germ cell chimerism was always low. Meiosis of XX germ cells in testis was detected through diakinesis. Lower ratios for germ cells than for somatic cells were obtained probably because they are mobile only for a short period of time in embryogeny, when they travel from yolk sac to gonadal primordia.     

Different rates of telomere attrition in peripheral lymphocytes in a pair of dizygotic twins with hematopoietic chimerism

Hematopoietic chimerism in dizygotic twins is due to placental vascular anastomoses and arises when hematopoietic stem cells from one twin home to the bone marrow of the other. We report a case of hematopoietic chimerism in a pair of 27-year-old dizygotic twins who each had a mixture of 46,XX and 46,XY blood lymphocytes, both with 98% male (XY) lymphocytes and 2% female (XX) lymphocytes. Analysis of telomere length by T/C FISH revealed that the female twin generally had longer telomeres than the male twin. Moreover, in the male sibling, the telomeres within the female lymphocytes were shortened to 87% of their original length, while the telomeres within the male lymphocytes were 33% longer in the female sibling. Thus, telomere length attrition in peripheral lymphocytes is determined mainly by the environment of the cell and less by intracellular factors.     

Deficiency of X and Y chromosomal pairing at meiotic prophase in spermatocytes of sterile interspecific hybrids between laboratory mice (Mus domesticus) and Mus spretus

The normal association between the X and Y chromosomes at metaphase I of meiosis, as seen in air-dried light microscope preparations of mouse spermatocytes, is frequently lacking in the spermatocytes of the sterile interspecific hybrid between the laboratory mouse strains C57BL/6 and Mus spretus. The purpose of this work is to determine whether the separate X and Y chromosomes in the hybrid are asynaptic, caused by failure to pair, or desynaptic, caused by precocious dissociation. Unpaired X-Y chromosomes were observed in air-dried preparations at diakinesis, just prior to metaphase I. Furthermore, immunocytology and electron microscopy studies of surface-spread pachytene spermatocytes indicate that the X and Y chromosomes frequently fail to initiate synapsis as judged by the failure to form a synaptonemal complex between the pairing regions of the X and Y Chromosomes. Several additional chromosomal abnormalities were observed in the hybrid. These include fold-backs of the unpaired X or Y cores, associations between the autosome and sex chromosome cores, and autosomal univalents. The occurrence of abnormal autosomal and XY-autosomal associations was also correlated with cell degeneration during meiotic prophase. The primary breakdown in hybrid spermatogenesis occurs at metaphase I (MI), with the appearance of degenerated cells at late MI. In those cells, the X and Y are decondensed rather than condensed as they are in normal mouse MI spermatocytes. These results, in combination with the previous genetic analysis of spermatogenesis in hybrids and backcrosses with fertile female hybrids, suggest that the spermatogenic breakdown in the interspecific hybrid is primarily correlated with the failure of XY pairing at meiotic prophase, asynapsis, followed by the degeneration of spermatocytes at metaphase I. Secondarily, the failure of XY pairing can be accompanied by failure of autosomal pairing, which appears to involve an abnormal sex vesicle and degeneration at pachytene or diplotene.by C. Heyting     

Mutant meiotic chromosome core components in mice can cause apparent sexual dimorphic endpoints at prophase or X–Y defective male-specific sterility

Genetic modifications causing germ cell death during meiotic prophase in the mouse frequently have sexually dimorphic phenotypes where oocytes reach more advanced stages than spermatocytes. To determine to what extent these dimorphisms are due to differences in male versus female meiotic prophase development, we compared meiotic chromosome events in the two sexes in both wild-type and mutant mice. We report the abundance and time course of appearance of structural and recombination-related proteins of fetal oocyte nuclei. Oocytes at successive days post coitus show rapid, synchronous meiotic prophase development compared with the continuous spermatocyte development in adult testis. Consequently, a genetic defect requiring 2–3 days from the onset of prophase to reach arrest registers pachytene as the developmental endpoint in oocytes. Pachytene spermatocytes, on the other hand, which normally accumulate during days 4–10 after the onset of prophase, will be rare, giving the appearance of an earlier endpoint than in oocytes. We conclude that these different logistics create apparent sexually dimorphic endpoints. For more pronounced sexual dimorphisms, we examined meiotic prophase of mice with genetic modifications of meiotic chromosome core components that cause male but not female sterility. The correlations between male sterility and alterations in the organization of the sex chromosome cores and X–Y chromatin may indicate that impaired signals from the XY domain (XY chromosome cores, chromatin, dense body and sex body) may interfere with the progression of the spermatocyte through prophase. Oocytes, in the absence of the X–Y pair, do not suffer such defects.     

The XYY condition in a wild mammal: an XY/XYY mosaic common shrew (Sorex araneus)

XY/XYY sex-chromosome mosaicism was demonstrated in both bone marrow and germ cells of a wild adult common shrew. Secondary sexual characteristics were those of a normal male, but the testes were small, and the sperm count was only about 3% of normal. Most of the seminiferous tubule cross-sections examined revealed serious spermatogenic impairment and a reduced diameter. A range of sex-chromosome pairing configurations was observed in XYY primary spermatocytes, including configurations involving the X and both Y chromosomes in a linear or radial array. The presence of metaphase II (MII) spreads with an XY sex-chromosome complement indicated that XYY primary spermatocytes could contribute products to MII. Following Burgoyne (1979) and Burgoyne and Biddle (1980), a number of models of spermatocyte loss were tested. The data indicated that there was an association between the sex-chromosome complement of primary spermatocytes and their contribution to MII. The best fit to the observed MII frequency data was provided by a model which assumed that all XYY primary spermatocytes with a univalent Y chromosome and a high proportion of XYY primary spermatocytes with an unpaired X chromosome failed to contribute products to MII.