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PubMed2005

Understanding the cytological diploidization mechanism of polyploid wild wheats.

The allohexaploid Aegilops species (2n = 6x = 42), Ae. neglecta 6x (UUXtXtNN), Ae. juvenalis (DcDcXcXcUU), and Ae. vavilovii (DcDcXcXcSsSs) regularly form bivalents at metaphase I. However, in Ae. crassa 6x (DcDcXcXcDD) 0.27 quadrivalents per cell were observed probably as a consequence of the partial homology displayed by the D and Dc genomes. Likewise, the synthetic amphiploid Ae. ventricosa-Secale cereale (DDNNRR) is fertile and displays a diploid-like behavior at metaphase I, despite its recent origin. The pattern of synapsis at late zygotene and pachytene in the natural and artificial allohexaploids was analyzed by whole-mount surface-spreading of synaptonemal complexes under an electron microscope. It revealed that chromosomes were mostly associated as bivalents in all cases, the mean of multivalents per nucleus ranging from 0.17 (Ae. neglecta 6x) to 1.03 (Ae. crassa 6x) in the natural species and 1.05 in the Ae. ventricosa-S. cereale amphiploid. It can be concluded that the mechanism controlling bivalent formation in these species and also in the synthetic amphiploid acts mainly at zygotene by restricting synapsis to homologous chromosomes, but also acts at pachytene by preventing chiasma formation in the homoeologous associations. These observations are discussed in relation to the origin and evolution of the mechanism of diploidization in the allopolyploid species of the Poaceae family.

Chromosome Pairing↗
PubMed2005

Meiotic abnormalities in infertile males.

Meiotic anomalies, as reviewed here, are synaptic chromosome abnormalities, limited to germ cells that cannot be detected through the study of the karyotype. Although the importance of synaptic errors has been underestimated for many years, their presence is related to many cases of human male infertility. Synaptic anomalies can be studied by immunostaining of synaptonemal complexes (SCs), but in this case their frequency is probably underestimated due to the phenomenon of synaptic adjustment. They can also be studied in classic meiotic preparations, which, from a clinical point of view, is still the best approach, especially if multiplex fluorescence in situ hybridization is at hand to solve difficult cases. Sperm chromosome FISH studies also provide indirect evidence of their presence. Synaptic anomalies can affect the rate of recombination of all bivalents, produce achiasmate small univalents, partially achiasmate medium-sized or large bivalents, or affect all bivalents in the cell. The frequency is variable, interindividually and intraindividually. The baseline incidence of synaptic anomalies is 6-8%, which may be increased to 17.6% in males with a severe oligozoospermia, and to 27% in normozoospermic males with one or more previous IVF failures. The clinical consequences are the production of abnormal spermatozoa that will produce a higher number of chromosomally abnormal embryos. The indications for a meiotic study in testicular biopsy are provided.

Chromosome Aberrations↗
PubMed2005

IgE-mediated rat mast cell triggering with tryptic and synthetic peptides of bovine beta-lactoglobulin.

BACKGROUND: Immunoglobulin E (IgE) epitopes of beta-lactoglubulin (betaLG) have been identified by ELISA inhibition methods using sera from allergic patients. However, the functional capacity of these epitopes to stimulate mast cells is unknown. It is the goal of the present study to identify bivalent IgE epitopes of betaLG able to trigger target mast cells. METHODS: Peptides were obtained either by purification from tryptic hydrolysates of betaLG or by synthesis. They were examined for their triggering activity in vitro on peritoneal 3H-serotonin-labeled rat mast cells passively sensitized with IgE anti-betaLG antibodies. In vivo, rats immunized with betaLG were administered peptides by gavage for intestinal rat mast cell protease II release. RESULTS: Compared with intact betaLG, purified or synthetic tryptic-like betaLG peptides have a sharply decreased allergenicity. Peptide 149-162 retains the highest bivalent IgE epitope-mediated triggering capacity. CONCLUSION: A functional bivalent IgE epitope was identified at the C terminal end of betaLG.

