Insights into genetic alterations of liver metastases from uveal melanoma

The liver is the organ usually affected by metastatic uveal melanoma (MUM). Current treatments are almost always ineffective and mortality remains high. In this study, copy number variations (CNVs) were identified in 12 metastatic and five matched primary UMs (PUMs). Our data revealed a wide spectrum of genetic alterations in MUM. Most common were amplifications of chromosome (chr.) 8q; alterations on chr. 3 included monosomy, isodisomy, and large regions of homozygosity (ROH). Genomic profiles of PUM‐MUM pairs varied in their degree of similarity and complexity. However, within the pairs, 135 genes were consistently altered. Protein expression of C‐MYC and BAP1 was examined by immunohistochemistry (IHC); a positive association between IHC and CNVs was seen for C‐MYC. This comprehensive catalogue of CNVs associated with MUM should facilitate the identification of key alterations that drive tumor growth. This would have the potential to select urgently needed novel, targeted, therapeutic regimens.     

Novel Y‐chromosome polymorphisms in Chinese domestic yak

Y‐chromosome‐specific haplotypes (Y‐haplotypes) constructed using single nucleotide polymorphisms (Y‐SNPs) in the MSY (male‐specific region of the Y‐chromosome) are valuable in population genetic studies. But sequence variants in the yak MSY region have been poorly characterized so far. In this study, we screened a total of 16 Y‐chromosome‐specific gene segments from the ZFY, SRY, UTY, USP9Y, AMELY and OFD1Y genes to identify Y‐SNPs in domestic yaks. Six novel Y‐SNPs distributed in the USP9Y (g.223C>T), UTY19 (g.158A>C and g.169C>T), AMELY2 (g.261C>T), OFD1Y9 (g.165A>G) and SRY4 (g.104G>A) loci, which can define three Y‐haplotypes (YH1, YH2 and YH3) in yaks, were discovered. YH1 was the dominant and presumably most ancient haplotype based on the comparison of UTY19 locus with other bovid species. Interestingly, we found informative UTY19 markers (g.158A>C and g.169C>T) that can effectively distinguish the three yak Y‐haplotypes. The nucleotide diversity was 1.7 × 10^?4 ± 0.3 × 10^?4, indicating rich Y‐chromosome diversity in yaks. We identified two highly divergent lineages (YH1 and YH2 vs. YH3) that share similar frequencies (YH1 +  YH2: 0.82–0.89, YH3: 0.11–0.18) among all three populations. In agreement with previous mtDNA studies, we supported the hypothesis that the two highly divergent lineages (YH1 and YH2 vs. YH3) derived from a single gene pool, which can be explained by the reunion of at least two paternal populations with the divergent lineages already accumulated before domestication. We estimated a divergence time of 408 110 years between the two divergent lineages, which is consistent with the data from mitochondrial DNA in yaks.     

Y-chromosome polymorphisms of the domestic Bactrian camel in China

Single-nucleotide polymorphisms (SNPs), microsatellites and copy number variation (CNV) were studied on the Y chromosome to understand the paternal origin and phylogenetic relationships for resource protection, rational development and utilization of the domestic Bactrian camel in China. Our sample set consisted of 94 Chinese domestic Bactrian camels from four regions (Inner Mongolia, Gansu, Qinghai and Xinjiang), we screened 29 Y-chromosome-specific loci for SNPs, analysed 40 bovine-derived microsatellite loci and measured CNVs of HSFY and SRY through Sanger sequencing, automated fluorescence-based microsatellite analysis and quantitative real-time PCR, respectively. A multicopy gene, SRY, was first found, and sequence variation was only detected in SRY in a screen of 29 loci in 13 DNA pools of individual camels. In addition, a TG repeat in the USP9Y gene was identified as the first polymorphic microsatellite in the camel Y chromosome, whereas microsatellite based on bovine sequences were not detected. The frequency of each allele varied among different populations. For the Nanjiang, Hexi and Alashan populations, a 243-bp allele was found. For the Sunite population, 241-bp, 243-bp and 247-bp alleles were detected, and the frequencies of these alleles were \(22.2\%\), \(44.5\%\) and \(33.3\%\), respectively; 241-bp and 243-bp alleles were found in other populations. Finally, CNVs in two Y-chromosomal genes were detected; CNV for HSFY and SRY ranged from 1 to 3 and from 1 to 9, respectively.     

