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PubMed1995

Evidence for a terminal differentiation process in the rat liver.

In rapidly renewing epithelia, such as skin and gut, as well as hemopoietic cells and stromal fibroblasts, the process of progenitor cell maturation, terminal differentiation and senescence from cells of a fetal phenotype is strikingly similar. To examine hepatocellular maturation, we studied embryonic, suckling and young adult rat liver cells with multiparametric fluorescence activated cell sorting (FACS), after exclusion of hemopoietic, endothelial, Kupffer, and nonviable cells. With maturation, cell granularity and autofluorescence exponentially increased from fetal liver to suckling and adult liver as the proportion of S phase cells progressively declined from 33.8% +/- 1.3% to 4.9% +/- 2.8% and 1.1% +/- 0.6% (P < 0.05), respectively. In liver from fetal and suckling rats, all hepatocytes were mononuclear and contained diploid DNA whereas 21.2% +/- 5.9% hepatocytes in adult liver were binucleated. Analysis of nuclear DNA content in adult hepatocytes demonstrated that 53.3% +/- 3.9% of the nuclei were diploid, 43.6% +/- 3.5% tetraploid and 0.5 +/- 0.6% octaploid. However, in the adult liver, small, mononuclear cells were also present with granularity and autofluorescence comparable to fetal hepatoblasts, as well as glucose-6-phosphatase activity, diploid DNA in 89.0% +/- 2.1% of the nuclei, and with increased granularity in culture. Since general features of terminal cellularity differentiation and senescence include cessation of mitotic activity, polyploidy and accumulation of autofluorescent secondary lysosomes, our data suggest that liver cells too undergo a process of terminal differentiation.

Animals↗
PubMed2002

Chromosome mapping of Xenopus tropicalis using the G- and Ag-bands: tandem duplication and polyploidization of larvae heads.

Developmental cytogenetic analyses of Xenopus tropicalis larvae from two origins were performed on stage 27-34 heads treated with colchicine. Standard G-band karyotyping using trypsin and chromosome mapping of 184 bands were examined. Although the main karyotype was 2n = 20, polyploidy (3n = 30 or 4n = 40) and aneuploidy were detected in each individual treated with colchicine, even those treated for only 1 h. The percentage of polyploid karyotypes was 10-20% across the total of measured metaphases. The mean mitotic index was 0.10. Chromosomal breaks and exchanges were detected at the secondary constriction of chromosomes 5 or 6. Ag-band detection showed clearly positive staining at the secondary constriction of chromosome 5, which corresponds to the nucleolar organizer region. Tandem duplication of negative G-bands at the secondary constriction of chromosome 6 and the short arm of chromosome 10 was suggested by this study. X. tropicalis thus provides a good model to study the mechanism and effects of chromosomal abnormalities, gene mapping and tissue specific gene expression in the developmental process.

Animals↗
PubMed2005

Inactivation of the Snf5 tumor suppressor stimulates cell cycle progression and cooperates with p53 loss in oncogenic transformation.

Snf5 (Ini1/Baf47/Smarcb1), a core member of the Swi/Snf chromatin remodeling complex, is a potent tumor suppressor whose mechanism of action is largely unknown. Biallelic loss of Snf5 leads to the onset of aggressive cancers in both humans and mice. We have developed an innovative and widely applicable analytical technique for cross-species validation of cancer models and show that the gene expression profiles of our Snf5 murine models closely resemble those of human Snf5-deficient rhabdoid tumors. We exploit this system to produce what we believe to be the first report documenting the effects on gene expression of inactivating a Swi/Snf subunit in normal mammalian cells and to identify the transcriptional pathways regulated by Snf5. We demonstrate that the tumor suppressor activity of Snf5 depends on its regulation of cell cycle progression; Snf5 inactivation leads to aberrant up-regulation of E2F targets and increased levels of p53 that are accompanied by apoptosis, polyploidy, and growth arrest. Further, conditional mouse models demonstrate that inactivation of p16Ink4a or Rb (retinoblastoma) does not accelerate tumor formation in Snf5 conditional mice, whereas mutation of p53 leads to a dramatic acceleration of tumor formation.

Animals↗
PubMed2002

Mutation of the Drosophila homologue of the Myb protooncogene causes genomic instability.