Amino Acid Sequence↗
PubMed2006

The high variability of subtelomeric heterochromatin and connections between nonhomologous chromosomes, suggest frequent ectopic recombination in rye meiocytes.

The position of telomeres, centromeres and subtelomeric heterochromatin (SH) has been studied by FISH in rye meiocytes. We compare the morphology of the signals from zygotene to telophase II mainly to determine differences in SH and telomere positions between plants with and without neocentromeres. Plants from two varieties were used: Paldang showing neocentromeres, and Puyo without neocentromeres but with two B chromosomes. In both varieties, at zygotene and pachytene the SH is observed forming clumps often including two or more bivalent ends. At diplotene the SH is stretched suggesting that it is close to the nuclear envelope. In these cases, the telomere signals are not stretched and lay behind the SH. Frequently, two or more bivalents are joined by conspicuous SH connections at diplotene strongly suggesting ectopic recombination. Probably as a result, differential distribution of the SH between recombinant homologues or the whole meiotic products is observed. From diplotene onwards, the large heterochromatic blocks cover the telomeres, the SH being the morphological end of the bivalents, both in plants with or without neocentromeres. The Bs are tightly associated only at the telomeric end of the long arm from diplotene to metaphase I. The high variability between homologous chromosomes and the frequent nonhomologous bindings of SH, strongly suggest that rye SH is in dynamic state and frequently changes in chromosome position during meiosis.

Chromosomes, Plant↗
PubMed1986

Recombination nodules in the oocytes of the chicken, Gallus domesticus.

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.

Animals↗
PubMed1987

The nature of the 1;29 translocation in cattle as revealed by synaptonemal complex analysis using electron microscopy.

Synaptonemal complex analyses were carried out by electron microscopy on surface-spread spermatocytes of one normal bull and two bulls that were heterozygous for the so-called 1;29 translocation. The autosomal bivalents of the normal karyotype, which could be arranged by size in a series, demonstrated kinetochores at the terminally located attachment plaques. One autosomal bivalent was clearly larger than the rest and apparently consisted of the long arm of the 1;29 translocation. The 1;29 translocation was the longest autosome in the set and had a kinetochore in a subtelocentric position. Some of the autosome pairs had nucleolus organizer regions in telomeric regions. The X and Y chromosomes, which were not paired at zygotene, demonstrated association in a very short segment at early pachytene; in no cells could a synaptonemal complex be seen between the X and Y. Very often the sex chromosomes were dissociated. At zygotene, a few, usually large, bivalents were unpaired proximally. This always also involved the proximal parts of the arms of the 1;29 translocation and their normal homologs. At early pachytene, the 1;29 trivalent, although to a less extensive degree, was also unpaired in the pericentric region. Configurations in which one chromosome, either 1 or 29, was completely paired with its corresponding arm in the 1;29 translocation chromosome also occurred. When unpaired proximally, the size of chromosome 1 agreed fairly well with the size of its corresponding arm, but the size of chromosome 29 was considerably larger than the corresponding arm of the 1;29 translocation chromosome. During late zygotene and early pachytene, the percent difference between chromosome 29 and its corresponding arm decreased, and at mid and late pachytene there had been a complete synaptic adjustment. The size difference and pairing behavior indicated that a deletion of the kinetochore and the most proximal segment of chromosome 29 had preceded the fusion with chromosome 1 into the 1;29 translocation. The unique structural appearance of the 1;29 translocation chromosome compared to that of other centric fusion translocations in cattle lends support to the theory of a monophyletic origin of the 1;29 translocation. The importance of the pairing behavior observed in governing recombination and chromosome disjunction is briefly discussed.

Animals↗
PubMed1988

An analysis of the chromomere map and chiasmata characteristics of human diplotene spermatocytes.

A complete chromomere map of early/mid diplotene human spermatocytes has been developed which permits identification of each bivalent. Bivalents 9, 16, 17, and 19 demonstrated unique cytogenetic characteristics at this meiotic stage. The mean chiasma frequency per spermatocyte was 45.33 +/- 4.52 (ranging from 32 to 58) with 28% of bivalents having one chiasma, 38% having two, and 27% having three. The remaining 7% had four or more chiasmata. Fifty-eight percent of chiasmata were located distally, 31% centrally, and 11% proximally. Univalents were rare. The availability of human diplotene spermatocyte maps permits exploration of many basic questions of recombination with accuracy.

