cDNA sequence and chromosome localization of pig α1,3 galactosyltransferase

Human serum contains natural antibodies (NAb), which can bind to endothelial cell surface antigens of other mammals. This is believed to be the major initiating event in the process of hyperacute rejection of pig to primate xenografts. Recent work has implicated galoctosyl 1,3 galactosyl 1,4 N-acetyl-glucosaminyl carbohydrate epitopes, on the surface of pig endothelial cells as a major target of human natural antibodies. This epitope is made by a specific galactosyltransferase (1,3 GT) present in pigs but not in higher primates. We have now cloned and sequenced a full-length pig 1,3 GT cDNA. The predicted 371 amino acid protein sequence shares 85% and 76% identity with previously characterized cattle and mouse 1,3 GT protein sequences, respectively. By using fluorescence and isotopic in situ hybridization, the GGTA1 gene was mapped to the region q2.10–q2.11 of pig chromosome 1, providing further evidence of homology between the subterminal region of pig chromosome 1q and human chromosome 9q, which harbors the locus encoding the AB0 blood group system, as well as a human pseudogene homologous to the pig GGTA1 gene.The nucleotide sequence data reported in this paper have been submitted to the GenBank nucleotide sequence database and have been assigned the accession number L36152     

A mitotically stable marker chromosome negative for whole chromosome libraries, centromere probes and chromosome specific telomere regions: a novel class of supernumerary marker chromosome?

A two year-old child presented with mild developmental delay. On karyotype analysis, a supernumerary small marker chromosome (SMC) was found in all cells examined. This SMC was approximately the size of an isochromosome 18p, being symmetrical with a central constriction. C-banding and silver staining were negative and FISH with all chromosome-specific paints, centromere probes and telomere probes showed no hybridization to the SMC; telomere repeat sequences were however present on both arms. Comparative genomic hybridization showed no amplification of any chromosome region. Flow sorting of the SMC and reverse painting onto normal metaphase spreads showed no hybridization to any chromosome, whereas reverse painting onto the patient's own metaphases showed hybridization to the SMC only. This SMC may thus represent either a complex amplicon of different genomic regions, or a multifold amplification of a very small region, with a neocentromere comprising an active kinetochore but no alphoid DNA. Prognostic implications for the proband were difficult to assess due to the absence of reports of similar marker chromosomes in the literature.     

Localization of CTβ and C K on mouse chromosome 6

In the mouse three lymphocyte gene families have been positioned on the proximal region of chromosome 6. Originally the immunoglobulin kappa light chain (Igk) and the thymocyte surface antigens Lyt-2 and Lyt-3 were assigned to chromosome 6, and recently the beta chain of the T-cell receptor for antigen was positioned proximal to Igk. Molecular clones which recognize the constant (C region of the beta chain of the T-cell receptor for antigen (C_T ) and the constant region of the immunoglobulin kappa (C _k ) chain were used to determine recombination frequencies with respect to the morphological marker hypodactyly (Hd). SJL/JLw mice were mated with C.B6.C3-Hd/+ mice, and the progeny expressing the Hd phenotype were mated with SJL/JLw mice. Backcross progeny which expressed the Hd phenotype were nephrectomized, and kidney DNA was examined by Southern hybridization for the polymorphic restriction endonuclease fragment (REF) patterns of the parental mice. Of the 88 progeny tested in this three-point cross, 3 C_T and 4 C _K homozygote REF patterns were detected. These homozygotes were mutually exclusive. This implies the following gene order: centromere-C _T -Hd-Igk and C _T1 would be 7.95±2.88 centimorgans from C _K .     

Characterisation of interstitial duplications and triplications of chromosome 15q11–q13

Chromosome 15 is frequently involved in the formation of structural rearrangements. We report the molecular characterisation of 16 independent interstitial duplications, including those of one individual who carried a duplication on both of her chromosomes 15, and three interstitial triplications of the Prader-Willi/Angelman syndrome critical region (PWACR). In all probands except one, the rearrangement was maternal in origin. In one family, the duplication was paternal in origin, yet appeared to segregate in a sibship of three with an abnormal phenotype that included developmental delay and a behavioural disorder. Ten duplications were familial, five de novo and one unknown. All 16 duplications, including two not visible by routine G-banding, were of an almost uniform size and shared the common deletion breakpoints of Prader-Willi syndrome and Angelman syndrome. Like deletions, the formation of duplications can occur in both male and female meiosis and involve both inter- and intrachromosomal events. This implies that at least some deletions and duplications are the reciprocal products of each other. We observed no instances of meiotic instability in the transmission of a duplication, although recombination within the PWACR occurred in two members of the same family between the normal and the duplicated chromosome 15 homologues. All three triplications arose de novo and included alleles from both maternal chromosomes 15. Triplication breakpoints were more variable and extended distally beyond the PWACR. The molecular characteristics of duplications and triplications suggest that they are formed by different mechanisms.     

