The terminal DNA structure of mammalian chromosomes

In virtually all eukaryotic organisms, telomeric DNA is composed of a variable number of short direct repeats. While the primary sequence of telomeric repeats has been determined for a great variety of species, the actual physical DNA structure at the ends of a bona fide metazoan chromosome with a centromere is unknown. It is shown here that an overhang of the strand forming the 3' ends of the chromosomes, the G-rich strand, is found at mammalian chromosome ends. Moreover, on at least some telomeres, the overhangs are > or = 45 bases long. Such surprisingly long overhangs were present on chromosomes derived from fully transformed tissue culture cells and normal G0-arrested peripheral leukocytes. Thus, irrespective of whether the cells were actively dividing or arrested, a very similar terminal DNA arrangement was found. These data suggest that the ends of mammalian and possibly all vertebrate chromosomes consist of an overhang of the G-rich strand and that these overhangs may be considerably larger than previously anticipated.     

Feasibility of analysis of macroevolutionary rearrangements of mammalian chromosomes]

In the Institute of Cytology and Genetics, Siberian Division, Russian Academy of Sciences, detailed studies on chromosome sets of humans and domestic animals were initiated and supported by D.I. Belyaev and started by S.I. Radzhabli. They believed that analysis of differentially stained chromosomes and mapping of the genomes of main commercial species provide for a better understanding of the processes that occurred during their evolution and domestication. Several new approaches to studying macroevolutionary karyotypic rearrangements associated with divergence of remote taxa, such as primates and paridigitate ungulates, are discussed.     

A proposed path by which genes common to mammalian X and Y chromosomes evolve to become X inactivated

Mammalian X and Y chromosomes evolved from an autosomal pair; the X retained and the Y gradually lost most ancestral genes. In females, one X chromosome is silenced by X inactivation, a process that is often assumed to have evolved on a broadly regional or chromosomal basis. Here we propose that genes or clusters common to both the X and Y chromosomes (X-Y genes) evolved independently along a multistep path, eventually acquiring dosage compensation on the X chromosome. Three genes studied here, and other extant genes, appear to be intermediates. ZFX, RPS4X and SMCX were monitored for X inactivation in diverse species by assaying CpG-island methylation, which mirrors X inactivation in many eutherians. ZFX evidently escaped X inactivation in proto-eutherians, which also possessed a very similar Y-linked gene; both characteristics were retained in most extant orders, but not in myomorph rodents. For RPS4X, escape from X inactivation seems unique to primates. SMCX escapes inactivation in primates and myomorphs but not in several other lineages. Thus, X inactivation can evolve independently for each of these genes. We propose that it is an adaptation to the decay of a homologous, Y-linked gene.     

Centromeres of human chromosomes

The centromere, recognized cytologically as the primary constriction, is essential for chromosomal attachment to the spindle and for proper segregation of mitotic and meiotic chromosomes. Considerable progress has been made in identifying both DNA and protein components of the centromere and kinetochore complex in mammalian chromosomes, including definition of specific motor proteins with demonstrable functions in chromosome movement. Searches for possible environmental influences on chromosome disjunction might logically be based on known components of the segregation apparatus, both intrinsic and extrinsic to the chromosomes themselves. This article reviews available information on both DNA and protein components of the centromere of mammalian, particularly human, chromosomes and summarizes our current understanding of their role(s) in facilitating normal chromosome behavior in mitosis and meiosis.     

M33, a mammalian homologue of Drosophila Polycomb localises to euchromatin within interphase nuclei but is enriched within the centromeric heterochromatin of metaphase chromosomes