An efficient method for measuring copy number variation applied to improvement of nematode resistance in soybean

Copy number variation (CNV) is implicated in important traits in multiple crop plants, but can be challenging to genotype using conventional methods. The Rhg1 locus of soybean, which confers resistance to soybean cyst nematode (SCN), is a CNV of multiple 31.2‐kb genomic units each containing four genes. Reliable, high‐throughput methods to quantify Rhg1 and other CNVs for selective breeding were developed. The CNV genotyping assay described here uses a homeologous gene copy within the paleopolyploid soybean genome to provide the internal control for a single‐tube TaqMan copy number assay. Using this assay, CNV in breeding populations can be tracked with high precision. We also show that extensive CNV exists within Fayette, a released, inbred SCN‐resistant soybean cultivar with a high copy number at Rhg1 derived from a single donor parent. Copy number at Rhg1 is therefore unstable within a released variety over a relatively small number of generations. Using this assay to select for individuals with altered copy number, plants were obtained with both increased copy number and increased SCN resistance relative to control plants. Thus, CNV genotyping technologies can be used as a new type of marker‐assisted selection to select for desirable traits in breeding populations, and to control for undesirable variation within cultivars.     

Microarray Analysis of Copy Number Variants on the Human Y Chromosome Reveals Novel and Frequent Duplications Overrepresented in Specific Haplogroups

Background

The human Y chromosome is almost always excluded from genome-wide investigations of copy number variants (CNVs) due to its highly repetitive structure. This chromosome should not be forgotten, not only for its well-known relevance in male fertility, but also for its involvement in clinical phenotypes such as cancers, heart failure and sex specific effects on brain and behaviour.

Results

We analysed Y chromosome data from Affymetrix 6.0 SNP arrays and found that the signal intensities for most of 8179 SNP/CN probes in the male specific region (MSY) discriminated between a male, background signals in a female and an isodicentric male containing a large deletion of the q-arm and a duplication of the p-arm of the Y chromosome. Therefore, this SNP/CN platform is suitable for identification of gain and loss of Y chromosome sequences. In a set of 1718 males, we found 25 different CNV patterns, many of which are novel. We confirmed some of these variants by PCR or qPCR. The total frequency of individuals with CNVs was 14.7%, including 9.5% with duplications, 4.5% with deletions and 0.7% exhibiting both. Hence, a novel observation is that the frequency of duplications was more than twice the frequency of deletions. Another striking result was that 10 of the 25 detected variants were significantly overrepresented in one or more haplogroups, demonstrating the importance to control for haplogroups in genome-wide investigations to avoid stratification. NO-M214(xM175) individuals presented the highest percentage (95%) of CNVs. If they were not counted, 12.4% of the rest included CNVs, and the difference between duplications (8.9%) and deletions (2.8%) was even larger.

Conclusions

Our results demonstrate that currently available genome-wide SNP platforms can be used to identify duplications and deletions in the human Y chromosome. Future association studies of the full spectrum of Y chromosome variants will demonstrate the potential involvement of gain or loss of Y chromosome sequence in different human phenotypes.     

Novel Y‐chromosome short tandem repeats in Sus scrofa and their variation in European wild boar and domestic pig populations

Y‐chromosome markers are important tools for studying male‐specific gene flow within and between populations, hybridization patterns and kinship. However, their use in non‐human mammals is often hampered by the lack of Y‐specific polymorphic markers. We identified new male‐specific short tandem repeats (STRs) in Sus scrofa using the available genome sequence. We selected four polymorphic loci (5–10 alleles per locus), falling in one duplicated and two single‐copy regions. A total of 32 haplotypes were found by screening 211 individuals from eight wild boar populations across Europe and five domestic pig populations. European wild boar were characterized by significantly higher levels of haplotype diversity compared to European domestic pigs (H_D = 0.904 ± 0.011 and H_D = 0.491 ± 0.077 respectively). Relationships among STR haplotypes were investigated by combining them with single nucleotide polymorphisms at two linked genes (AMELY and UTY) in a network analysis. A differentiation between wild and domestic populations was observed (F_ST = 0.229), with commercial breeds sharing no Y haplotype with the sampled wild boar. Similarly, a certain degree of geographic differentiation was observed across Europe, with a number of local private haplotypes and high diversity in northern populations. The described Y‐chromosome markers can be useful to track male inheritance and gene flow in wild and domestic populations, promising to provide insights into evolutionary and population genetics in Sus scrofa.     

