XY pair associates with the synaptonemal complex of autosomal male-sterile translocations in pachytene spermatocytes of the mouse (Mus musculus)

Analysis of the chromosome behaviour at pachytene has been performed by means of the silver staining technique visualizing the synaptonemal complexes (SCs) in male mice heterozygous for the male-sterile translocations T(5;12)31H, T(16;17)43H and T(7;19)145H, respectively. The T(9;17)138Ca male heterozygotes and T43H/T43H homozygous males were used as fertile controls. The sterile mice displayed a high frequency (about 60%) of pachytene spermatocytes with autosomal translocation configuration located in close vicinity of the XY pair. The dense round body (XAB), normally located near the X-chromosome axis in fertile males, exhibited abnormal affinity to translocation configuration in the sterile translocation heterozygotes. The incomplete synapsis of autosomes involved in translocation configuration was observed in more than 70% of the pachytene spermatocytes with the male-sterile translocations but in less than 20% of the cells from T138Ca fertile male.s. A hypothesis relating the spermatogenic arrest of carriers of male-sterile rearrangements to the presumed interference with X chromosome inactivation in male meiosis is discussed.     

小麂、黑麂、赤麂精母细胞联会复合体的比较研究

本工作以界面铺张——硝酸银染色技术,对小麂(Muntiacus reeuesi)、黑麂(M.crinifrons)和赤麂(M.muntjak)的精母细胞联会复合体(Syna ptonemal complex,SC)进行亚显微结构的比较研究。结果表明: 1.SC的平均相对长度和臂比指数同有丝分裂细胞相应染色体的数值有很好的一致性。根据SC的相对长度和臂比指数绘制了三种麂的SC组型图。雄性黑麂减数分裂前期形成一个复杂的易位多价体,意味着其核型的演化过程涉及两次染色体易位和一次臂间倒位。 2.在减数分裂前期,性染色体的形态和行为同常染色体的有明显差异,如性染色体嗜银性较强,配对延迟等。XY的配对起始于早粗线期,在中粗线期,Y的全长均同X配对;XY-SC开始解体于晚粗线期。 3.在粗线期,X染色体未配对区域出现自身折叠,形成“发夹”状结构。这种“发夹”结构的形成,可能是在性染色体的进化过程中,X染色体通过不对称易位得到的重复片段在减数分裂前期同源配对的一种细胞学表现。     

2例男性育性障碍患者的联会复合体分析

运用表面铺展联会复合体（synaptonemal complex,SC）的电镜技术对2例男性育性障碍患者的联会复合体进行分析,发现患者1的部分粗线期精母细胞（20%）中出现配对紊乱（一些SCs中间未完全配对,未配对区轴心出现增粗,类似性染色体配对行为;有些SC仅两端配对,形成很短的两段SC,中间大部分未配对,在未配对区出现断裂,似乎是一个三价体和单价体）现象,大部分细胞配对正常。患者2的几乎100%生精细胞被阻断在减数分裂的前期阶段,显示联会异常如SC粉碎化、SC侧生组分膨化等现象。该文对男性育性障碍的机理进行了讨论。 Abstract:Synaptonemal complexes (SCs) of male fertility impairment were analysed in two patiants.In case one,his testicular histology was normal and there were 20% abnormal pachytene spermatoeytes,showing unsynaptic and broken SCs.It resulted in spermatogensis with unbalanced chromosomes and pregnancy wastages.In case two,he had no sperms basically.G-banded chromosome analysis of lymphocytes showed normal chromosomal karyotype,but triple fragment,lateral element swelling and fragment breakages of SCs were observed in his spermatocytes.The mechanisms of infertility,impairment of fertility for the observed men were discussed in this paper.     

Banding studies and synaptonemal complex analysis of an X-autosome translocation in the domestic pig

In a litter of nine domestic pigs, a translocation between the X-chromosome and chromosome 13 was found in six individuals: four males and two females. The translocation was presumed to have originated in the dam. Banding studies indicated that the breaks preceding the translocation had occurred in a distal GTG-negative band of the long arm of the X, 15-30% of the length of Xq from the telomere, and proximally in chromosome 13, 15-25% from the centromere. The normal X of the females invariably replicated its DNA late. Synaptonemal complex analysis of spermatocytes demonstrated a quadrivalent in 75 of 85 analyzable cells (88.2%), and in 10 cells (11.8%) one trivalent and one univalent were found. Extensive nonhomologous pairings were visualized in the pachytene stage by applying an 'overlap' test measuring the sex chromosomes and collating their pairings. An arrest in male meiosis was verified histologically; no meiotic stages later than pachytene developed. This resulted in sterility, with considerable testicular hypoplasia. The records of female fertility were available only for the dam and did not show any deviations from the average of the herd.     