Both experimental and clinical data have shown that coagulation disorders are common in patients with cancer although clinical symptoms occur rarely. A prethrombotic state is probably involved in the mechanism of metastatic spread. Anticoagulant treatment, with either warfarin or heparin, has been shown to have a positive influence in small cell lung cancer. The purpose of this study was to evaluate the prethrombotic state as a possible marker of the outcome of lung cancer. Pretreatment prothrombin time (PT), partial thromboplastin time (PTT), antithrombin III (AT-III), platelet blood count (P), fibrinogen (F) and D-dimer (DD) were prospectively recorded in a series of 286 consecutive patients with a new primary lung cancer. Other recorded variables (32 in all) consisted of a set of anthropometric, clinical, physical, laboratory, radiological and pathological data. All patients were carefully followed up, and their subsequent clinical course recorded. Spearman rank correlation tests between coagulation factors were weakly significant, or more often non-significant. The best correlation index was that between PT and PTT (ra = -0.25). Univariate analyses of survival showed that a prolonged value of PT (P = 0.00167) and higher values of F (P = 0.00143) and DD (P = 0.0005) were associated with a poor prognosis. A few, weak relationships between well-known prognostic variables and coagulation abnormalities were also found. Because of the weakness of this correlation pattern, coagulation factors emerged in all the Cox's regression analyses as important predictors of survival, regardless of the number and type of cofactors used. A prethrombotic state (depicted by a prolongation of PT and increase of DD) is confirmed in this study as an aggravating condition in lung cancer. Studies attempting to reverse possible haemostatic abnormalities with the use of anticoagulants are justified by the present data.     

Visual classification of banded human chromosomes. I. Karyotyping compared with classification of isolated chromosomes

Visual karyotyping and visual classification of isolated chromosomes was carried out by seven investigators on 22 trypsin banded metaphases of average quality. The karyotyping experiment resulted in an average error rate of 0-1% (zero-0-4%) and the classification of isolated chromosomes resulted in an error rate of 3% (2-5%). The B and F group chromosomes were found to be most difficult to classify when isolated, while no errors were made of the no. 1 and the X chromosome. Large differences were seen in the resulting error pattern for the individual investigators both with regard to their total error rate and also the chromosome types which they most frequently misclassified. Based upon these error patterns it is suggested that more than 95% of the chromosomes in an average quality material contain features upon which a reliable visual classification can be made. Thus there may be a potential possibility that these chromosomes may be classified by computer on the basis of these features. The fact that visual karyotyping is much more reliable than visual classification of isolated chromosomes indicated that computer classification of chromosomes should include programming capable of making appropriate comparison between the chromosomes in the metaphase and at the same time take into account the expected presence of 23 chromosome pairs for normal cells. This would simulate the human performance of visual karyotyping and make a classification possible of at least some of the remaining 5% difficult chromosomes.     

Alphoid repetitive DNA in human chromosomes

The thesis describes the first extensive DNA sequence analysis that demonstrated that the tandemly repeated alphoid DNA in the centromere of the human chromosomes consists of distinct subfamilies and in a number equal to or exceeding the number of chromosomes. The expected presence of only one or a few distinct subfamily on individual chromosomes was supported by the characterization of an extremely well-defined subfamily specific for chromosome 7 and represented in the original collection of subfamilies. The pattern of chromosome-specificity breaks down among the acrocentric chromosomes where chromosomes 13 and 21 were found to share one and chromosomes 14 and 22 to share another specific subfamily. By in situ hybridization these subfamilies were shown not to be shared by other chromosomes. The remarkable pairwise pattern of sequence homogenization was present also in the chimpanzee genome raising the question of its biological role. However, the subfamilies on these human and chimpanzee chromosomes are not orthologous but were shown to originate from two evolutionarily different repeat families. It follows that dramatic sequence evolution has occurred in one or both species during or after separation. The sequence evolution might even occur at a higher rate in humans. This possibility was studied in orthologous alphoid sequences on the X chromosome of humans and the great apes. The analysis supports the general view that our closest relative is the chimpanzee and indicates that the rate of recombination is increased in the human repeat DNA. A "molecular clock" running faster in this DNA may have evolutionary implications. Finally, the usefulness of alphoid subfamilies as chromosome-specific markers is illustrated in a cytogenetic dissection of the centromeric region of Robertsonian translocations. The breakpoints were located to satellite III DNA leaving these chromosomes dicentric. The order of the different tandem DNAs on the p-arm of the acrocentric chromosomes could also be established.     