Adult↗
PubMed1988

Pachytene pairing in relation to sperm and oocyte numbers in a male-fertile reciprocal translocation in the mouse.

In adult males carrying the male-fertile reciprocal translocation T(2;4)13H, body weights, testis weights, and sperm counts were higher in heterozygotes than in homozygotes. Heterozygotes whose mothers were C3H/He exceeded their reciprocal counterparts in the same criteria. At 3-4 days of age, no significant differences between homozygous and heterozygous females were found in body weight, ovarian volume, or oocyte numbers, although mean oocyte volumes were somewhat larger in heterozygotes than in homozygotes. In homozygous males and females the synaptonemal complexes of rearranged chromosomes appeared as bivalents that were indistinguishable from normal bivalents. In most gametocytes of heterozygotes, the translocation was present in the form of a quadrivalent. The degree of pairing failure was greater in oocytes than in spermatocytes. Terminal asynapsis of quadrivalents was very rare in spermatocytes, but it affected one quarter of the oocytes. Only very few translocation configurations were associated with the XY bivalent. It is concluded that the number of sperm produced in male heterozygotes can match the general increase in vigor by the formation of a high level of fully paired quadrivalents, whereas a greater degree of terminal asynapsis in the quadrivalents of oocytes may indicate a slightly more deleterious effect of this translocation on oogenesis.

Animals↗
PubMed1989

Synaptic adjustment in Peromyscus beatae (Rodentia: Cricetidae) heterozygous for interstitial heterochromatin.

Chromosomal pairing and chiasma formation were studied two individuals of Peromyscus beatae heterozygous for the presence of a large block of interstitial heterochromatin. Although the modified chromosome was of medium size, analysis of C-banded diakinetic configurations revealed that it was the homolog of one of the smallest autosomes. Analysis of silver stained synaptonemal complexes indicated that synapsis was either unidirectional from initiation at one set of telomeres or was bidirectional from initiation at both sets of telomeres. Each pattern resulted in characteristic heteromorphic pairing configurations (interstitial asynapsis or terminally positioned unpaired segments) in early pachynema. These configurations underwent synaptic adjustment and, by mid-pachynema, the lateral elements of the polymorphic bivalent either appeared typical of homomorphic bivalents or exhibited regional heteropycnosis in one or both axes. Synaptonemal complex data for Peromyscus and many other mammalian species reflect an apparent need for fully paired, linear bivalents prior to the end of pachynema.

Animals↗
PubMed1989

Meiosis in trisomic female mice with Robertsonian translocations. II. Chromosome behaviour at first and second meiotic metaphases.

First and second meiotic metaphases (MI and MII, respectively) from female mice of Robertsonian translocation (Rb) stock, trisomic for chromosome 16 (Ts16) or 19 (Ts19), were studied. The mature trisomic oocytes were derived from explanted fetal ovaries that had been cultured and then transplanted so as to mature heterotopically. Multivalent configurations involving the Rb chromosomes and the additional trisomic acrocentric were analysed. Pentavalent configurations occurred in 74.5% of 98 Ts16 MI and 44.2% of 249 Ts19 MI oocytes; quadrivalents (with a univalent acrocentric) were found in 9.2% of Ts16 MI and 10.8% of Ts19 MI oocytes. In 1% of Ts16 MI and 4% of Ts19 MI oocytes, there were two Rb bivalents and a univalent trisomic acrocentric. Rb trivalents and Rb bivalents occurred together in 14.3% of Ts16 MI and 39.4% of Ts19 MI oocytes. Chiasma frequencies were similar in trisomic and chromosomally balanced MI. Chiasma position, distribution, and localization were nearly identical, whether they were found in Rb multivalents or acrocentric bivalents, but one control group (from chromosomally balanced Ts19 littermates) had significantly more terminal chiasmata. Within the triple homologous region of 8% of Rb pentavalents, two chiasmata were observed in the same relative position in the two sister chromatids of one of the three homologs, suggesting a lapse in chiasma position interference. Assortment at MI anaphase was influenced by secondary nondisjunction of the Rb. The ratio of balanced to unbalanced MII oocytes was 1:4 in both trisomies.