Molecular cytogenetic characterization of a familial balanced reciprocal translocation t(11;18) (q13.3; q23) associated with pregnancy wastage

A new male patient associated with a pregnancy wastage was detected in China. Cytogenetic analyses including G-banding, chromosome painting and observation of synaptonemal complexes (SCs) demonstrated that the pregnancy wastage was associated with a balanced reciprocal translocation t(11;18) (q13.3; q23). The proband was the carrier of the translocation and his karyotype was 46,XY,t(11;18)(11pter-->11q13.3:: 18q23-->18qter; 18pter-->18q23::11q13.3-->11qter). The pedigree was analyzed based on a G-banded karyotype of the nine familial members. The translocation chromosomes came from the proband's mother. The result of the SC observation in the proband showed that each of the spermatocytes displayed one quadrivalent during their pachytene stages. In the quadrivalents, there existed homologous and nonhomologous synapses and the latter occurred widely during early, middle and late pachytene stages. The reasons and genetic basis of the pregnancy wastage are discussed.     

Inter-sex variation in synaptonemal complex lengths largely determine the different recombination rates in male and female germ cells

Meiotic chromosomes in human oocytes are packaged differently than in spermatocytes at the pachytene stage of meiosis I, when crossing-over takes place. Thus the meiosis-specific pairing structure, the synaptonemal complex (SC), is considerably longer in oocytes in comparison to spermatocytes. The aim of the present study was to examine the influence of this length factor on meiotic recombination in male and female human germ cells. The positions of crossovers were identified by the DNA mismatch repair protein MLH1. Spermatocytes have approximately 50 crossovers per cell in comparison to more than 70 in oocytes. Analyses of inter-crossover distances (and presumptively crossover interference) along SCs suggested that while there might be inter-individual variation, there was no consistent difference between sexes. Thus the higher rate of recombination in human oocytes is not a consequence of more closely spaced crossovers along the SCs. The rate of recombination per unit length of SC is higher in spermatocytes than oocytes. However, when the so-called obligate chiasma is excluded from the analysis, then the rates of recombination per unit length of SC are essentially identical in the two sexes. Our analyses indicate that the inter-sex difference in recombination is largely a consequence of the difference in meiotic chromosome architecture in the two sexes. We propose that SC length per se, and therefore the size of the physical platform for crossing-over (and not the DNA content) is the principal factor determining the difference in rate of recombination in male and female germ cells. A preliminary investigation of SC loop size by fluorescence in situ hybridization (FISH) indicated loops may be shorter in oocytes than in spermatocytes.     

Synaptonemal complex karyotype of zebrafish

Meiotic cells of zebrafish have been prepared to show synaptonemal complexes (SCs) by light and electron microscopy. Completely paired SCs from both spermatocytes and oocytes were measured to produce an SC karyotype. The SC karyotype resembles the somatic karyotype of zebrafish and has no recognisable sex bivalent. Measurements of total SC length indicate that SCs grow longer and develop centromeres during pachytene. Oocytes consistently have longer SCs than spermatocytes, presumably correlated with the reported higher recombination frequency in females than in males.     