How Y chromosomes become genetically inert

We have investigated the mechanistic aspects of inactivation of the major larval cuticle protein genes (Lcp1-4) in Drosophila miranda during Y chromosome evolution. The Lcp genes are located on the X2 and neo-Y chromosomes in D. miranda but are autosomally inherited in all other Drosophila species investigated so far. In the neo-Y chromosome all four Lcp loci are embedded within a dense cluster of transposable elements. The X2 Lcp1-4 loci are expressed, while the Y chromosomal Lcp3 locus shows only reduced activity and the Lcp1, Lcp2, and Lcp4 are completely inactive. Our results suggest that Lcp1 and Lcp3 loci on the degenerating Y chromosome of D. miranda are silenced by neighboring transposable elements. These observations support our assumption that the first step in Y chromosome degeneration is the successive silencing of Y chromosomal loci caused by trapping and accumulation of transposons.     

Sex chromosomes: evolving dosage compensation

Dosage compensation of some X-linked genes varies among mammals. Inactivation of an X-linked copy of a gene in females appears to correlate with lack of an active homologue on the Y chromosome, implying that dosage compensation evolves in response to the loss of function of genes on the Y.     

Histones and G banding of chromosomes

Polylysine, polyarginine, and histones H1, H2A, H2B, and H3 inhibit Giemsa staining and chromosome banding by binding to DNA and preventing side stacking of the positively charged thiazine dyes to the negatively charged phosphate groups on DNA. This is a nonspecific effect and does not of itself provide evidence for a role of histones in G banding. The question of whether histones are involved in chromosome banding is reviewed.     

Hypomethylation of human sperm pronuclear chromosomes

Using the methylase SssI enzyme, we have analyzed the degree of in situ methylation of human sperm pronuclear chromosomes obtained by fertilizing hamster oocytes with human sperm. Untreated (control) sperm chromosome complements showed a higher degree of in situ methylation, compared to sperm complements previously treated with 5-azadeoxycytidine or lymphocyte chromosomes. This indicates that human sperm pronuclear chromosomes have a lower degree of genomic methylation compared to that of other somatic cells. The similarity in the degree of in situ methylation of the euchromatic and heterochromatic regions of chromosomes 1, 9, 15, and 16 and the Y chromosome in human sperm does not support the existence of a possible correlation between hypomethylation and heterochromatin decondensation.     

A mammalian telomerase-associated protein

The telomerase ribonucleoprotein catalyzes the addition of new telomeres onto chromosome ends. A gene encoding a mammalian telomerase homolog called TP1 (telomerase-associated protein 1) was identified and cloned. TP1 exhibited extensive amino acid similarity to the Tetrahymena telomerase protein p80 and was shown to interact specifically with mammalian telomerase RNA. Antiserum to TP1 immunoprecipitated telomerase activity from cell extracts, suggesting that TP1 is associated with telomerase in vivo. The identification of TP1 suggests that telomerase-associated proteins are conserved from ciliates to humans.     

Ultrastructural characterization of the sex chromosomes during spermatogenesis of spiders having holocentric chromosomes and a long diffuse stage

An ultrastructural study has been made of spermatogenesis in two species of primitive spiders having holocentric chromosomes (Dysdera crocata, male X0 and Sergestria florentia X1X2O). Analysis of the meiotic prophase shows a scarcity or absence of typical leptotene to pachytene stages. Only in D. crocata have synaptonemal complex (SC) remnants been seen, and these occurred in nuclei with an extreme chromatin decondensation. In both species typical early prophase stages have been replaced by nuclei lacking SC and with their chromatin almost completely decondensed, constituting a long and well-defined diffuse stage. Only nucleoli and the condensed sex chromosomes can be identified. - In S. florentina paired non-homologous sex chromosomes lack a junction lamina and thus clearly differ from the sex chromosomes of more evolved spiders with an X1X20 male sex determination mechanism. In the same species, sex chromosomes can be recognized during metaphase I due to their special structural details, while in D. crocata the X chromosome is not distinguishable from the autosomes at this stage. - The diffuse stage and particularly the structural characteristics of the sex chromosomes during meiotic prophase are reviewed and discussed in relation to the meiotic process in other arachnid goups.