Vertebrates have three related Myb genes. The c-Myb protooncogene is required for definitive hematopoiesis in mice and when mutated causes leukemias and lymphomas in birds and mammals. The A-Myb gene is required for spermatogenesis and mammary gland proliferation in mice. The ubiquitously expressed B-Myb gene is essential for early embryonic development in mice and is directly regulated by the p16/cyclin D/Rb family/E2F pathway along with many critical S-phase genes. Drosophila has a single Myb gene most closely related to B-Myb. We have isolated two late-larval lethal alleles of Drosophila Myb. Mutant imaginal discs show an increased number of cells arrested in M phase. Mutant mitotic cells display a variety of abnormalities including spindle defects and increased polyploidy and aneuploidy. Remarkably, some mutant cells have an aberrant S- to M-phase transition in which replicating chromosomes undergo premature histone phosphorylation and chromosomal condensation. These results suggest that the absence of Drosophila Myb causes a defect in S phase that may result in M-phase abnormalities. Consistent with a role for Drosophila Myb during S phase, we detected Dm-Myb protein in S-phase nuclei of wild-type mitotic cells as well as endocycling cells, which lack both an M phase and cyclin B expression. Moreover, we found that the Dm-Myb protein is concentrated in regions of S-phase nuclei that are actively undergoing DNA replication. Together these findings imply that Dm-Myb provides an essential nontranscriptional function during chromosomal replication.

Aneuploidy↗
PubMed2002

The hidden duplication past of Arabidopsis thaliana.

Analysis of the genome sequence of Arabidopsis thaliana shows that this genome, like that of many other eukaryotic organisms, has undergone large-scale gene duplications or even duplications of the entire genome. However, the high frequency of gene loss after duplication events reduces colinearity and therefore the chance of finding duplicated regions that, at the extreme, no longer share homologous genes. In this study we show that heavily degenerated block duplications that can no longer be recognized by directly comparing two segments because of differential gene loss, can still be detected through indirect comparison with other segments. When these so-called hidden duplications in Arabidopsis are taken into account, many homologous genomic regions can be found in five to eight copies. This finding strongly implies that Arabidopsis has undergone three, but probably no more, rounds of genome duplications. Therefore, adding such hidden blocks to the duplication landscape of Arabidopsis sheds light on the number of polyploidy events that this model plant genome has undergone in its evolutionary past.

Arabidopsis↗
PubMed2024

Genome evolution of the ancient hexaploid Platanus &#xd7; acerifolia (London planetree).

Whole-genome duplication (WGD; i.e., polyploidy) and chromosomal rearrangement (i.e., genome shuffling) significantly influence genome structure and organization. Many polyploids show extensive genome shuffling relative to their pre-WGD ancestors. No reference genome is currently available for Platanaceae (Proteales), one of the sister groups to the core eudicots. Moreover, Platanus &#xd7; acerifolia (London planetree; Platanaceae) is a widely used street tree. Given the pivotal phylogenetic position of Platanus and its 2-y flowering transition, understanding its flowering-time regulatory mechanism has significant evolutionary implications; however, the impact of Platanus genome evolution on flowering-time genes remains unknown. Here, we assembled a high-quality, chromosome-level reference genome for P. &#xd7; acerifolia using a phylogeny-based subgenome phasing method. Comparative genomic analyses revealed that P. &#xd7; acerifolia (2n = 42) is an ancient hexaploid with three subgenomes resulting from two sequential WGD events; Platanus does not seem to share any WGD with other Proteales or with core eudicots. Each P. &#xd7; acerifolia subgenome is highly similar in structure and content to the reconstructed pre-WGD ancestral eudicot genome without chromosomal rearrangements. The P. &#xd7; acerifolia genome exhibits karyotypic stasis and gene sub-/neo-functionalization and lacks subgenome dominance. The copy number of flowering-time genes in P. &#xd7; acerifolia has undergone an expansion compared to other noncore eudicots, mainly via the WGD events. Sub-/neo-functionalization of duplicated genes provided the genetic basis underlying the unique flowering-time regulation in P. &#xd7; acerifolia. The P. &#xd7; acerifolia reference genome will greatly expand understanding of the evolution of genome organization, genetic diversity, and flowering-time regulation in angiosperms.

Polyploidy↗
PubMed1983

Polyploid nuclei in human artery wall smooth muscle cells.

Although polyploid nuclei have long been known to be present in many adult human tissues, the ploidy of smooth muscle cells in human artery wall has never been determined. We measured DNA content in individual smooth muscle cell nuclei of artery wall specimens by two means: Feulgen microdensitometry and flow microfluorimetry. A significant percentage of nuclei were polyploid; most of these were tetraploid, although higher levels were also found. The frequency of polyploidy varied with age from less than 1% at birth to a mean of 7% in adult aortic, carotid, and iliac vessels. Atherosclerotic plaques had a lower tetraploid content than the underlying media, whereas normal intima was similar to the corresponding media. The increase in frequency of hyperploid smooth muscle cell nuclei correlates with the normal growth, development, and aging of human artery wall. We suggest that the regular existence of a subset of polyploid smooth muscle cells may indicate an important functional role for this phenotype.