Animals↗
PubMed1990

Pachytene pairing and oocyte numbers in mice with two single Robertsonian translocations and the male-sterile compound with monobrachial homology.

Oocyte numbers and synaptonemal complexes were studied in two Robertsonian translocations, Rb(6.15)1Ald and Rb(4.6)2Bnr, and their male-sterile compound. Oocyte numbers in the compound were lower than those of either parent, and there was a marked difference between reciprocal crosses. Synaptonemal complexes of homozygous females appeared as 19 bivalents, those of single heterozygotes as 18 bivalents and a trivalent, and those of compound heterozygotes as 17 bivalents and a quadrivalent. Most trivalents were fully paired, whereas the majority of quadrivalents exhibited terminal asynapsis. About one-half of all oocytes had other pairing abnormalities, probably reflecting reduced survivability. Whereas all fully paired quadrivalents were present in cells not showing any pairing anomalies, one-half of the quadrivalents with terminal asynapsis were seen in oocytes with other anomalies. It is suggested that in oocytes destined for atresia, there is a predisposition to synaptic failure of translocation configurations. Additional oocytes are likely to break down because of the deleterious effect of the compound translocation on gametogenesis. This effect seems to be more pronounced in Rb1Ald/Rb2Bnr spermatocytes than in oocytes.

Animals↗
PubMed1992

Synapsis, recombination, and meiotic segregation in the mesquite lizard, Sceloporus grammicus, complex. I. Pericentric inversion heteromorphism of the F5 cytotype.

Chromosomal pairing and recombination were analyzed in male specimens of Sceloporus grammicus heterozygous for a large pericentric inversion of macrochromosome 4. Analysis of silver-stained synaptonemal complexes (SCs) in surface-spread nuclei revealed that homologously paired inversion loops were not formed. Synapsis of the inverted segments proceeded directly to nonhomologous straight pairing. In some nuclei, this resulted in a configuration that could not be distinguished from homozygous bivalents of similar size. Examination of Giemsa- and silver-stained diakinetic nuclei indicated that crossing-over was limited to the noninverted (homologous) portion of the heteromorphic bivalent. Analysis of secondary spermatocytes (metaphase II configurations) revealed normal disjunction and balanced segregation of the elements of the heteromorphic bivalent. These observations indicate that the inversion heteromorphism does not lead to the production of unbalanced gametes.

Animals↗
PubMed1993

A balanced autosomal reciprocal translocation in an azoospermic bull.

This appears to be the first reported case of a bull with a balanced autosomal reciprocal translocation associated with azoospermia. The analysis includes somatic chromosome banding, conventional meiotic analysis, and electron microscopy of synaptonemal complexes (SCs). The karyotype of the bull was found to be 60,XY,rcp(8;13)(q11;q24). Electron microscopy of SCs in microspread pachytene spermatocytes revealed a high incidence of terminal asynapsis of the smallest arm of the quadrivalent. Most quadrivalents with such asynapsis and few with nonhomologous synapsis showed associations with the XY sex bivalent, leading to complete meiotic arrest at late pachynema. Except for one diakinetic cell, no diplotene and subsequent stages were encountered in air-dried meiotic preparations. The presence of degenerating primary spermatocytes in SC preparations, as well as in testicular sections, and the absence of spermatozoa in ejaculates confirm the chromosomally derived male sterility of the bull. X-chromosome reactivation, evidenced by the cytomorphological reversal of associated sex bivalents, appeared to be the initial step in the degeneration of spermatocytes. Consequently, the formation of a separate, fully developed XY body, which was previously demonstrated on the periphery of spermatocyte nuclei in fertile bulls, could not be attained in this case. Extensive end-to-end association of autosomal bivalents in meiotically arrested, as well as degenerating, spermatocytes was a consistent and unique observation of this study. Such associations may lead to enhanced reactivation of the X chromosome.