Genome-wide detection of copy number variants in European autochthonous and commercial pig breeds by whole-genome sequencing of DNA pools identified breed-characterising copy number states

In this study, we identified copy number variants (CNVs) in 19 European autochthonous pig breeds and in two commercial breeds (Italian Large White and Italian Duroc) that represent important genetic resources for this species. The genome of 725 pigs was sequenced using a breed-specific DNA pooling approach (30–35 animals per pool) obtaining an average depth per pool of 42×. This approach maximised CNV discovery as well as the related copy number states characterising, on average, the analysed breeds. By mining more than 17.5 billion reads, we identified a total of 9592 CNVs (~683 CNVs per breed) and 3710 CNV regions (CNVRs; 1.15% of the reference pig genome), with an average of 77 CNVRs per breed that were considered as private. A few CNVRs were analysed in more detail, together with other information derived from sequencing data. For example, the CNVR encompassing the KIT gene was associated with coat colour phenotypes in the analysed breeds, confirming the role of the multiple copies in determining breed-specific coat colours. The CNVR covering the MSRB3 gene was associated with ear size in most breeds. The CNVRs affecting the ELOVL6 and ZNF622 genes were private features observed in the Lithuanian Indigenous Wattle and in the Turopolje pig breeds respectively. Overall, the genome variability unravelled here can explain part of the genetic diversity among breeds and might contribute to explain their origin, history and adaptation to a variety of production systems.     

Sex‐chromosome differentiation parallels postglacial range expansion in European tree frogs (Hyla arborea)

Occasional XY recombination is a proposed explanation for the sex‐chromosome homomorphy in European tree frogs. Numerous laboratory crosses, however, failed to detect any event of male recombination, and a detailed survey of NW‐European Hyla arborea populations identified male‐specific alleles at sex‐linked loci, pointing to the absence of XY recombination in their recent history. Here, we address this paradox in a phylogeographic framework by genotyping sex‐linked microsatellite markers in populations and sibships from the entire species range. Contrasting with postglacial populations of NW Europe, which display complete absence of XY recombination and strong sex‐chromosome differentiation, refugial populations of the southern Balkans and Adriatic coast show limited XY recombination and large overlaps in allele frequencies. Geographically and historically intermediate populations of the Pannonian Basin show intermediate patterns of XY differentiation. Even in populations where X and Y occasionally recombine, the genetic diversity of Y haplotypes is reduced below the levels expected from the fourfold drop in copy numbers. This study is the first in which X and Y haplotypes could be phased over the distribution range in a species with homomorphic sex chromosomes; it shows that XY‐recombination patterns may differ strikingly between conspecific populations, and that recombination arrest may evolve rapidly (<5000 generations).     

Evaluation of non‐invasive prenatal testing to detect chromosomal aberrations in a Chinese cohort

The aim of this study was to evaluate the clinical feasibility of non‐invasive prenatal testing (NIPT) to detect foetal copy number variations (CNVs). Next‐generation sequencing for detecting foetal copy number variations (CNVs) was performed on the collected samples from 161 pregnancies with ultrasound anomalies and negative NIPT results for aneuploidy. The performance of NIPT for detecting chromosome aberrations was calculated. The sensitivity and specificity of NIPT for detecting CNVs > 1 Mb were 83.33% and 99.34%; the PPV and negative predictive rate (NPV) were 90.91% and 98.68%. Non‐invasive prenatal testing can be performed to detect chromosomal aberrations in first trimester with high performance for CNVs, and occasional discordant cases are unavoidable.     