Cell Nucleus↗
PubMed1995

Bidirectional interlocus concerted evolution following allopolyploid speciation in cotton (Gossypium).

Polyploidy is a prominent process in plant evolution; yet few data address the question of whether homeologous sequences evolve independently subsequent to polyploidization. We report on ribosomal DNA (rDNA) evolution in five allopolyploid (AD genome) species of cotton (Gossypium) and species representing their diploid progenitors (A genome, D genome). Sequence data from the internal transcribed spacer regions (ITS1 and ITS2) and the 5.8S gene indicate that rDNA arrays are homogeneous, or nearly so, in all diploids and allopolyploids examined. Because these arrays occur at four chromosomal loci in allopolyploid cotton, two in each subgenome, repeats from different arrays must have become homogenized by interlocus concerted evolution. Southern hybridization analysis combined with copy-number estimation demonstrate that this process has gone to completion in the diploids and to completion or near-completion in all allopolyploid species and that it most likely involves the entire rDNA repeat. Phylogenetic analysis demonstrates that interlocus concerted evolution has been bidirectional in allopolyploid species--i.e., rDNA from four polyploid lineages has been homogenized to a D genome repeat type, whereas sequences from Gossypium mustelinum have concerted to an A genome repeat type. Although little is known regarding the functional significance of interlocus concerted evolution of homeologous sequences, this study demonstrates that the process occurs for tandemly repeated sequences in diploid and polyploid plants. That interlocus concerted evolution can occur bidirectionally subsequent to hybidization and polyploidization has significant implications for phylogeny reconstruction, especially when based on rDNA sequences.

Base Sequence↗
PubMed1995

Rapid genome change in synthetic polyploids of Brassica and its implications for polyploid evolution.

Although the evolutionary success of polyploidy in higher plants has been widely recognized, there is virtually no information on how polyploid genomes have evolved after their formation. In this report, we used synthetic polyploids of Brassica as a model system to study genome evolution in the early generations after polyploidization. The initial polyploids we developed were completely homozygous, and thus, no nuclear genome changes were expected in self-fertilized progenies. However, extensive genome change was detected by 89 nuclear DNA clones used as probes. Most genome changes involved loss and/or gain of parental restriction fragments and appearance of novel fragments. Genome changes occurred in each generation from F2 to F5, and the frequency of change was associated with divergence of the diploid parental genomes. Genetic divergence among the derivatives of synthetic polyploids was evident from variation in genome composition and phenotypes. Directional genome changes, possibly influenced by cytoplasmic-nuclear interactions, were observed in one pair of reciprocal synthetics. Our results demonstrate that polyploid species can generate extensive genetic diversity in a short period of time. The occurrence and impact of this process in the evolution of natural polyploids is unknown, but it may have contributed to the success and diversification of many polyploid lineages in both plants and animals.

Biological Evolution↗
PubMed1996

A change of ploidy can modify epigenetic silencing.

A silent transgene in Arabidopsis thaliana was reactivated in an outcross but not upon selfing of hemizygous plants. This result could only be explained by assuming a genetic difference between the transgene-free gametes of the wild-type and hemizygous transgenic plants, respectively, and led to the discovery of ploidy differences between the parental plants. To investigate whether a change of ploidy by itself can indeed influence gene expression, we performed crosses of diploid or tetraploid plants with a strain containing a single copy of a transgenic resistance gene in an active state. We observed reduced gene expression of the transgene in triploid compared with diploid hybrids. This led to loss of the resistant phenotype at various stages of seedling development in part of the population. The gene inactivation was reversible. Thus, an increased number of chromosomes can result in a new type of epigenetic gene inactivation, creating differences in gene expression patterns. We discuss the possible impact of this finding for genetic diploidization in the light of widespread, naturally occurring polyploidy and polysomaty in plants.

Arabidopsis↗
PubMed1996

Hypothesis: "Rogue cell"-type chromosomal damage in lymphocytes is associated with infection with the JC human polyoma virus and has implications for oncopenesis.