Genetic analysis of transgenome structure and size of chromosome—mediated gene transfer lines

The TK-selected chromosome-mediate gene transfer lines were analysed using DNA dot blot method G-11 banding and in situ hybridization.The results showed that CMGT can provide a wide variety of intermediate size of the transgenome from greater than 80,000kb to less than 2,000kb,Some of transfectants are intergrated into mouse chromosome which can be detected by G-11 banding and in situ hybridization.     

Mapping through somatic cell hybrids and cDNA probes of protein C to chromosome 2, factor X to chromosome 13, and α1-acid glycoprotein to chromosome 9

Summary The previously unassigned gene coding for the anticoagulatory protein C has been mapped on chromosome 2 using a cDNA probe and genomic blots from a human-hamster somatic cell hybrid panel. The assignments of the genes coding for the coagulation factor X to chromosome 13, and for 1-acid glycoprotein to chromosome 9 have been confirmed using a similar direct approach.     

Y chromosome and male infertility: what is a normal Y chromosome?

The human Y chromosome contains a number of genes and gene families that are essential for germ cell development and maintenance. Many of these genes are located in highly repetitive elements that are subject to rearrangements. Deletion of azoospermia factor (AZF) regions AZFa, AZFb, and AZFc are found in approximately 10-15% of men with severe forms of spermatogenic failure. Several partial AZFc deletions have been described. One of these, which removes around half of all the genes within the AZFc region, appears to be present as an inconsequential polymorphism in populations of northern Eurasia. A second deletion, termed gr/gr, also results in the absence of several AZFc genes and it may be a genetic risk factor for spermatogenic failure. However, the link between these partial deletions and fertility is unclear. The gr/gr deletion is not a single deletion but a combination of deletions that vary in size and complexity and result in the absence of different genes. There are also regional or ethnic differences in the frequency of gr/gr deletions. In some Y-chromosome lineages, these deletions appear to be fixed and may have little influence on spermatogenesis. Most of these data (gene content and Y chromosome structure) have been deduced from the reference Y chromosome sequence deposited in NCBI. However, recently there have been attempts to define these types of structural rearrangements in the general population. These have highlighted the considerable degree of structural diversity that exist. Trying to correlate these changes with the phenotypic variability is a major challenge and it is likely that there will not be a single reference (or normal) Y chromosome sequence but many.     

Assignment of the gene for carboxypeptidase A to human chromosome 7q22→qter and to mouse chromosome 6

Summary A rat cDNA probe for preprocarboxypeptidase A was used to follow the segregation of the human gene for carboxypeptidase A (CPA) in 49 human x mouse somatic cell hybrids using Southern filter hybridization techniques. CPA was assigned to human chromosome 7q22qter. Similarly, the probe was used to follow the segregation of the mouse gene for carboxypeptidase A (Cpa) in 19 mouse x Chinese hamster somatic cell hybrids. Cpa was assigned to mouse chromosome 6. The gene for carboxypeptidase A forms part of a syntenic group that is conserved in man and mouse.Preliminary chromosomal assignments of carboxypeptidase A in man and mouse have been made in abstract (Honey et al. 1983a, b)     

Localization of anti-topoisomerase IIα antibodies under normal conditions and after artificial chromosome condensation and decondensation

By means of immunofluorescence method, localization of DNA-topoisomerase IIα (Topo IIα) in interphase nuclei and chromosomes at different stages of mitosis was studied in situ under normal conditions and after treatment with condensing and decondensing solutions. In non-isolated mitotic M-HeLa cell chromosomes, Topo IIα was uniformly distributed along chromatids after fixation and permeabilization in situ. After treatment of cells with decondensing solutions (10 mM Tris; 0.1 mM CaCl₂ in 10 mM Tris; 0.3 mM CaCl₂ in 10 mM Tris; 15% DMEM; 75 mM KCl), Topo IIα was evenly distributed along chromatids in prophase, prometaphase and metaphase; its concentration was the highest in the pericentromere region. After treatment of cells with condensing solutions containing 0.7 mM, 1 mM, 2 mM or 3 mM CaCl₂ in 10 mM Tris, Topo IIα was not detected in prophase, metaphase and anaphase. However, in late telophase anti-Topo IIα antibodies were found in reforming nuclei under identical conditions. After sequential treatment with condensing and decondensing solutions, the distribution patterns of Topo IIα in chromosomes were the same as after treatment with only decondensing solutions. In anaphase and telophase, Topo IIα was evenly distributed along chromatids, while in prophase, prometaphase and metaphase it was predominantly localized in the pericentromere region. After the treatment of cells with condensing solutions chromosome staining was not observed, apparently due to “masking” of binding sites for anti-Topo IIα antibodies. Homogenous distribution of Topo IIα along chromatids in non-isolated chromosomes was preserved after the treatment of cells with hypotonic solutions; however, under these conditions Topo IIα concentration was higher in centromeres.     

