Losses of chromosome arms 4q, 8p, 13q and gain of 8q are correlated with increasing chromosomal instability in hepatocellular carcinoma

OBJECTIVE: Chromosomal instability is a key feature in hepatocellular carcinoma (HCC). Array comparative genomic hybridization (aCGH) revealed recurring structural aberrations, whereas fluorescence in situ hybridization (FISH) indicated an increasing number of numerical aberrations in dedifferentiating HCC. Therefore, we examined whether there was a correlation between structural and numerical aberrations of chromosomal instability in HCC. METHODS AND RESULTS: 27 HCC (5 well, 10 moderately, 12 lower differentiated) already cytogenetically characterized by aCGH were analyzed. FISH analysis using probes for chromosomes 1, 3, 7, 8 and 17 revealed 1.46-4.24 signals/nucleus, which correlated with the histological grade (well vs. moderately,p < 0.0003; moderately vs. lower, p < 0.004). The number of chromosomes to each other was stable with exceptions only seen for chromosome 8. Loss of 4q and 13q, respectively, were correlated with the number of aberrations detected by aCGH (p < 0.001, p < 0.005; Mann-Whitney test). Loss of 4q and gain of 8q were correlated with an increasing number of numerical aberrations detected by FISH (p < 0.020, p < 0.031). Loss of 8p was correlated with the number of structural imbalances seen in aCGH (p < 0.048), but not with the number of numerical changes seen in FISH. CONCLUSION: We found that losses of 4q, 8p and 13q were closely correlated with an increasing number of aberrations detected by aCGH, whereas a loss of 4q and a gain of 8q were also observed in the context of polyploidization, the cytogenetic correlate of morphological dedifferentiation.

Phylogeny of horse chromosome 5q in the genus Equus and centromere repositioning

Horses, asses and zebras belong to the genus Equus and are the only extant species of the family Equidae in the order Perissodactyla. In a previous work we demonstrated that a key factor in the rapid karyotypic evolution of this genus was evolutionary centromere repositioning, that is, the shift of the centromeric function to a new position without alteration of the order of markers along the chromosome. In search of previously undiscovered evolutionarily new centromeres, we traced the phylogeny of horse chromosome 5, analyzing the order of BAC markers, derived from a horse genomic library, in 7 Equus species (E. caballus, E. hemionus onager, E. kiang, E. asinus, E. grevyi, E. burchelli and E. zebra hartmannae). This analysis showed that repositioned centromeres are present in E. asinus (domestic donkey, EAS) chromosome 16 and in E. burchelli (Burchell's zebra, EBU) chromosome 17, confirming that centromere repositioning is a strikingly frequent phenomenon in this genus. The observation that the neocentromeres in EAS16 and EBU17 are in the same chromosomal position suggests that they may derive from the same event and therefore, E. asinus and E. burchelli may be more closely related than previously proposed; alternatively, 2 centromere repositioning events, involving the same chromosomal region, may have occurred independently in different lineages, pointing to the possible existence of hot spots for neocentromere formation. Our comparative analysis also showed that, while E. caballus chromosome 5 seems to represent the ancestral configuration, centric fission followed by independent fusion events gave rise to 3 different submetacentric chromosomes in other Equus lineages.

The bovine dilated cardiomyopathy locus maps to a 1.0-Mb interval on chromosome 18

Cardiomyopathies are myocardial diseases that lead to cardiac dysfunction, heart failure, arrhythmia, and sudden death. In human medicine, cardiomyopathies frequently warrant heart transplantation in children and adults. Bovine dilated cardiomyopathy (BDCMP) is a heart muscle disorder that has been observed during the last 30 years in cattle of Holstein-Friesian origin. In Switzerland BDCMP affects Swiss Fleckvieh and Red Holstein breeds. BDCMP is characterized by a cardiac enlargement with ventricular remodeling and chamber dilatation. The common symptoms in affected animals are subacute subcutaneous edema, congestion of the jugular veins, and tachycardia with gallop rhythm. A cardiomegaly with dilatation and hypertrophy of all heart chambers, myocardial degeneration, and fibrosis are typical postmortem findings. It was shown that all BDCMP cases reported worldwide traced back to a red factor-carrying Holstein-Friesian bull, ABC Reflection Sovereign. An autosomal recessive mode of inheritance was proposed for BDCMP. Recently, the disease locus was mapped to a 6.7-Mb interval MSBDCMP06-BMS2785 on bovine Chr 18 (BTA18). In the present study the BDCMP locus was fine mapped by using a combined strategy of homozygosity mapping and association study. A BAC contig of 2.9 Mb encompassing the crucial interval was constructed to establish the correct marker order on BTA18. We show that the disease locus is located in a gene-rich interval of 1.0 Mb and is flanked by the microsatellite markers DIK3006 and MSBDCMP51.