Chromosomal location and gene paucity of the male specific region on papaya Y chromosome

Sex chromosomes in flowering plants evolved recently and many of them remain homomorphic, including those in papaya. We investigated the chromosomal location of papaya’s small male specific region of the hermaphrodite Y (Y^h) chromosome (MSY) and its genomic features. We conducted chromosome fluorescence in situ hybridization mapping of Y^h-specific bacterial artificial chromosomes (BACs) and placed the MSY near the centromere of the papaya Y chromosome. Then we sequenced five MSY BACs to examine the genomic features of this specialized region, which resulted in the largest collection of contiguous genomic DNA sequences of a Y chromosome in flowering plants. Extreme gene paucity was observed in the papaya MSY with no functional gene identified in 715 kb MSY sequences. A high density of retroelements and local sequence duplications were detected in the MSY that is suppressed for recombination. Location of the papaya MSY near the centromere might have provided recombination suppression and fostered paucity of genes in the male specific region of the Y chromosome. Our findings provide critical information for deciphering the sex chromosomes in papaya and reference information for comparative studies of other sex chromosomes in animals and plants. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Dmrt1 polymorphism and sex‐chromosome differentiation in Rana temporaria

Sex‐determination mechanisms vary both within and among populations of common frogs, opening opportunities to investigate the molecular pathways and ultimate causes shaping their evolution. We investigated the association between sex‐chromosome differentiation (as assayed from microsatellites) and polymorphism at the candidate sex‐determining gene Dmrt1 in two Alpine populations. Both populations harboured a diversity of X‐linked and Y‐linked Dmrt1 haplotypes. Some males had fixed male‐specific alleles at all markers (“differentiated” Y chromosomes), others only at Dmrt1 (“proto‐” Y chromosomes), while still others were genetically indistinguishable from females (undifferentiated X chromosomes). Besides these XX males, we also found rare XY females. The several Dmrt1 Y haplotypes differed in the probability of association with a differentiated Y chromosome, which we interpret as a result of differences in the masculinizing effects of alleles at the sex‐determining locus. From our results, the polymorphism in sex‐chromosome differentiation and its association with Dmrt1, previously inferred from Swedish populations, are not just idiosyncratic features of peripheral populations, but also characterize highly diverged populations in the central range. This implies that an apparently unstable pattern has been maintained over long evolutionary times.     

The Drosophila simulans Y chromosome interacts with the autosomes to influence male fitness

The Y chromosome should degenerate because it cannot recombine. However, male‐limited transmission increases selection efficiency for male‐benefit alleles on the Y, and therefore, Y chromosomes should contribute significantly to variation in male fitness. This means that although the Drosophila Y chromosome is small and gene‐poor, Y‐linked genes are vital for male fertility in Drosophila melanogaster and the Y chromosome has large male fitness effects. It is unclear whether the same pattern is seen in the closely related Drosophila simulans. We backcrossed Y chromosomes from three geographic locations into five genetic backgrounds and found strong Y and genetic background effects on male fertility. There was a significant Y‐background interaction, indicating substantial epistasis between the Y and autosomal genes affecting male fertility. This supports accumulating evidence that interactions between the Y chromosome and the autosomes are key determinants of male fitness.     

Copy number variations at the Prader-Willi syndrome region on chromosome 15 and associations with obesity in whites

Obesity is a serious health problem with strong genetic determination. Copy number variation (CNV) is a common type of genomic variant associated with some complex human diseases. However, it is not clear how CNVs contribute to the etiology of obesity. In this study, we examined 1,000 unrelated US whites to search for CNVs that may predispose to obesity. We focused our analyses on the Prader‐Willi syndrome (PWS) critical region (chromosome 15q11–q13), because the PWS region is a hotspot for CNV generation and obesity is one of the major clinical manifestations for chromosome abnormalities at this region. We constructed a map containing 39 CNVs at the PWS critical region with CNV occurrence rates higher than 1%. Among them, three CNVs were significantly associated with body fat mass (P < 0.05), with a higher copy number (CN) associated with an increase of 5.08–9.77 kg in body fat mass. These three CNVs are close to two known PWS genes, NDN (necdin homolog) and C15orf2 (chromosome 15 open reading frame 2), and partially overlap with another obesity gene PWRN1 (Prader‐Willi region nonprotein‐coding RNA 1). Interestingly, our recently published whole genome association scan study using the same sample by examining single‐nucleotide polymorphisms (SNPs) did not find any significant associations at these CNV regions, suggesting the importance of examining both CNVs and SNPs for better understanding of genetic basis of obesity. Further studies are warranted to validate these CNVs and their importance to obesity.     

