Expression analysis of NF1-mutated alleles in a rare compound heterozygous spinal NF1 patient by digital PCR.

BACKGROUD: Neurofibromatosis type 1 (NF1) is a heterogeneous neurocutaneous disorder. Spinal neurofibromatosis (SNF) is a distinct clinical entity of NF1, characterized by bilateral neurofibromas involving all spinal nerve roots. Although both forms are caused by intragenic heterozygous variants of NF1, missense variants have been associated with SNF, according to a dominant inheritance model causing haploinsufficiency. Most patients carry pathogenic variants in one of the NF1 alleles; nevertheless, patients with both NF1-mutated copies have been described. Interestingly, all NF1 variants carried by the known SNF compound heterozygotes were missense/splicing variants or in-frame insertion-deletions. AIMS: To investigate whether there is a differential expression of NF1 variant alleles in an NF1 compound heterozygous SNF patient possibly contributing to clinical phenotype. MATERIALS & METHODS: We performed an allele-specific expression study, by chip-based digital PCR, in an SNF family carrying two NF1 missense variants. We evaluated the expression levels of the two NF1-mutated alleles both carried by the compound heterozygous SNF patient and his relatives. RESULTS: Both alleles were expressed at comparable levels in the patient and hyper-expressed compared to the wild-type alleles of healthy controls. DISCUSSION: Here we provide new insights into expression studies of NF1-mutated transcripts suggesting that a novel pathogenetic mechanism, caused by gain-of-function variants, could be associated with SNF. CONCLUSIONS: Further studies should be performed in larger cohorts, opening new perspectives in the NF1 pathogenesis comprehension.

ZFP57 dictates allelic expression switch of target imprinted genes.

ZFP57 is a master regulator of genomic imprinting. It has both maternal and zygotic functions that are partially redundant in maintaining DNA methylation at some imprinting control regions (ICRs). In this study, we found that DNA methylation was lost at most known ICRs in Zfp57 mutant embryos. Furthermore, loss of ZFP57 caused loss of parent-of-origin-dependent monoallelic expression of the target imprinted genes. The allelic expression switch occurred in the ZFP57 target imprinted genes upon loss of differential DNA methylation at the ICRs in Zfp57 mutant embryos. Specifically, upon loss of ZFP57, the alleles of the imprinted genes located on the same chromosome with the originally methylated ICR switched their expression to mimic their counterparts on the other chromosome with unmethylated ICR. Consistent with our previous study, ZFP57 could regulate the NOTCH signaling pathway in mouse embryos by impacting allelic expression of a few regulators in the NOTCH pathway. In addition, the imprinted Dlk1 gene that has been implicated in the NOTCH pathway was significantly down-regulated in Zfp57 mutant embryos. Our allelic expression switch models apply to the examined target imprinted genes controlled by either maternally or paternally methylated ICRs. Our results support the view that ZFP57 controls imprinted expression of its target imprinted genes primarily through maintaining differential DNA methylation at the ICRs.

Random Allelic Expression in Inherited Retinal Disease Genes.

Inherited retinal diseases (IRDs) are a significant contributor to visual loss in children and young adults, falling second only to diabetic retinopathy. Understanding the pathogenic mechanisms of IRDs remains paramount. Some autosomal genes exhibit random allelic expression (RAE), similar to X-chromosome inactivation. This study identifies RAE genes in IRDs. Genes in the Retinal Information Network were cross-referenced with the recent literature to identify expression profiles, RAE, or biallelic expression (BAE). Loss-of-function intolerance (LOFI) was determined by cross-referencing the existing literature. Molecular and biological pathways that are significantly enriched were evaluated using gene ontology. A total of 184 IRD-causing genes were evaluated. Of these, 31 (16.8%) genes exhibited RAE. LOFI was exhibited in 6/31 (19.4%) of the RAE genes and 18/153 (11.8%) of the BAE genes. Brain tissue exhibited BAE in 107/128 (83.6%) genes for both sexes. The molecular pathways significantly enriched among BAE genes were photoreceptor activity, tubulin binding, and nucleotide/ribonucleotide binding. The biologic pathways significantly enriched for RAE genes were equilibrioception, parallel actin filament bundle assembly, photoreceptor cell outer segment organization, and protein depalmitoylation. Allele-specific expression may be a mechanism underlying IRD phenotypic variability, with clonal populations of embryologic precursor cells exhibiting RAE. Brain tissue preferentially exhibited BAE, possibly due to selective pressures against RAE. Pathways critical for cellular and visual function were enriched in BAE, which may offer a survival benefit.

