Transcription of a centromere-enriched retroelement and local retention of its RNA are significant features of the CENP-A chromatin landscape.

Centromeres depend on chromatin containing the conserved histone H3 variant CENP-A for function and inheritance, while the role of centromeric DNA repeats remains unclear. Retroelements are prevalent at centromeres across taxa and represent a potential mechanism for promoting transcription to aid in CENP-A incorporation or for generating RNA transcripts to maintain centromere integrity. Here, we probe into the transcription and RNA localization of the centromere-enriched retroelement G2/Jockey-3 (hereafter referred to as Jockey-3 ) in Drosophila melanogaster , currently the only in vivo model with assembled centromeres. We find that Jockey-3 is a major component of the centromeric transcriptome and produces RNAs that localize to centromeres in metaphase. Leveraging the polymorphism of Jockey-3 and a de novo centromere system, we show that these RNAs remain associated with their cognate DNA sequences in cis , suggesting they are unlikely to perform a sequence-specific function at all centromeres. We show that Jockey-3 transcription is positively correlated with the presence of CENP-A, and that recent Jockey-3 transposition events have occurred preferentially at CENP-A-containing chromatin. We propose that Jockey-3 contributes to the epigenetic maintenance of centromeres by promoting chromatin transcription, while inserting preferentially within these regions, selfishly ensuring its continued expression and transmission. Given the conservation of retroelements as centromere components through evolution, our findings have broad implications in understanding this association in other species.

Zenker's diverticulitis: a bitter pill to swallow.

Acute oesophageal obstruction from food bolus impaction is often triggered by underlying oesophageal pathology, both benign and malignant. These can be readily detected with standard investigations such as oesophagoscopy or computed tomography. Zenker's diverticulum (ZD) is a benign condition frequently presenting with chronic dysphagia or may be asymptomatic. We report the case of an 81-year-old man with a previously undiagnosed 1-cm ZD causing complete oesophageal obstruction secondary to localized oedema from an impacted ibuprofen tablet. Although initial clinical, endoscopic and radiological findings were equivocal and suspicious for upper oesophageal malignancy, symptoms rapidly settled in response to systemic corticosteroids. The diagnosis was later confirmed on barium swallow with no other clinical, radiological or histopathological abnormalities identified. In conclusion, ZD is an uncommon cause of acute oesophageal obstruction which may occur in diverticula of all sizes. Surgery should be performed in patients with recurrent symptoms or large diverticula.

Association of BMI with Clinicopathological Features of Papillary Thyroid Cancer: A Systematic Review and Meta-Analysis.

BACKGROUND: Papillary thyroid cancer (PTC) is the most common subtype of thyroid cancer. The incidence of PTC is rising in tandem with an obesity epidemic. Associations have been demonstrated between increased body mass index (BMI) and worse oncological outcomes in a number of malignancies. However, research on this topic in PTC to date has been inconsistent, often due to limited data. This study aimed to measure the association between BMI and potentially adverse clinicopathological features of PTC. METHODS: A meta-analysis of studies reporting outcomes after surgical treatment of PTC was performed. PubMed, Embase and the Cochrane Library were searched systematically to identify studies which provided data on BMI and clinicopathologic features of PTC. Relevant data were extracted and synthesis performed using adjusted odds ratios where available and crude values when not. Data were analysed by inverse variance using random and fixed effects models. RESULTS: Data on 35,237 patients from 15 studies met the criteria for inclusion. Obesity was associated with larger tumour size (MD = 0.17 cm [0.05, 0.29]), increased rates of multifocality (OR = 1.41 [1.16, 1.70]), extrathyroidal extension (OR = 1.70 [1.39, 2.07]) and nodal spread (OR = 1.18 [1.07, 1.30]). Associations were more pronounced as BMI increased. There was no association between BMI and bilaterality, vascular invasion or metastatic spread. CONCLUSION: Increased BMI is significantly associated with multiple potentially adverse features of PTC. The effect on long-term oncological outcomes requires further evaluation.

Esperanto for histones: CENP-A, not CenH3, is the centromeric histone H3 variant.