Arachnomelia in Brown Swiss cattle maps to chromosome 5

Arachnomelia in Brown Swiss cattle is a monogenic autosomal recessive inherited congenital disorder of the skeletal system giving affected calves a spidery look (OMIA ID 000059). Over a period of 20 years 15 cases were sampled in the Swiss and Italian Brown cattle population. Pedigree data revealed that all affected individuals trace back to a single acknowledged carrier founder sire. A genome scan using 240 microsatellites spanning the 29 bovine autosomes showed homozygosity at three adjacent microsatellite markers on bovine Chr 5 in all cases. Linkage analysis confirmed the localization of the arachnomelia mutation in the region of the marker ETH10. Fine-mapping and haplotype analysis using a total of 34 markers in this region refined the critical region of the arachnomelia locus to a 7.19-Mb interval on bovine Chr 5. The disease-associated IBD haplotype was shared by 36 proven carrier animals and allows marker-assisted selection. As the corresponding human and mouse chromosome segments do not contain any clear functional candidate genes for this disorder, the mutation causing arachnomelia in the Brown Swiss cattle might help to identify an unknown gene in bone development.

Molecular phylogenetic analyses identify Alpine differentiation and dysploid chromosome number changes as major forces for the evolution of the European endemic Phyteuma (Campanulaceae)

Phyteuma is a chromosomally and ecologically diverse vascular plant genus and constitutes an excellent system for studying both the role of chromosomal change for species diversification and the evolution of high-mountain biota. This kind of research is, however, hampered by the lack of a sound phylogenetic framework exacerbated by the notoriously low predictive power of traditional taxonomy with respect to phylogenetic relationships in Campanulaceae. Based on a comprehensive taxon sampling and analyses of nuclear and plastid sequence and AFLP fingerprint data, Phyteuma is confirmed as a monophyletic group sister to the monotypic Physoplexis, which is in line with their peculiar flower morphologies. Within Phyteuma two clades, largely corresponding to previously recognized sections, are consistently found. The traditional circumscription of taxonomic series is largely rejected. Whereas distinctness of the currently recognized species is mostly corroborated, some interspecific relationships remain ambiguous due to incongruences between nuclear and plastid data. Major forces for diversification and evolution of Phyteuma are descending dysploidy (i.e., a decrease in chromosome base number) as well as allopatric and ecological differentiation within the Alps, the genus' center of species diversity. (C) 2013 Elsevier Inc. All rights reserved.

Novel pseudo-staphylococcal cassette chromosome mec element (ψSCCmec57395) in methicillin-resistant Staphylococcus pseudintermedius CC45

Genetic characterization of methicillin-resistant Staphylococcus pseudintermedius (MRSP) from Thailand and Israel revealed the presence of a predominant atypical clonal lineage which was not typeable by SmaI-PFGE and SCCmec typing. All the atypical isolates (n = 34) belonged to CC45 (30 ST45 and 2 ST179 isolates, 1 ST57 isolate, and 1 ST85 isolate). The isolates originated from healthy and diseased dogs and cats, as well as from the environment of one clinic. Cfr9I-pulsed-field gel electrophoresis (Cfr9I-PFGE) and dru typing permitted the further distinction of CC45 isolates from the two different countries. Microarray analysis identified genes that confer resistance to β-lactams (mecA; blaZ), aminoglycosides [aac(6')-Ie-aph(2')-Ia; aph(3')-III; ant(6)-Ia], macrolides and lincosamides [erm(B)], tetracyclines [tet(M)], trimethoprim [dfr(G)], streptothricin (sat4), and chloramphenicol (catpC221). Fluoroquinolone resistance was attributed to specific amino acid substitutions, i.e., Ser84Leu in GyrA and Ser80Ile and Asp84Asn in GrlA. A novel pseudo-staphylococcal cassette chromosome (ΨSCCmec57395) element was identified in MRSP strain 57395 (sequence type ST45) by whole-genome sequencing. The 12,282-bp ΨSCCmec57395 element contained a class C1 mec gene complex but no ccr genes. In addition to the methicillin resistance gene mecA, ΨSCCmec57395 also carried determinants of resistance to heavy metals, such as arsenic, cadmium, and copper. Bsu36I restriction analysis of the ΨSCCmec57395 element amplified by long-range PCR revealed the presence of ΨSCCmec57395 in the 33 additional isolates of MRSP CC45. The ΨSCCmec57395 element represents a new class of SCCmec and has been identified in MRSP of CC45, which is a predominant clonal lineage in Israel and Thailand.