Allelic expression imbalance in articular cartilage and subchondral bone refined genome-wide association signals in osteoarthritis.

OBJECTIVES: To present an unbiased approach to identify positional transcript single nucleotide polymorphisms (SNPs) of osteoarthritis (OA) risk loci by allelic expression imbalance (AEI) analyses using RNA sequencing of articular cartilage and subchondral bone from OA patients. METHODS: RNA sequencing from 65 articular cartilage and 24 subchondral bone from OA patients was used for AEI analysis. AEI was determined for all genes present in the 100 regions reported by the genome-wide association studies (GWAS) catalog that were also expressed in cartilage or bone. The count fraction of the alternative allele (φ) was calculated for each heterozygous individual with the risk SNP or with the SNP in linkage disequilibrium (LD) with it (r2 > 0.6). Furthermore, a meta-analysis was performed to generate a meta-φ (null hypothesis median φ = 0.49) and P-value for each SNP. RESULTS: We identified 30 transcript SNPs (28 in cartilage and two in subchondral bone) subject to AEI in 29 genes. Notably, 10 transcript SNPs were located in genes not previously reported in the GWAS catalog, including two long intergenic non-coding RNAs (lincRNAs), MALAT1 (meta-φ = 0.54, FDR = 1.7×10-4) and ILF3-DT (meta-φ = 0.6, FDR = 1.75×10-5). Moreover, 12 drugs were interacting with seven genes displaying AEI, of which seven drugs have been already approved. CONCLUSIONS: By prioritizing proxy transcript SNPs that mark AEI in cartilage and/or subchondral bone at loci harbouring GWAS signals, we present an unbiased approach to identify the most likely functional OA risk-SNP and gene. We identified 10 new potential OA risk genes ready for further translation towards underlying biological mechanisms.

Characterization of Genes That Exhibit Genotype-Dependent Allele-Specific Expression and Its Implications for the Development of Maize Kernel.

Heterosis or hybrid vigor refers to the superior phenotypic traits of hybrids relative to their parental inbred lines. An imbalance between the expression levels of two parental alleles in the F1 hybrid has been suggested as a mechanism of heterosis. Here, based on genome-wide allele-specific expression analysis using RNA sequencing technology, 1689 genes exhibiting genotype-dependent allele-specific expression (genotype-dependent ASEGs) were identified in the embryos, and 1390 genotype-dependent ASEGs in the endosperm, of three maize F1 hybrids. Of these ASEGs, most were consistent in different tissues from one hybrid cross, but nearly 50% showed allele-specific expression from some genotypes but not others. These genotype-dependent ASEGs were mostly enriched in metabolic pathways of substances and energy, including the tricarboxylic acid cycle, aerobic respiration, and energy derivation by oxidation of organic compounds and ADP binding. Mutation and overexpression of one ASEG affected kernel size, which indicates that these genotype-dependent ASEGs may make important contributions to kernel development. Finally, the allele-specific methylation pattern on genotype-dependent ASEGs indicated that DNA methylation plays a potential role in the regulation of allelic expression for some ASEGs. In this study, a detailed analysis of genotype-dependent ASEGs in the embryo and endosperm of three different maize F1 hybrids will provide an index of genes for future research on the genetic and molecular mechanism of heterosis.

Conservation Study of Imprinted Genes in Maize Triparental Heterozygotic Kernels.