Animals↗
PubMed1981

Nonantibody binding of serum proteins to 5S anti-Rh fragments produced by chymotrypsin.

Chymotrypsin hydrolysis of the IgG anti-Rh antibodies Ri results in both bivalent and univalent antibody fragments. The bivalent fragments coated on Rh-positive erythrocytes are agglutinable by albumin and other serum proteins in 3% polyethylene glycol. The bivalent structure of the 5S fragment is essential for expression of this site since 5S fragments produced by trypsin and pepsin are also agglutinable, while univalent fragments produced by papain and subtilisin are not. The agglutination by albumin of the 5S fragments is not caused by residual enzyme. The reaction appears to be irreversible in that once albumin has reacted with the 5S fragment, either in the fluid phase or at the cell surface, fresh addition of albumin and PEG will not result in agglutination. The nonantibody reaction of albumin and the other serum proteins with these 5S IgG fragments is believed to be caused by hydrophobic bonding involving the intrachain disulfide in the 5S fragment and hydrophobic areas of other proteins.

Antibodies↗
PubMed1984

DNA-anti-DNA immune complexes. Antibody protection of a discrete DNA fragment from DNase digestion in vitro.

We examined the ability of DNase I to digest DNA that was contained with DNA-anti-DNA immune complexes. IgG isolated from the sera of 20 patients with systemic lupus erythematosus (SLE) and containing antibodies to DNA was incubated with double-stranded DNA to form immune complexes. Excess DNase was added, and digestion of DNA was monitored by the conversion of DNA to TCA soluble products. IgG from 8 of the 20 SLE patients protected DNA from degradation by DNase in direct proportion to the amount of DNA bound to IgG as measured in the Farr binding assay. Using IgG from these sera, we showed that the DNA protected from degradation remained bound to IgG during digestion and was 35-45 base pairs in size. The size of this fragment is the same as that which has been proposed to be the minimal size necessary for monogamous bivalent binding of IgG to DNA. We therefore compared the ability of F(ab')2 and Fab' to protect DNA from DNase digestion and demonstrated that the bivalent F(ab')2 fragments were protective, but that the univalent Fab' fragments were not. These results suggest that some antibodies to DNA that bind to DNA via monogamous bivalent binding can protect a 35-45-base pair DNA fragment from DNase digestion. The implications of this finding are discussed with regard to the in vivo behavior and potential pathogenicity of small DNA-anti-DNA immune complexes.

Animals↗
PubMed2002

Thrombogenicity of beta 2-glycoprotein I-dependent antiphospholipid antibodies in a photochemically induced thrombosis model in the hamster.

We previously showed that beta(2)-glycoprotein I (beta(2)GPI)-dependent lupus anticoagulants (LAs) form bivalent antigen-antibody complexes with high affinity for phospholipids; these complexes are responsible for their in vitro anticoagulant effect. We now studied the role of these bivalent complexes in arterial thrombosis in the hamster. Three monoclonal antibodies (mAbs) raised against human beta(2)GPI were selected on the basis of their cross-reactivity with hamster beta(2)GPI. Two of these, one with LA activity, 5H2, and one with only anticardiolipin properties, 11E8, were infused at 0 to 10 mg/kg prior to photochemically induced vessel damage. 5H2 promoted thrombus formation dose dependently, raising the thrombus size from 6.0 arbitrary units (AU) in controls (n = 9) to 65.0 AU in the high-dose group (10 mg/kg, n = 6, P =.007). The LA(-) mAb 11E8 and mAb 27A8, reactive with human beta(2)GPI exclusively, did not significantly promote thrombus formation. In a second set of experiments, intact mAb 5H2 was compared to its fragments. Intact mAb 5H2 at 3.3 mg/kg and the equimolar dose of F(ab')(2) fragments (2.2 mg/kg) promoted thrombus formation equally well (55.8 AU, n = 8 and 62.5 AU, n = 7, respectively); mAb 5H2-derived Fab' fragments were inactive. Immunohistochemical analysis showed platelet-rich thrombi, with 5H2 or its F(ab')(2) fragments mainly bound to individual platelets. Our results indicate that bivalent immune complex formation plays an important role in the genesis of arterial thrombosis by certain antiphospholipid antibodies. Cellular activation via the Fc portion of these immune complexes, however, is not essential, because F(ab')(2) fragments of 5H2 still promote thrombus formation.