The first centromeric protein identified in any species was CENP-A, a divergent member of the histone H3 family that was recognised by autoantibodies from patients with scleroderma-spectrum disease. It has recently been suggested to rename this protein CenH3. Here, we argue that the original name should be maintained both because it is the basis of a long established nomenclature for centromere proteins and because it avoids confusion due to the presence of canonical histone H3 at centromeres.

Making a long story short: noncoding RNAs and chromosome change.

As important as the events that influence selection for specific chromosome types in the derivation of novel karyotypes, are the events that initiate the changes in chromosome number and structure between species, and likewise polymorphisms, variants and disease states within species. Although once thought of as transcriptional 'noise', noncoding RNAs (ncRNAs) are now recognized as important mediators of epigenetic regulation and chromosome stability. Here we highlight recent work that illustrates the influence short and long ncRNAs have as participants in the function and stability of chromosome regions such as centromeres, telomeres, evolutionary breakpoints and fragile sites. We summarize recent evidence that ncRNAs can facilitate chromosome change and present mechanisms by which ncRNAs create DNA breaks. Finally, we present hypotheses on how they may create novel karyotypes and thus affect chromosome evolution.

Centromere conversion and retention in somatic cell hybrids.

The generation of somatic cell hybridization-derived cell lines between highly divergent species affords the opportunity to examine the concept of 'genome dominance' in the context of genetic and epigenetic changes. While whole-scale genome dominance has been well documented in natural hybrids among closely related species, an examination of centromere position and sequence retention in 2 marsupial-eutherian hybrids has revealed a mechanism for 'centromere dominance' as a driving force in the generation of stable somatic cell hybrids following an initial period of genomic instability. While one somatic cell hybrid cell line appeared to retain marsupial centromere sequences which remained competent to recruit the centromere-specific histone variant CENP-A in a Chinese hamster background, fusion events between marsupial and mouse-derived chromosomes in another hybrid line led to a centromere sequence conversion from one species to the other. We postulate that the necessity to maintain an epigenetically defined centromere following genome hybridization may be responsible for retention of specific chromosomes and may result in rapid sequence turnover to facilitate the recruitment of CENP-A containing histones.

Divergent patterns of breakpoint reuse in Muroid rodents.

Multiple Genome Rearrangement (MGR) analysis was used to define the trajectory and pattern of chromosome rearrangement within muroid rodents. MGR was applied using 107 chromosome homologies between Mus, Rattus, Peromyscus, the muroid sister taxon Cricetulus griseus, and Sciurus carolinensis as a non-Muroidea outgroup, with specific attention paid to breakpoint reuse and centromere evolution. This analysis revealed a high level of chromosome breakpoint conservation between Rattus and Peromyscus and indicated that the chromosomes of Mus are highly derived. This analysis identified several conserved evolutionary breakpoints that have been reused multiple times during karyotypic evolution in rodents. Our data demonstrate a high level of reuse of breakpoints among muroid rodents, further supporting the "Fragile Breakage Model" of chromosome evolution. We provide the first analysis of rodent centromeres with respect to evolutionary breakpoints. By analyzing closely related rodent species we were able to clarify muroid rodent karyotypic evolution. We were also able to derive several high-resolution ancestral karyotypes and identify rearrangements specific to various stages of Muroidea evolution. These data were useful in further characterizing lineage-specific modes of chromosome evolution.

Peromyscus maniculatus--Mus musculus chromosome homology map derived from reciprocal cross species chromosome painting.

The Mus musculus and Rattus norvegicus genomes have been extensively studied, yet despite the emergence of Peromyscus maniculatus as an NIH model for genome sequencing and biomedical research much remains unknown about the genome organization of Peromyscines. Contrary to their phylogenetic relationship, the genomes of Rattus and Peromyscus appear more similar at the gross karyotypic level than either does to Mus. We set out to define the chromosome homologies between Peromyscus, Mus and Rattus. Reciprocal cross-species chromosome painting and G-band homology assignments were used to delineate the conserved chromosome homology map between P. maniculatus and M. musculus. These data show that each species has undergone extensive chromosome rearrangements since they last shared a common ancestor 25 million years ago (mya). This analysis coupled with an inferred homology map with Rattus revealed a high level of chromosome conservation between Rattus and Peromyscus and indicated that the chromosomes of Mus are highly derived.