Genomic imprinting is a classic epigenetic phenomenon related to the uniparental expression of genes. Imprinting variability exists in seeds and can contribute to observed parent-of-origin effects on seed development. Here, we conducted allelic expression of the embryo and endosperm from four crosses at 11 days after pollination (DAP). First, the F1 progeny of B73(♀) × Mo17(♂) and the inducer line CAU5 were used as parents to obtain reciprocal crosses of BM-C/C-BM. Additionally, the F1 progeny of Mo17(♀) × B73(♂) and CAU5 were used as parents to obtain reciprocal crosses of MB-C/C-MB. In total, 192 and 181 imprinted genes were identified in the BM-C/C-BM and MB-C/C-MB crosses, respectively. Then, by comparing the allelic expression of these imprinted genes in the reciprocal crosses of B73 and CAU5 (BC/CB), fifty-one Mo17-added non-conserved genes were identified as exhibiting imprinting variability. Fifty-one B73-added non-conserved genes were also identified by comparing the allelic expression of imprinted genes identified in BM-C/C-BM, MB-C/C-MB and MC/CM crosses. Specific Gene Ontology (GO) terms were not enriched in B73-added/Mo17-added non-conserved genes. Interestingly, the imprinting status of these genes was less conserved across other species. The cis-element distribution, tissue expression and subcellular location were similar between the B73-added/Mo17-added conserved and B73-added/Mo17-added non-conserved imprinted genes. Finally, genotypic and phenotypic analysis of one non-conserved gene showed that the mutation and overexpression of this gene may affect embryo and kernel size, which indicates that these non-conserved genes may also play an important role in kernel development. The findings of this study will be helpful for elucidating the imprinting mechanism of genes involved in maize kernel development.

The Defects of Epigenetic Reprogramming in Dox-Dependent Porcine-iPSCs.

Porcine-induced pluripotent stem cells (piPSCs) are of great significance to animal breeding and human medicine; however, an important problem is that the maintenance of piPSCs mainly depends on exogenous expression of pluripotent transcription factors (TFs), and germline transmission-competent piPSCs have not yet been successfully established. In this study, we explore the defect of epigenetic reprogramming during piPSCs formation, including chromatin accessibility, DNA methylation, and imprinted gene expression, with high-throughput sequencing (ATAC-seq, WGBS, RNA-seq, and Re-seq) methods. We found the somatic features were successfully silenced by connecting closed chromatin loci with downregulated genes, while DNA methylation has limited effects on somatic silence. However, the incomplete chromatin remodeling and DNA demethylation in pluripotency genes hinder pluripotent activation, resulting in the low expression of endogenous pluripotency genes. In addition, the expression of potential imprinted genes was abnormal, and many allelic-biased expressed genes in porcine embryonic fibroblasts (PEFs) were erased, accompanied by establishment of new allelic-biased expressed genes in piPSCs. This study reveals the aberrant epigenetic reprogramming during dox-dependent piPSCs formation, which lays the foundation for research of porcine-iPSC reprogramming and genome imprinting.

Identification of Clinical Variants beyond the Exome in Inborn Errors of Metabolism.

Inborn errors of metabolism (IEM) constitute a huge group of rare diseases affecting 1 in every 1000 newborns. Next-generation sequencing has transformed the diagnosis of IEM, leading to its proposed use as a second-tier technology for confirming cases detected by clinical/biochemical studies or newborn screening. The diagnosis rate is, however, still not 100%. This paper reports the use of a personalized multi-omics (metabolomic, genomic and transcriptomic) pipeline plus functional genomics to aid in the genetic diagnosis of six unsolved cases, with a clinical and/or biochemical diagnosis of galactosemia, mucopolysaccharidosis type I (MPS I), maple syrup urine disease (MSUD), hyperphenylalaninemia (HPA), citrullinemia, or urea cycle deficiency. Eight novel variants in six genes were identified: six (four of them deep intronic) located in GALE, IDUA, PTS, ASS1 and OTC, all affecting the splicing process, and two located in the promoters of IDUA and PTS, thus affecting these genes' expression. All the new variants were subjected to functional analysis to verify their pathogenic effects. This work underscores how the combination of different omics technologies and functional analysis can solve elusive cases in clinical practice.