家鸡联会复合体的亚显微结构分析

本文以表面铺展——硝酸银染色技术,对家鸡的联会复合体(Syneptonemal Complex,SC)作亚显微结构分析。根据对10个精母细胞和10个卵母细胞SC的测量结果,绘制组型图。发现雌雄家鸡的常染色体的SC组型相同。在精母细胞中,性染色体(ZZ)的行为与常染色体相似。在卵母细胞中,性染色体ZW的长度不同,长轴为Z,短轴为W,两者之间只有部分配对,形成SC。从早粗线期到晚粗线期,由同源配对调整为非同源配对。另外,在一只雌鸡中,第一次观察到,有些细胞的常染色体能正常配对,而性染色体完全不配对的现象。     

Delayed Formation of Chromosome Aberrations in Mouse Pachytene Spermatocytes Treated with Triethylenemelamine (Tem)

Induction of chromosome aberrations in pachytene spermatocytes of mice by 2 mg/kg TEM was compared with induction by 400 R X rays. These doses induced comparably high dominant lethal effects in pachytene spermatocytes of mice. Cytological analysis at diakinesis–metaphase I stage showed that whereas 76.4% of the cells treated with X rays at pachytene stage had aberrations, the frequencies observed in two TEM experiments were only 0.8 and 2.2%. On the other hand, 5% of the progeny from TEM-treated pachytene spermatocytes were found to be translocation heterozygotes. This is the first report on the recovery of heritable translocations from treated spermatocytes of mice. The aberration frequencies observed for TEM in diakinesis–metaphase I were much too low to account for all the lethal mutations and heritable translocations. Thus, the formation of the bulk of aberrations induced by TEM in pachytene spermatocytes was delayed—a marked contrast to the more immediate formation of X-ray-induced aberrations. It is postulated that the formation of the bulk of TEM-induced aberrations in pachytene spermatocytes and in certain postmeiotic stages occurs sometime during spermiogenesis, and not through the operation of postfertilization pronuclear DNA synthesis.     

■普斑限性与■虎斑限性品系家蚕的联会复合体分析

作者以去污剂铺展——硝酸银染色技术,对家蚕普斑限性及虎斑限性品系雌蚕的联会复合体进行亚显微观察分析,未发现品系有异形ZW;但在品系中发现ZW呈现明显的末端不对称性。联会复合体分析表明该品系的ZW两侧线的相对长度相差达5.2％,较长的侧线为Z染色体,较短的侧线是断裂后的W染色体和第Ⅳ连锁群带有Ze片染的相互转座。ZW-SC在晚粗线期开始分离,较其他SC分离显著提前。这可能与易位导致ZW的不同源区段增长有关。     

A pachytene analysis of two male-fertile paracentric inversions in chromosome 1 of the mouse and in the male-sterile double heterozygote

Meiotic studies have been made at pachytene on two paracentric inversions in chromosome 1 of the mouse. Surface-spread preparations of primary spermatocytes have been analysed at the light microscope level in males heterozygous for the inversions In(1)1Rk and In(1)12Rk and in the double heterozygote In(1)1RK/In(1)12Rk. In singly heterozygous form, neither inversion produces any serious effect on male fertility. In the double heterozygote, spermatogenesis is arrested in the majority of cells at the spermatocyte stage and males are rendered totally sterile by azoospermia. In the double heterozygote, a complex loop, indicating the inversion bivalent, is found in 90% of pachytene cells analysed. In the In(1)1Rk/+ heterozygote, a looped bivalent was seen in 47 per cent of pachytene cells but in In(1)12Rk/+ no cells containing loops could be found. -80% of pachytene spermatocytes from the In(1)1Rk/In (1)12Rk double heterozygote showed apposition of the inversion bivalent to the sex bivalent. Such an association was rarely seen in pachytene cells of either of the fertile single heterozygotes. Spermatogenic failure in the double heterozygote may be related to interference, by the inversion bivalent, with X chromosome inactivation at meiotic prophase.     

Immunocytogenetics. III. Analysis of trivalent and multivalent configurations in mouse pachytene spermatocytes by human autoantibodies to synaptonemal complexes and kinetochores

Mice heterozygous for one or more Robertsonian (Rb) translocation chromosomes have been used to analyze synaptonemal complex (SC) configurations and kinetochore arrangements in trivalents and multivalents. Rb heterozygosity without arm homologies leads to the formation of heteromorphic trivalents in meiosis I; alternating homology of the chromosome arms produces ringlike or chainlike multivalents. Immunofluorescence double-labeling with human antibodies to SCs and kinetochores was performed on surface-spread pachytene spermatocytes. Both Rb bivalents and Rb trivalents clearly showed that metacentrics possess only one centromere. In heteromorphic trivalent SCs, the nonhomologous kinetochores of the two acrocentrics were closely paired in a cis-configuration and juxtaposed opposite the kinetochore of the metacentric; the latter appeared to be an integral part of the longitudinal SC axis. Meiotic multivalents of interpopulation hybrids included up to 36 chromosome arms. In multivalent SCs, the kinetochores always lay together, with the SC arms arranged away from the central centromere cluster. The paracentromeric regions of the Rb chromosomes appeared to remain unsynapsed on both sides of the centromeres. The SC arms were often linked by end-to-end associations. Following desynapsis of the multivalent SC, the kinetochores of the Rb metacentrics showed a highly nonrandom topologic distribution within the nucleus, reminiscent of their arrangement during synapsis.     