Methylation perturbations in retroelements within the genome of a Mus interspecific hybrid correlate with double minute chromosome formation.

A reduction in the DNA modification of cytosine methylation has been linked directly to chromosome rearrangements concomitant with retroelement amplification in several marsupial hybrid genomes. While phenotypes observed for interspecific eutherian hybrids are suggestive of methylation perturbations and retroelement instability, no link between retroelements, DNA methylation, and chromosome instability has yet been identified. Previous studies in eutherian hybrids, however, have been limited to a gross examination of methylation using methylation-sensitive restriction enzyme analysis or focused on single-copy genes and/or have avoided examination of repetitive DNA. Methylation changes and retroelements are proposed as mechanisms for double minute chromosome formation and oncogene amplification, both present in the genome of a Mus hybrid model, thus making it an ideal system to evaluate methylation status more closely. We have used the PCR-based methodologies methylation-sensitive amplicon subtraction (MS-AS) and methylation-sensitive representational difference analysis (MS-RDA) to detect differentially methylated sequences between three complex genomes and to isolate methylation perturbations in a Mus musculusxMus caroli hybrid. This novel application of MS-AS and MS-RDA resulted in the isolation of differentially methylated retroelements surrounding the locus on Chromosome 10 responsible for double minute chromosome formation within this interspecific eutherian hybrid.

Physical mapping of the IGF2 locus in the South American opossum Monodelphis domestica.

The South American opossum Monodelphis domestica has been a model organism for marsupials for many years and has recently been the subject of a large-scale genome sequencing effort that will provide the foundation for comparative studies of gene function and regulation. Genomic imprinting is one mechanism of gene regulation that has received increasing attention due to the impact of imprinting defects on development and disease. We have mapped the imprinted insulin-like growth factor II (IGF2) gene of M. domestica as a first step in understanding the regulatory mechanisms involved in genomic imprinting in this marsupial.

Retention of latent centromeres in the Mammalian genome.

The centromere is a cytologically defined entity that possesses a conserved and restricted function in the cell: it is the site of kinetochore assembly and spindle attachment. Despite its conserved function, the centromere is a highly mutable portion of the chromosome, carrying little sequence conservation across taxa. This divergence has made studying the movement of a centromere, either within a single karyotype or between species, a challenging endeavor. Several hypotheses have been proposed to explain the permutability of centromere location within a chromosome. This permutability is termed "centromere repositioning" when described in an evolutionary context and "neocentromerization" when abnormalities within an individual karyotype are considered. Both are characterized by a shift in location of the functional centromere within a chromosome without a concomitant change in linear gene order. Evolutionary studies across lineages clearly indicate that centromere repositioning is not a rare event in karyotypic evolution and must be considered when examining the evolution of chromosome structure and syntenic order. This paper examines the theories proposed to explain centromere repositioning in mammals. These theories are interpreted in light of evidence gained in human studies and in our presented data from the marsupial model species Macropus eugenii, the tammar wallaby.

Centromere dynamics and chromosome evolution in marsupials.

The eukaryotic centromere poses an interesting evolutionary paradox: it is a chromatin entity indispensable to precise chromosome segregation in all eukaryotes, yet the DNA at the heart of the centromere is remarkably variable. Its important role of spindle attachment to the kinetochore during meiosis and mitosis notwithstanding, recent studies implicate the centromere as an active player in chromosome evolution and the divergence of species. This is exemplified by centromeric involvement in translocations, fusions, inversions, and centric shifts. Often species are defined karyotypically simply by the position of the centromere on certain chromosomes. Little is known about how the centromere, either as a functioning unit of chromatin or as a specific block of repetitive DNA sequences, acts in the creation of these types of chromosome rearrangements in an evolutionary context. Macropodine marsupials (kangaroos and wallabies) offer unique insights into current theories expositing centromere emergence during karyotypic diversification and speciation.

Contrasting modes of natural selection acting on pigmentation genes in the Drosophila dunni subgroup.