Single‐copy gene‐based chromosome painting in cucumber and its application for chromosome rearrangement analysis in Cucumis

Chromosome painting based on fluorescence in situ hybridization (FISH) has played an important role in chromosome identification and research into chromosome rearrangements, diagnosis of chromosome abnormalities and evolution in human and animal species. However, it has not been applied widely in plants due to the large amounts of dispersed repetitive sequences in chromosomes. In the present work, a chromosome painting method for single‐copy gene pools in Cucumis sativus was successfully developed. Gene probes with sizes above 2 kb were detected consistently. A cucumber karyotype was constructed based on FISH using a cocktail containing chromosome‐specific gene probes. This single‐copy gene‐based chromosome painting (ScgCP) technique was performed by PCR amplification, purification, pooling, labeling and hybridization onto chromosome spreads. Gene pools containing sequential genes with an interval less than 300 kb yielded painting patterns on pachytene chromosomes. Seven gene pools corresponding to individual chromosomes unambiguously painted each chromosome pair of C. sativus. Three mis‐aligned regions on chromosome 4 were identified by the painting patterns. A probe pool comprising 133 genes covering the 8 Mb distal end of chromosome 4 was used to evaluate the potential utility of the ScgCP technique for chromosome rearrangement research through cross‐species FISH in the Cucumis genus. Distinct painting patterns of this region were observed in C. sativus, C. melo and C. metuliferus species. A comparative chromosome map of this region was constructed between cucumber and melon. With increasing sequence resources, this ScgCP technique may be applied on any other sequenced species for chromosome painting research.     

No evidence that Y‐chromosome differentiation affects male fitness in a Swiss population of common frogs

The canonical model of sex‐chromosome evolution assigns a key role to sexually antagonistic (SA) genes on the arrest of recombination and ensuing degeneration of Y chromosomes. This assumption cannot be tested in organisms with highly differentiated sex chromosomes, such as mammals or birds, owing to the lack of polymorphism. Fixation of SA alleles, furthermore, might be the consequence rather than the cause of recombination arrest. Here we focus on a population of common frogs (Rana temporaria) where XY males with genetically differentiated Y chromosomes (nonrecombinant Y haplotypes) coexist with both XY° males with proto‐Y chromosomes (only differentiated from X chromosomes in the immediate vicinity of the candidate sex‐determining locus Dmrt1) and XX males with undifferentiated sex chromosomes (genetically identical to XX females). Our study finds no effect of sex‐chromosome differentiation on male phenotype, mating success or fathering success. Our conclusions rejoin genomic studies that found no differences in gene expression between XY, XY° and XX males. Sexual dimorphism in common frogs might result more from the differential expression of autosomal genes than from sex‐linked SA genes. Among‐male variance in sex‐chromosome differentiation seems better explained by a polymorphism in the penetrance of alleles at the sex locus, resulting in variable levels of sex reversal (and thus of X‐Y recombination in XY females), independent of sex‐linked SA genes.     

Copy number variations and polymorphisms in HSP90AB1 and risk of systemic lupus erythematosus and efficacy of glucocorticoids

The aim of our study was to assess the associations of HSP90AB1 copy number variations (CNVs) with systemic lupus erythematosus (SLE) risk and glucocorticoids (GCs) efficacy, as well as the relationship between HSP90AB1 single‐nucleotide polymorphisms (SNPs) and GCs efficacy. HSP90AB1 CNVs and SLE risk were analysed in 519 patients and 538 controls. Patients treated with GCs were followed up for 12 weeks and were divided into sensitive and insensitive groups to investigate the effects of CNVs (419 patients) and SNPs (457 patients) on the efficacy of GCs. Health‐related quality of life (HRQoL) was also measured by SF‐36 at baseline and week 12 to explore the relationship between CNVs/SNPs and HRQoL improvements in Chinese SLE patients. Our results indicated a statistically significant association between HSP90AB1 CNVs and SLE (P_BH = 0.039), and this association was more pronounced in the female subgroup (P_BH = 0.039). However, we did not detect association of HSP90AB1 CNVs/SNPs with efficacy of GCs. But we found a marginal association between SNP rs13296 and improvement in Role‐emotional, while this association was not strong enough to survive in the multiple testing corrections. Collectively, our findings suggest that the copy number of HSP90AB1 is associated with SLE susceptibility. But copy number and polymorphisms of HSP90AB1 may not be associated with efficacy of GCs.