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.     

Pachytene analysis in males heterozygous for a familial translocation (9;12;13) (q22; q22; q32) ascertained through a child with partial trisomy 9

A family with four male and three female heterozygotes for a three-way translocation (9;12;13) (q22; q22; q32) in three generations was ascertained through a chromosomally imbalanced newborn with an additional derivative chromosome 9 resulting from nondisjunction. Three heterozygous males from two generations with apparent differences in their fertility status were investigated using pachytene spreads and testicular histology. Pachytene analysis in all three individuals, the fertile (II-2) as well as the subfertile (III-7) and infertile (III-9), showed a hexavalent with central nonpairing around the translocation breakpoints in nearly all spermatocytes. Thus, the observed hexavalent configurations in pachytene do not seem to have caused impaired fertility. This rather may have been the result of sperm carrying unbalanced chromosome sets. However, the observed difference in fertility between the heterozygous fertile male in generation II and his two heterozygous sons remains unexplained.     

六种鱼的精母细胞联会复合体的电镜观察

我们以界面铺张——硝酸银染色技术,对鲈形目三种鱼(尼罗罗非鱼、莫桑比克罗非鱼、刺鳅)和鲤形目(鱼句)亚科三种鱼(花(鱼骨)、黑鳍鳈、麦穗鱼)的精母细胞联会复合体进行了电镜观察研究。系统考察了鱼类常染色体SC的亚显微结构、形成过程和配对行为,比较分析了刺鳅的性染色体SC的异配形态和行为,并绘制了鲈形目三种鱼的SC组型模式图。     

X-Y chromosome univalency in the testes of hyperthermic mice: II. Light and electron microscopic analysis of synaptonemal complexes

Hyperthermia-induced X-Y dissociation has been observed in diakinesis-metaphase I sper-matocytes but not in pachytene spermatocytes, which have been implicated as highly susceptible to heat stress. To determine X-Y dissociation in pachytene spermatocytes and to compare levels of dissociation between pachytene and diakinesis-metaphase I spermatocytes male ICR mice were exposed to 35°C ± 0.07°C and 65% ± 0.14% relative humidity for 2 or 4 days. Control mice were housed at 24°C ± 0.04°C and 43% ± 0.58% relative humidity. Mice were killed 0, 3, 5, 8, or 10 days after stress and the testes processed for meiotic chromosome display at diakinesis-metaphase I and synaptonemal complex display at pachynema. Twenty-five to thirty cells per mouse at both stages of meiosis were observed with light microscopy, and pachytene spreads with transmission electron microscopy to determine heat-stress effects on synaptonemal complex structure. Statistical analyses revealed significant linear increases in X-Y dissociation with prolonged exposure to heat at pachynema and diakinesis-metaphase I. Levels of pachytene dissociation were one-half the levels of dissociation at diakinesis-metaphase I. The resolvable structure of the lateral elements of the synaptonemal complex was not affected by heat stress.     

Meiosis in gray voles of the subgenus microtus (rodentia, arvicolinae) and in their hybrids