Loss of DAXX and ATRX are associated with chromosome instability and reduced survival of patients with pancreatic neuroendocrine tumors

BACKGROUND & AIMS Sporadic pancreatic neuroendocrine tumors (pNETs) are rare and genetically heterogeneous. Chromosome instability (CIN) has been detected in pNETs from patients with poor outcomes, but no specific genetic factors have been associated with CIN. Mutations in death domain-associated protein gene (DAXX) or ATR-X gene (ATRX) (which both encode proteins involved in chromatin remodeling) have been detected in 40% of pNETs, in association with activation of alternative lengthening of telomeres. We investigated whether loss of DAXX or ATRX, and consequent alternative lengthening of telomeres, are related to CIN in pNETs. We also assessed whether loss of DAXX or ATRX is associated with specific phenotypes of pNETs. METHODS We collected well-differentiated primary pNET samples from 142 patients at the University Hospital Zurich and from 101 patients at the University Hospital Bern (both located in Switzerland). Clinical follow-up data were obtained for 149 patients from general practitioners and tumor registries. The tumors were reclassified into 3 groups according to the 2010 World Health Organization classification. Samples were analyzed by immunohistochemistry and telomeric fluorescence in situ hybridization. We correlated loss of DAXX, or ATRX, expression, and activation of alternative lengthening of telomeres with data from comparative genomic hybridization array studies, as well as with clinical and pathological features of the tumors and relapse and survival data. RESULTS Loss of DAXX or ATRX protein and alternative lengthening of telomeres were associated with CIN in pNETs. Furthermore, loss of DAXX or ATRX correlated with tumor stage and metastasis, reduced time of relapse-free survival, and decreased time of tumor-associated survival. CONCLUSIONS Loss of DAXX or ATRX is associated with CIN in pNETs and shorter survival times of patients. These results support the hypothesis that DAXX- and ATRX-negative tumors are a more aggressive subtype of pNET, and could lead to identification of strategies to target CIN in pancreatic tumors.

Characterization of new staphylococcal cassette chromosome mec (SCCmec) and topoisomerase genes in fluoroquinolone- and methicillin-resistant Staphylococcus pseudintermedius

Fluoroquinolone- and methicillin-resistant Staphylococcus pseudintermedius isolates harbor two new staphylococcal cassette chromosome mec (SCCmec) elements that belong to class A, allotype 3 (SCCmec II-III), and to the new allotype 5 (SCCmec VII). Analysis of the complete nucleotide sequences of the topoisomerase loci gyrB/gyrA and grlB/grlA revealed mutations involved in fluoroquinolone resistance.

Cytomegalovirus Serology and Replication Remain Associated With Solid Organ Graft Rejection and Graft Loss in the Era of Prophylactic Treatment

BACKGROUND Cytomegalovirus (CMV) replication has been associated with more risk for solid organ graft rejection. We wondered whether this association still holds when patients at risk receive prophylactic treatment for CMV. METHODS We correlated CMV infection, biopsy-proven graft rejection, and graft loss in 1,414 patients receiving heart (n=97), kidney (n=917), liver (n=237), or lung (n=163) allografts reported to the Swiss Transplant Cohort Study. RESULTS Recipients of all organs were at an increased risk for biopsy-proven graft rejection within 4 weeks after detection of CMV replication (hazard ratio [HR] after heart transplantation, 2.60; 95% confidence interval [CI], 1.34-4.94, P<0.001; HR after kidney transplantation, 1.58; 95% CI, 1.16-2.16, P=0.02; HR after liver transplantation, 2.21; 95% CI, 1.53-3.17, P<0.001; HR after lung transplantation, 5.83; 95% CI, 3.12-10.9, P<0.001. Relative hazards were comparable in patients with asymptomatic or symptomatic CMV infection. The CMV donor or recipient serological constellation also predicted the incidence of graft rejection after liver and lung transplantation, with significantly higher rates of rejection in transplants in which donor or recipient were CMV seropositive (non-D-/R-), compared with D- transplant or R- transplant (HR, 3.05; P=0.002 for liver and HR, 2.42; P=0.01 for lung transplants). Finally, graft loss occurred more frequently in non-D- or non-R- compared with D- transplant or R- transplant in all organs analyzed. Valganciclovir prophylactic treatment seemed to delay, but not prevent, graft loss in non-D- or non-R- transplants. CONCLUSION Cytomegalovirus replication and donor or recipient seroconstellation remains associated with graft rejection and graft loss in the era of prophylactic CMV treatment.