Genes that encode for divergent adaptive traits may have genealogies that contrast with those from loci that are not functionally involved in differentiation. Here, we examine DNA sequence variation among the species of the eastern Caribbean Drosophila dunni subgroup at two loci, yellow and dopa decaboxylase (Ddc), which both play integral roles in pigmentation patterning of adult Drosophila. Phylogenetic analyses of these loci produce gene genealogies with topologies that mirror those described for other nuclear genes: the six morphologically distinct species within the subgroup are divided into only three lineages, with one lineage containing four species that share extensive ancestral polymorphism. At the Ddc locus these major lineages are delineated only by silent site variation. We observe a significantly higher rate of synonymous site divergence than non-synonymous divergence, consistent with strong purifying selection acting on the locus. In contrast, the yellow locus exhibits patterns of amino acid divergence and nucleotide diversity that are consistent with recent diversifying selection acting in two different lineages. This selection appears to be targeting amino acid variants in the signal sequence of the Yellow protein, a region which is tightly constrained among members of the larger D. cardini radiation. This result highlights not only the potential importance of yellow in the evolution of divergent pigmentation patterns among members of the D. dunni subgroup, but also hints that variation in signal peptide sequences may play a role in phenotypic diversification.

A centromere-specific retroviral element associated with breaks of synteny in macropodine marsupials.

Studies of chromosome evolution have focused heavily on the evolution of conserved syntenic, gene-rich domains. It is obvious, however, that the centromere plays an equally important role in chromosome evolution, through its involvement in fissions, centric fusions, translocations, inversions and centric shifts. It is unclear how the centromere, either as a functioning unit of the chromosome or as a DNA sequence motif, has been involved in these processes. Marsupials of the family Macropodidae (kangaroos, wallabies, rat kangaroos and potoroos) offer unique insights into current theories expositing centromere emergence during karyotypic diversification and speciation. Tracing the genomic distribution of centromeric sequences in a model macropodine (subfamily Macropodinae: kangaroos and wallabies) species, Macropus eugenii (tammar wallaby), indicates these sequences have played an important role in chromosome evolution through possible segmental duplications associated with phylogenetically conserved breaks of synteny, pericentromeric and subtelomeric regions. Hybrids between different kangaroo species provide evidence that the centromere is unstable within this group of mammals and is involved in a large number of chromosome aberrations. A better understanding of the genetic and epigenetic factors that define centromeres and how centromeres may mediate changes in chromosome architecture are critical not only to our understanding of basic cellular functioning but also to our understanding of the process of speciation.

Characterization and isolation of novel microsatellites from the Drosophila dunni subgroup.

We have isolated and characterized 77 novel microsatellites from two species, Drosophila dunni and Drosophila nigrodunni, which are closely related Caribbean-island endemics from the Drosophila cardini species group. These species are very distantly related to all other Drosophila from which microsatellites have previously been characterized. We find that the average length of microsatellites isolated in these species is quite small, with an overall mean length of 9.8 repeat units for dinucleotide microsatellites in the two study species. The nucleotide composition of dinucleotides differs between the two species: D. nigrodunni has a predominance of (AC/GT)n repeats, whereas D. dunni has equal numbers of (AC/GT)n and (AG/CT)n repeats. Tri- and tetranucleotide repeats are not abundant in either species. We assayed the variability of eight microsatellites in a closely related third species, Drosophila arawakana, using wild-caught individuals from the island of Guadeloupe. We found the microsatellites to be extremely variable in this population, with observed heterozygosities ranging from 0.541 to 0.889. DNA amplification trials suggest that these eight microsatellites are widely conserved across the D. cardini group, with five of the eight producing amplification products in every species tested. However, the loci are very poorly conserved over greater phylogenetic distances. DNA amplification of the microsatellite loci was unreliable in members of the closely related Drosophila quinaria, Drosophila calloptera, Drosophila guarani and Drosophila tripunctata species groups. Furthermore, these microsatellites could not be detected in the genome of Drosophila melanogaster, despite the conservation of microsatellite flanking regions at some loci. These data indicate that Drosophila microsatellite loci are quite short lived over evolutionary timescales relative to many other taxa.

Interspecific hybridization induced amplification of Mdm2 on double minutes in a Mus hybrid.