Isolation and characterization of an α-satellite repeated sequence from human chromosome 22

We constructed a library in IL47.1 with DNA isolated from flow-sorted human chromosome 22. Over 50% of the recombinants contained the same highly repetitive sequence. When this sequence was used to probe Southern blots of EcoRI-digested genomic DNA, a ladder of bands with increments of about 170 bp was observed. This sequence comigrates with satellite III in Ag⁺/Cs₂SO₄ gradients and may account for at least part of the 170 bp Hae III ladder seen in isolated satellite III DNA. Partial sequence analysis revealed homology to the 171 bp monomeric repeat unit of -R1-DNA and the X specific -satellite consensus sequence. After low stringency in situ hybridization, silver grains were found over the centromeres of a number of chromosomes. Under high stringency conditions, however, the labeling was concentrated over the centromeric region of chromosome 22. This localization was confirmed using DNA from a panel of human/hamster cell lines which showed that the homologous 2.1 and 2.8 kb EcoR1 restriction fragments were chromosome 22 specific. These clones therefore contain chromosome 22 derived -satellite sequences analogous to other chromosome-specific satellite sequences described previously.     

Age- and tissue-specific variation of X chromosome inactivation ratios in normal women

The incidence of skewed X-inactivation in normal women is controversial, with up to 10-fold differences being reported by different authors. In order to clarify this issue, we have conducted a survey of the X-inactivation patterns in 270 informative females from various age groups, using the androgen receptor gene/polymerase chain reaction assay. Results obtained by using DNA extracted from blood samples show that the incidence of severe skewing (defined here as ratios > or = 90:10) is relatively common and increases with age (P<0.05), occurring in 7% of women under 25 years of age, and 16% of women over 60. In order to study tissue-specific patterns of X-inactivation, samples of both buccal and urinary epithelia were also obtained from 88 of the females studied. Although there was a significant association of the X-inactivation ratios between each tissue in most individuals, wide variations were apparent in some cases, making accurate extrapolations between tissues impossible. The degree of correlation between each tissue also fell markedly with age. Our data are consistent with the hypothesis that the major factors in the aetiology of skewed X-inactivation are secondary selection processes.     

＂Micro-deletions＂ of the human Y chromosome and their relationship with male infertility

The Y chromosome evolves from an autochromosome and accumulates male-related genes including sex-determining region of Y-chromosome (SRY) and several spermatogenesis-related genes.The human Y chromosome (60 Mb long) is largely composed of repeti-tive sequences that give it a heterochromatic appearance,and it consists of pseudoautosomal,euchromatic,and heterochromatic regions.Located on the two extremities of the Y chromosome,pseudoautosomal regions 1 and 2 (PAR1 and PAR2,2.6 Mb and 320 bp long,re-spectively) are homologs with the termini of the X chromosome.The euchromatic region and some of the repeat-rich heterochromatic parts of the Y chromosome are called "male-specific Y" (MSY),which occupy more than 95% of the whole Y chromosome.After evolu-tion,the Y chromosome becomes the smallest in size with the least number of genes but with the most number of copies of genes that are mostly spermatogenesis-related.The Y chromosome is characterized by highly repetitive sequences (including direct repeats,inverted repeats,and palindromes) and high polymorphism.Several gene rearrangements on the Y chromosome occur during evolution owing to its specific gene structure.The consequences of such rearrangements are not only loss but also gain of specific genes.One hundred and fifty three haplotypes have been discovered in the human Y chromosome.The structure of the Y chromosome in the GenBank belongs to haplotype R1.There are 220 genes (104 coding genes,111 pseudogenes,and 5 other uncategorized genes) according to the most recent count.The 104 coding genes encode a total of about 48 proteins/protein families (including putative proteins/protein families).Among them,16 gene products have been discovered in the azoospermia factor region (AZF) and are related to spermatogenesis.It has been dis-covered that one subset of gene rearrangements on the Y chromosome,"micro-deletions",is a major cause of male infertility in some populations.However,controversies exist about different Y chromosome haplotypes.Six AZFs of the Y chromosome have been discov-ered including AZFa,AZFb,AZFc,and their combinations AZFbc,AZFabc,and partial AZFc called AZFc/gr/gr.Different deletions in AZF lead to different content spermatogenesis loss from teratozoospermia to infertility in different populations depending on their Y hap-lotypes.This article describes the structure of the human Y chromosome and investigates the causes of micro-deletions and their relation-ship with male infertility from the view of chromosome evolution.After analysis of the relationship between AZFc and male infertility,we concluded that spermatogenesis is controlled by a network of genes,which may locate on the Y chromosome,the autochromosomes,or even on the X chromosome.Further investigation of the molecular mechanisms underlying male fertility/infertifity will facilitate our knowledge of functional genomics.     