Two Loci on Chromosome 5 Are Associated with Serum IgE Levels in Labrador Retrievers

Crosslinking of immunoglobulin E antibodies (IgE) bound at the surface of mast cells and subsequent mediator release is considered the most important trigger for allergic reactions. Therefore, the genetic control of IgE levels is studied in the context of allergic diseases, such as asthma, atopic rhinitis, or atopic dermatitis (AD). We performed genome-wide association studies in 161 Labrador Retrievers with regard to total and allergen-specific immunoglobulin E (IgE) levels. We identified a genome-wide significant association on CFA 5 with the antigen-specific IgE responsiveness to Acarus siro. We detected a second genome-wide significant association with respect to the antigen-specific IgE responsiveness to Tyrophagus putrescentiae at a different locus on chromosome 5. A. siro and T. putrescentiae both belong to the family Acaridae and represent so-called storage or forage mites. These forage mites are discussed as major allergen sources in canine AD. No obvious candidate gene for the regulation of IgE levels is located under the two association signals. Therefore our studies offer a chance of identifying a novel mechanism controlling the host's IgE response.

Whole genome scan identifies several chromosomal regions linked to equine sarcoids.

Despite the evidence for a genetic predisposition to develop equine sarcoids (ES), no whole genome scan for ES has been performed to date. The objective of this explorative study was to identify chromosome regions associated with ES. The studied population was comprised of two half-sibling sire families, involving a total of 222 horses. Twenty-six of these horses were affected with ES. All horses had been previously genotyped with 315 microsatellite markers. Quantitative trait locus (QTL) signals were suggested where the F statistic exceeded chromosome-wide significance at P < 0.05. The QTL analyses revealed significant signals reaching P < 0.05 on equine chromosome (ECA) 20, 23 and 25, suggesting a polygenic character for this trait. The candidate regions identified on ECA 20, 23 and 25 include genes regulating virus replication and host immune response. Further investigation of the chromosome regions associated with ES and of genes potentially responsible for the development of ES could form the basis for early identification of susceptible animals, breeding selection or the development of new therapeutic targets.

FUS protein interacts with U7 snRNP and plays a role in replication-dependant histone genes expression

The U7 small nuclear ribonucleoprotein (U7 snRNP) is an essential factor mediating the unique 3’end processing of non-polyadenylated, replication-dependent histone mRNAs in metazoans. These histone genes expression and processing of their transcripts are cell cycle-regulated mechanisms that recruit a number of specific proteins as well as common factors required for expression and maturation of polyadenylated mRNAs. However, despite all the knowledge we have so far, there are still gaps in understanding of core histone RNA 3’ end processing, its coupling to transcription and regulation during cell cycle. To further elucidate this phenomena we used affinity chromatography based on tagged version of U7 snRNA molecule to identify proteins associated with U7 snRNP/U7 snRNA that could be potentially involved in core histone genes expression in human cells. Mass spectrometric analysis of affinity-purified fraction revealed, among others, multifunctional RNA/DNAbinding protein FUS/TLS (fused in sarcoma/translocated in liposarcoma) as a new factor interacting with U7 snRNA/RNP. Co-immunoprecipitation and RIP experiments confirmed the binding between FUS and the U7 RNA/snRNP. Interestingly, FUS:U7 snRNA interaction seems to be activated in S phase where the core histone genes are expressed. Moreover, FUS co-fractionates in 10-50% continuous glycerol gradient with other factors involved in histone premRNAs 3’end processing. However, this unique 3’end maturation was not disturbed upon FUS knockdown. Instead, we found that FUS depletion leads to a de-regulation of expression from selected histone promoters, suggesting that FUS is rather involved in regulation of core histone genes transcription. Thus, FUS bound to U7 snRNP can play a role in coupling between transcription and 3’end processing of replication dependant histone mRNAs.

The Host Nonsense-Mediated mRNA Decay Pathway Restricts Mammalian RNA Virus Replication

In addition to classically defined immune mechanisms, cell-intrinsic processes can restrict virus infection and have shaped virus evolution. The details of this virus-host interaction are still emerging. Following a genome-wide siRNA screen for host factors affecting replication of Semliki Forest virus (SFV), a positive-strand RNA (+RNA) virus, we found that depletion of nonsense-mediated mRNA decay (NMD) pathway components Upf1, Smg5, and Smg7 led to increased levels of viral proteins and RNA and higher titers of released virus. The inhibitory effect of NMD was stronger when virus replication efficiency was impaired by mutations or deletions in the replicase proteins. Consequently, depletion of NMD components resulted in a more than 20-fold increase in production of these attenuated viruses. These findings indicate that a cellular mRNA quality control mechanism serves as an intrinsic barrier to the translation of early viral proteins and the amplification of +RNA viruses in animal cells.