The hemagglutination inhibition antibody titers against the JC and BK polyoma viruses (JCV and BKV, respectively) are significantly elevated in individuals exhibiting "rogue" cells among their cultured lymphocytes. However, the elevation is so much greater with respect to JCV that the BKV elevation could readily be explained by cross reactivity to the capsid protein of these two closely related viruses. The JCV exhibits high sequence homology with the simian papovavirus, simian virus 40 (SV40), and inoculation of human fetal brain cells with JCV produces polyploidy and chromosomal damage very similar to that produced by SV40. We suggest, by analogy with the effects of SV40, that these changes are due to the action of the viral large tumor antigen, a pluripotent DNA binding protein that acts in both transcription and replication. The implications of these findings for oncogenesis are briefly discussed.

Adolescent↗
PubMed1989

Endpolyploidy and digestion in the midgut of Rhodnius prolixus Stål (Hemiptera: Reduviidae).

The Feulgen-DNA content of the midgut cell nuclei of Rhodnius prolixus is estimated by microspectrophotometry. Midgut cells contain 23.35-32.16 pg DNA/nucleus, which represents an increase of up to 26c when compared to diploid R. prolixus brain tissue. A high copy number cell population disappears from the anterior intestinal cells after feeding, and the presence of a new population of low copy number cells indicates that there may be some regeneration of the midgut epithelium in adult insects, or that non-mitotic division of the high ploidy cells occurs. Polyploidy of the midgut cells is thought to be important in the rapid post-feeding synthesis of digestive proteinases, whereby enzymes are produced without the immediate need for DNA synthesis.

Animals↗
PubMedDate not supplied

Hypoxia and induced mutations in Syrian (golden) hamsters. The origin and perpetuation of a species.

We were able to identify anomalous chromosome patterns-monosomy, trisomy, and polyploidy-in metaphase plates prepared from embryos of Syrian hamsters (Mesocricetus auratus) whose mothers had been exposed to low-pressure hypoxia during the hours after copulation. These females were exposed in a low-pressure chamber for 4 hours at pressures equivalent to 30,000 feet of altitude before (estimated) ovulation to create acid shifts of pH equilibrium in the milieu of newly released eggs during passage of sperm upward, during fertilization itself, and during early cleavage stages. The D group chromosomes were most frequently involved in both monosomy and hyperdiploidy. The easily identifiable X chromosome also appeared to be susceptible to aneuploid formation. Chromosomally abnormal patterns most frequently among sibling embryos conceived by aging mothers exposed to low-pressure hypoxia. The significance of induced mutations is discussed.

Animals↗
PubMed1975

Chromosomal aberrations in rats fed irradiated wheat.

The effects on chromosomal aberrations in bone-marrow cells of feeding irradiated wheat to well-fed and malnourished rats were investigated. Rats maintained on low levels of dietary protein (5 per cent) had a higher incidence of breaks and deletions in their bone-marrow chromosomes compared with rats maintained on adequate levels of protein (18 per cient), showing that the level of dietary protein per se can produce structural abnormalities. Feeding of irradiated wheat to rats was associated with increase in the number of polyploid cells in the bone-marrow. The level of protein in the diet did not appear to influence polyploidy.

Animals↗
PubMed2005

Placing paleopolyploidy in relation to taxon divergence: a phylogenetic analysis in legumes using 39 gene families.

Young polyploid events are easily diagnosed by various methods, but older polyploid events become increasingly difficult to identify as chromosomal rearrangements, tandem gene or partial chromosome duplications, changes in substitution rates among duplicated genes, pseudogenization or locus loss, and interlocus interactions complicate the means of inferring past genetic events. Genomic data have provided valuable information about the polyploid history of numerous species, but on their own fail to show whether related species, each with a polyploid past, share a particular polyploid event. A phylogenetic approach provides a powerful method to determine this but many processes may mislead investigators. These processes can affect individual gene trees, but most likely will not affect all genes, and almost certainly will not affect all genes in the same way. Thus, a multigene approach, which combines the large-scale aspect of genomics with the resolution of phylogenetics, has the power to overcome these difficulties and allow us to infer genomic events further into the past than would otherwise be possible. Previous work using synonymous distances among gene pairs within species has shown evidence for large-scale duplications in the legumes Glycine max and Medicago truncatula. We present a case study using 39 gene families, each with three or four members in G. max and the putative orthologues in M. truncatula, rooted using Arabidopsis thaliana. We tested whether the gene duplications in these legumes occurred separately in each lineage after their divergence (Hypothesis 1), or whether they share a round of gene duplications (Hypothesis 2). Many more gene family topologies supported Hypothesis 2 over Hypothesis 1 (11 and 2, respectively), even after synonymous distance analysis revealed that some topologies were providing misleading results. Only ca. 33% of genes examined support either hypothesis, which strongly suggests that single gene family approaches may be insufficient when studying ancient events with nuclear DNA. Our results suggest that G. max and M. truncatula, along with approximately 7000 other legume species from the same clade, share an ancient round of gene duplications, either due to polyploidy or to some other process.