The results of light and electron microscopic (EM) studies of meiosis in Microtus arvalis males of the karyoform "arvalis" (2n = 46, NFa = 80), in hybrids between the chromosomal forms arvalis and obscurus (2n = 46, NFa = 68), in M. rossiaemeridionalis voles (2n = 54, NFa = 54), and in a hybrid between the species M. rossiaemeridionalis and M. kermanensis (2n = 54, NFa = 54) are presented. SC (synaptonemal complex) karyotypes of the parental forms and the hybrids were constructed on the basis of measurements of the length ofautosomal SCs revealed by the EM analysis in spermatocytes at the stage of middle pachytene. The SC karyotypes of M. arvalis and the hybrids female obscurus x male arvalis consist of 22 synaptonemal complexes of autosomal bivalents and the axial elements of the synaptonemal complexes of the sex chromosomes X and Y. The SC karyotypes of M. rossiaemeridionalis and the hybrid M. rossiaemeridionalis x M. kermanensis consist of 26 synaptonemal complexes of autosomal bivalents and a sex bivalent; they differ only in the length of the Y chromosome axis (Y chromosome in the hybrid was inherited from M. kermanensis). Asynaptic configurations of the autosomal SCs were not observed in the hybrids. The SC axial elements of the X and Y chromosomes in the parental forms and in the hybrids were located close to each other throughout pachytene, but they did not form a synaptic region. The normal synapsis in sterile hybrids (M. rossiaemeridionalis x M. kermanensis) and the behavior of the sex chromosomes in meiosis in fertile and sterile hybrids are discussed in the context of specific features of meiosis and reproductive isolation.     

Spermatocytes of the caddisfly Potamophylax rotundipennis (Trichoptera, Insecta): a fine structure study with emphasis on synaptonemal complex plates associated with chromatin

Electron microscopy of ultrathin sections was used to study the restructuring of primary spermatocytes in a caddisfly, Potamophylax rotundipennis (Limnephilidae). Spindle structure was also examined using light microscopy of dividing spermatocytes lysed in a microtubule-stabilizing buffer. The bulk of pachytene spermatocytes was usual in that the nuclei contained tripartite synaptonemal complexes (SCs). The SCs were attached end-on to the inner face of the nuclear envelope and loosely surrounded by electron-dense chromatin. Cells of this type gave rise to late prophase I spermatocytes, where SCs were missing and chromatin condensation was advanced. By metaphase I, a conventional bipolar spindle apparatus assembled, bivalents were aligned at the spindle equator, and membrane sheets were scattered throughout the spindle matrix. Prominent interzone spindles were typical of telophase spermatocytes. However, a subset of prophase I spermatocytes possessed unusual forms of SCs. The analysis of short series of ultrathin sections through the nuclei revealed plates composed of synaptonemal complex material. These elements will be referred to as 'SC plates'. Within the SC plates, the tripartite organization typical of regular SCs was preserved. The chromatin surrounding the SC plates was highly condensed. The SC plates ended abruptly within the nuclear lumen and did not reach the nuclear envelope. Finally, branching of SC plates was common. In light of the bizarre organization of SC material and its relation to the chromatin, and because spermatocytes with SC plates do not readily fit into the regular development of male germ cells in the caddisfly, we venture the suggestion that the SC plates are not physiological intermediates of SC disassembly. The affected cells most probably fail to complete meiosis.     

Synaptonemal complex analysis of mouse chromosomal rearrangements

Electron microscopy of surface-spread spermatocytes from mice heterozygous for a tandem duplication shows the heteromorphic synaptonemal complex (SC) to comprise two lateral elements of unequal length, the longer of which is buckled out in a characteristic loop, representing the unsynapsed portion of the duplication. The loop is a regular feature of late zygotene-early pachytene nuclei; it is longest at these early stages, but, through equalization of the two axes as a consequence of synaptic adjustment, it is replaced by a normal appearing SC at late pachytene. Because equalization, as indicated by a decrease in the percent difference between axes, may begin shortly after completion of synapsis, estimates of duplication segment length are restricted to a sample selected for least adjustment. — Although the mean position of the loop is constant at various pachytene substages, individual positions vary widely from cell to cell, consistent with the behavior expected of a duplication, but not of a deletion or an inversion. The length of the segment that is duplicated is estimated to be 22% of the normal chromosome, the midpoint of the segment is mapped at 0.61 of the chromosome distal to the kinetochore, and the ends of the segment are mapped at 0.50 to 0.72. Measurements of G-banded mitotic chromosomes give comparable values: duplication length, 24%; midpoint, 0.60, and segment ends, 0.48 and 0.71. This agreement constitutes further validation of the SC/spreading method for detecting and analyzing chromosomal rearrangements at pachytene and substantiates the fidelity with which the axes and SCs represent the behavior of chromosomes in synapsis.