Adenosine Diphosphate↗
PubMed2006

A monoclonal antibody against CD148, a receptor-like tyrosine phosphatase, inhibits endothelial-cell growth and angiogenesis.

Angiogenesis contributes to a wide range of neoplastic, ischemic, and inflammatory disorders. Definition of the intrinsic molecular controls in angiogenic vessel growth promises novel therapeutic approaches for angiogenesis-related diseases. CD148 (also named DEP-1/PTP eta) is a receptor-like protein tyrosine phosphatase that is abundantly expressed in vascular endothelial cells. To explore a role of CD148 in endothelial vessel formation, we generated a monoclonal antibody, Ab1, against the ectodomain sequence of CD148 and examined its effects on endothelial-cell growth and vessel formation. Here we report that a bivalent, but not a monovalent, form of the Ab1 antibody inhibits endothelial-cell growth and blocks angiogenesis in mouse cornea in vivo. We further demonstrate that (1) bivalent Ab1 arrests cell-cycle progression of CD148-transfected CHO cells at G(0)/G(1) phase, (2) coexpression of catalytically inactive CD148 mutants attenuates the Ab1-cell growth inhibition, and (3) bivalent Ab1 suppresses phosphorylation of ERK1/2 kinases and Met tyrosine kinase as activated CD148 does, with an increase in CD148-associated tyrosine phosphatase activity. Taken together, these findings demonstrate that Ab1-induced ectodomain oligomerization arrests endothelial-cell growth through catalytic activity of the CD148 cytoplasmic domain. The present study defines CD148 as a valuable molecular target for antiangiogenesis therapy.

Animals↗
PubMed2004

Ubiquitin ligase Rad18Sc localizes to the XY body and to other chromosomal regions that are unpaired and transcriptionally silenced during male meiotic prophase.

In replicative damage bypass (RDB) in yeast, the ubiquitin-conjugating enzyme RAD6 interacts with the ubiquitin ligase RAD18. In the mouse, these enzymes are represented by two homologs of RAD6, HR6a and HR6b, and one homolog of RAD18, Rad18Sc. Expression of these genes and the encoded proteins is ubiquitous, but there is relatively high expression in the testis. We have studied the subcellular localization by immunostaining Rad18Sc and other RDB proteins in mouse primary spermatocytes passing through meiotic prophase in spermatogenesis. The highest Rad18Sc protein level is found at pachytene and diplotene, and the protein localizes mainly to the XY body, a subnuclear region that contains the transcriptionally inactivated X and Y chromosomes. In spermatocytes that carry translocations for chromosomes 1 and 13, Rad18Sc protein concentrates on translocation bivalents that are not fully synapsed. The partly synapsed bivalents are often localized in the vicinity of the XY body, and show a very low level of RNA polymerase II, indicating that the chromatin is in a silent configuration similar to transcriptional silencing of the XY body. Thus, Rad18Sc localizes to unsynapsed and silenced chromosome segments during the male meiotic prophase. All known functions of RAD18 in yeast are related to RDB. However, in contrast to Rad18Sc, expression of UBC13 and poleta, known to be involved in subsequent steps of RDB, appears to be diminished in the XY body and regions containing the unpaired translocation bivalents. Taken together, these observations suggest that the observed subnuclear localization of Rad18Sc may involve a function outside the context of RDB. This function is probably related to a mechanism that signals the presence of unsynapsed chromosomal regions and subsequently leads to transcriptional silencing of these regions during male meiotic prophase.

Animals↗