Computational Biology↗
PubMed2000

Orbit, a novel microtubule-associated protein essential for mitosis in Drosophila melanogaster.

We describe a Drosophila gene, orbit, that encodes a conserved 165-kD microtubule-associated protein (MAP) with GTP binding motifs. Hypomorphic mutations in orbit lead to a maternal effect resulting in branched and bent mitotic spindles in the syncytial embryo. In the larval central nervous system, such mutants have an elevated mitotic index with some mitotic cells showing an increase in ploidy. Amorphic alleles show late lethality and greater frequencies of hyperploid mitotic cells. The presence of cells in the hypomorphic mutant in which the chromosomes can be arranged, either in a circular metaphase or an anaphase-like configuration on monopolar spindles, suggests that polyploidy arises through spindle and chromosome segregation defects rather than defects in cytokinesis. A role for the Orbit protein in regulating microtubule behavior in mitosis is suggested by its association with microtubules throughout the spindle at all mitotic stages, by its copurification with microtubules from embryonic extracts, and by the finding that the Orbit protein directly binds to MAP-free microtubules in a GTP-dependent manner.

Alleles↗
PubMed2001

Drosophila aurora B kinase is required for histone H3 phosphorylation and condensin recruitment during chromosome condensation and to organize the central spindle during cytokinesis.

Aurora/Ipl1-related kinases are a conserved family of enzymes that have multiple functions during mitotic progression. Although it has been possible to use conventional genetic analysis to dissect the function of aurora, the founding family member in Drosophila (Glover, D.M., M.H. Leibowitz, D.A. McLean, and H. Parry. 1995. Cell. 81:95-105), the lack of mutations in a second aurora-like kinase gene, aurora B, precluded this approach. We now show that depleting Aurora B kinase using double-stranded RNA interference in cultured Drosophila cells results in polyploidy. aurora B encodes a passenger protein that associates first with condensing chromatin, concentrates at centromeres, and then relocates onto the central spindle at anaphase. Cells depleted of the Aurora B kinase show only partial chromosome condensation at mitosis. This is associated with a reduction in levels of the serine 10 phosphorylated form of histone H3 and a failure to recruit the Barren condensin protein onto chromosomes. These defects are associated with abnormal segregation resulting from lagging chromatids and extensive chromatin bridging at anaphase, similar to the phenotype of barren mutants (Bhat, M.A., A.V. Philp, D.M. Glover, and H.J. Bellen. 1996. Cell. 87:1103-1114.). The majority of treated cells also fail to undertake cytokinesis and show a reduced density of microtubules in the central region of the spindle. This is accompanied by a failure to correctly localize the Pavarotti kinesin-like protein, essential for this process. We discuss these conserved functions of Aurora B kinase in chromosome transmission and cytokinesis.

Adenosine Triphosphatases↗
PubMed1960

Is there "metabolic" DNA in the mouse seminal vesicle?

THIS STUDY WAS DESIGNED TO ANSWER THE QUESTION: Is H(3)-thymidine uptake by nuclei of the mouse seminal vesicle evidence for DNA synthesis and mitosis, or does it signify some "metabolic" function of DNA unrelated to chromosome duplication? Mice were given an intraperitoneal injection of H(3)-thymidine. Six hours later Feulgen squashes of the seminal vesicle epithelium were made and covered with autoradiographic stripping film. The silver grains above labeled nuclei were counted, and the Feulgen dye contents of these same nuclei were determined photometrically after removal of the grains from the emulsion. Unlabeled nuclei were also measured. The dye contents of non-radioactive nuclei form a unimodal distribution, indicating that polyploidy is absent from this tissue. The radioactive nuclei fall into two groups. In the first, the average dye content is the same as that of the cold nuclei (2C). In the second, the values range from 2C to 4C. In the 2C to 4C group the grain count is proportional to the dye content, showing that incorporation is correlated with synthesis. The radioactive 2C nuclei arose by mitosis during the course of the experiment. This is shown by the following facts: (1) They frequently occur in pairs. (2) They average smaller than unlabeled 2C nuclei. (3) Their average grain count is approximately half that of the 4C nuclei. (4) Labeled division figures are found. (5) A mitotic rate estimated from the number of labeled 2C nuclei accords reasonably well with one based on the number of observed mitoses. Since the incorporation of thymidine accompanies DNA synthesis and precedes mitosis, there is no reason to postulate a special "metabolic" DNA in this tissue.

Animals↗