We report a case of interspecific hybridization induced amplification of Chromosome 10 on double minutes (dm) in the karyotype of a hybrid Mus embryo. Stable, non-mosaic dm were previously found in tissues of a 16.5-day Mus Musculus x Mus Caroli hybrid (Graves, 1984). Dm in tissues of the hybrid was of interest to us because of previous reports of genomic instability in interspecific hybrids (O'Neill et al., 1998) and thus we decided to characterize the dm in the hybrid karyotypes. Whole chromosome painting of the hybrid cell lines showed amplification of Chromosome 10 sequences. Southern analysis with a probe for the candidate gene Mdm2 showed amplification of the paternal allele of this oncogene. Overexpression of Mdm2 was confirmed by a western analysis that also showed an associated inactivation of the tumor suppressor, Trp53. Evidence indicates that the event leading to the instability observed was an early adaptive response to stress on the genome, i.e. interspecific hybridization.

Chromosome evolution in kangaroos (Marsupialia: Macropodidae): cross species chromosome painting between the tammar wallaby and rock wallaby spp. with the 2n = 22 ancestral macropodid karyotype.

Marsupial mammals show extraordinary karyotype stability, with 2n = 14 considered ancestral. However, macropodid marsupials (kangaroos and wallabies) exhibit a considerable variety of karyotypes, with a hypothesised ancestral karyotype of 2n = 22. Speciation and karyotypic diversity in rock wallabies (Petrogale) is exceptional. We used cross species chromosome painting to examine the chromosome evolution between the tammar wallaby (2n = 16) and three 2n = 22 rock wallaby species groups with the putative ancestral karyotype. Hybridization of chromosome paints prepared from flow sorted chromosomes of the tammar wallaby to Petrogale spp., showed that this ancestral karyotype is largely conserved among 2n = 22 rock wallaby species, and confirmed the identity of ancestral chromosomes which fused to produce the bi-armed chromosomes of the 2n = 16 tammar wallaby. These results illustrate the fission-fusion process of karyotype evolution characteristic of the kangaroo group.

Whatever happened to SRY?

The mammalian sex-determining gene, SRY, was identified by positional cloning approximately 10 years ago. Since its discovery, intense research into this gene has been directed on two main fronts: elucidation of its function in development of the testis and examination of its singular evolutionary history. The role or SRY as the testis-determining factor (TDF) places it at a crucial point in the highly conserved morphogenetic process of vertebrate gonadogenesis. None of the genes that directly activate SRY nor any of its immediate downstream targets have yet been positively identified. Several genes, however, such as SF1, DAX1, and SOX9, whose spatial and temporal expression profiles overlap with that of SRY, are strongly implicated as co-regulators of gonadogenesis. Molecular genetic manipulation of these genes in mice has shown that they are indispensable to sexual development. Remarkably, its key position in this cascade of gene action has not protected SRY from strong yet poorly understood selective forces that have caused it to evolve rapidly in mammals. The evolution of SRY has been characterized not only by rapid sequence divergence within mammals, but also by structural changes such as intron insertion, gene amplification, and deletion.

Undermethylation associated with retroelement activation and chromosome remodelling in an interspecific mammalian hybrid.

Genetic models predict that genomic rearrangement in hybrids can facilitate reproductive isolation and the formation of new species by preventing gene flow between the parent species and hybrid (sunflowers are an example). The mechanism underlying hybridization-induced chromosome remodelling is as yet unknown, although mobile element activity has been shown to be involved in DNA rearrangement in some dysgenic Drosophila hybrids. It has been proposed that DNA methylation evolved as a means of repressing the movement of mobile elements (the host defence model). If such a protective mechanism were to fail, mobile elements could be activated, and could cause major and rapid genome alterations. Here we demonstrate the occurrence of genome-wide undermethylation, retroviral element amplification and chromosome remodelling in an interspecific mammalian hybrid (Macropus eugenii x Wallabia bicolor). Atypically extended centromeres of Macropus eugenii derived autosomes in the hybrid were composed primarily of an unmethylated, amplified retroviral element not detectable in either parent species. These results, taken with the observation of deficient methylation and de novo chromosome change in other mammalian hybrids, indicate that the failure of DNA methylation and subsequent mobile-element activity in hybrids could facilitate rapid karyotypic evolution.