Alteration of chromosome numbers by generation of minichromosomes -- is there a lower limit of chromosome size for stable segregation?

Practical applications of minichromosomes, generated by de novo composition or by truncation of natural chromosomes, rely on stable transmission of these chromosomes. Functional centromeres, telomeres and replication origins are recognized as prerequisites for minichromosome stability. However, it is not yet clear whether, and if yes, to what degree the chromatin content has a qualitative or quantitative impact on stable chromosome transmission. A small translocation chromosome, which arose after X-irradiation of a reconstructed field bean karyotype, comprised approximately 5% of the haploid metaphase complement and was found to consist of three pieces of duplicated chromatin and a wild-type centromere. This chromosome was stably transmitted through all meristematic and pollen grain mitoses but was frequently lost during meiosis (66% loss in hemizygous and 33% in homozygous condition). This minichromosome was only a little smaller than stably segregating translocation chromosomes (comprising approximately 6% of the genome) of a euploid field bean karyotype. The duplications specific for this minichromosome did not influence meiotic segregation when associated with non-duplicated chromatin of other chromosomes. In comparison with minichromosomes of other species, the possibility of a lower size limit for a stable chromosome transmission must therefore be considered which might be based, for instance, on insufficient lateral support of centromeres or on insufficient bivalent stability due to the incapability of chiasma formation.     

Involvement of an “all-or-none event” in the triggering of chromosome segregation and of S phase in procaryotic and eucaryotic cells

A structural model of the triggering of mitosis is described. It is proposed that the number of structural effectors varies discontinuously both just before mitosis through an “all-or-none event” and during the mitotic process itself. The effector persists from mitosis to mitosis, but is not active in G2 due to the polymerization of soluble monomers. The “all-or-none event” which triggers mitosis is postulated to involve the doubling of the number of the structural effectors which are then temporarily in an active state, thus initiating mitosis. The subsequent segregation during mitosis of the active structural effectors to the two dividing nuclei allows the initiation of S phase as soon as the chromosomal replicative machinery is ready.     

Characteristics of βA chromosome haplotypes in Japanese

DNA polymorphism patterns linked to the ^A-globin gene were analyzed in healthy Japanese using four different restriction endonucleases. The chromosomes with the ^A-globin gene were mapped through an evaluation of the presence of seven different restriction sites (HincII 5 to ; HindIII in ^G and ^A; HincII in, and 3 to, ₁; AvaII in ; Bam-HI 3 to ). Among 36 chromosomes analyzed, 20 chromosomes had a haplotype of [+–––––+]. Among 55 individuals examined, 7 possessed a homozygous haplotye of [+–––––+]. All Japanese with the ^AT-globin gene had a subhaplotype of [–++–+] 5 to the -globin gene. Their major haplotypes were [–++–+–+] and [–++–++–]. It was expected that the presence of the ^AT-globin gene in Japanese may be deduced from subhaplotypes 5 to the -globin gene.     

Direct assignment of orosomucoid to human chromosome 9 and α2HS-glycoprotein to chromosome 3 using human fetal liver x rat hepatoma hybrids

Summary The production of plasma proteins has been monitored in somatic cell hybrids between a rat hepatoma cell line (7777) and human fetal liver cells. Production of 14 plasma proteins was assayed in concentrated serum-free culture supernatants by electroimmunoassay. 2HS-glycoprotein (AHSG) was produced by 10 of 19 hybrids; concordancy for presence or absence of protein production was 100% for human chromosome 3. Orosomucoid (ORM) was produced in 8 of 19 hybrids, with a concordancy for presence or absence of protein of 94.7% with human chromosome 9. The chromosome location for genes for these two proteins, previously assigned by linkage studies, is confirmed by direct assignment. These studies have also suggested possible chromosomal assignments for loci for ₁ and C1 esterase inhibitor. Other genes for proteins which could not be assigned to specific chromosomes using these hybrids were: complement C3, ceruloplasmin, hemopexin, inter--trypsin inhibitor, prealbumin, retinol-binding protein, transferrin and apolipoproteins CII, B, and sinking-pre-beta [Lp(a)].     

Population cytology of structural and numerical chromosome variants in theAloïneae (Liliaceae)

The distribution of interchange heterozygotes is analysed in one population of each of three species in the tribeAloïneae (Liliaceae). While one population is shown to be clonal the other two indicate that natural selection eliminates interchange homozygotes and maintains the chromosomal uniformity of the tribe. Analysis of a fourth population containing aneuploids and chromosome deletions shows that mutant plants are capable of interbreeding, but that spread of the aberrations is limited.