Integrative genome-wide analyses reveal the transcriptional aberrations in Japanese esophageal squamous cell carcinoma.

Esophageal squamous cell carcinoma (ESCC) is a malignant disease. At present, the genomic profiles of ESCC are known to a considerable extent, and DNA methylation and gene expression profiles have been mainly used for the classification of ESCC subtypes, but integrative genomic, transcriptomic, and epigenomic analyses remain insufficient. Therefore, we performed integrative analyses using whole-exome sequencing, DNA methylation, and RNA sequencing (RNA-seq) analyses of Japanese patients with ESCC. In cancer-related genes, such as NOTCH family genes, RTK/PI3K pathway genes, and NFE2L2 pathway genes, variants and copy number amplification were detected frequently. Japanese ESCC cases were clustered into two mutational signatures: an APOBEC-associated signature and an age-related signature. In imprinted genes, DNA methylation was aberrant in gene promoter regions and correlated well with gene expression profiles. Nonsynonymous single-nucleotide variants and allelic expression imbalance were detected frequently in FAT family genes. Our integrative genome-wide analyses, including DNA methylation and allele-specific gene expression profiles, revealed altered gene regulation of imprinted genes and FAT family genes in ESCC.

Brain-specific monoallelic expression of bovine UBE3A is associated with genomic position.

Genomic imprinting is a rare epigenetic process in mammalian cells that leads to monoallelic expression of a gene with a parent-specific pattern. The UBE3A (ubiquitin protein ligase E3A) gene is imprinted with maternal allelic expression in the brain but biallelically expressed in all other tissues in humans. The silencing of the paternal UBE3A allele is thought to be caused by the paternally expressed antisense RNA transcript of UBE3A-ATS. The aberrant imprinted expression of the UBE3A is associated with several neurodevelopmental syndromes and psychological disorders. Cattle are a valuable model species in determining the genetic etiology of sporadic human disorder, and maternal expression of UEB3A has been revealed by next-generation sequencing study in the bovine conceptus. In this study, we investigated the allelic expression of UBE3A and UBE3A-ATS in adult bovine somatic tissues. To confirm the splicing pattern of bovine UBE3A, five 5' alternative transcripts (MT210534-MT210538) were first obtained from bovine brain tissue by RT-PCR. Based on 10 SNP genotypes, we found that the brain-specific monoallelic expression of bovine UBE3A did not occur along the entire locus, and there was a shift from biallelic expression to monoallelic expression in exon 14 of the UBE3A gene. However, the brain-specific monoallelic expression of bovine UBE3A-ATS occurred in the entire gene. These observations demonstrated that the monoallelic expression did not occur along the bovine UBE3A entire locus and was associated with the genomic position.

Haplotype-resolved genomes of geminivirus-resistant and geminivirus-susceptible African cassava cultivars.

BACKGROUND: Cassava is an important food crop in tropical and sub-tropical regions worldwide. In Africa, cassava production is widely affected by cassava mosaic disease (CMD), which is caused by the African cassava mosaic geminivirus that is transmitted by whiteflies. Cassava breeders often use a single locus, CMD2, for introducing CMD resistance into susceptible cultivars. The CMD2 locus has been genetically mapped to a 10-Mbp region, but its organization and genes as well as their functions are unknown. RESULTS: We report haplotype-resolved de novo assemblies and annotations of the genomes for the African cassava cultivar TME (tropical Manihot esculenta), which is the origin of CMD2, and the CMD-susceptible cultivar 60444. The assemblies provide phased haplotype information for over 80% of the genomes. Haplotype comparison identified novel features previously hidden in collapsed and fragmented cassava genomes, including thousands of allelic variants, inter-haplotype diversity in coding regions, and patterns of diversification through allele-specific expression. Reconstruction of the CMD2 locus revealed a highly complex region with nearly identical gene sets but limited microsynteny between the two cultivars. CONCLUSIONS: The genome maps of the CMD2 locus in both 60444 and TME3, together with the newly annotated genes, will help the identification of the causal genetic basis of CMD2 resistance to geminiviruses. Our de novo cassava genome assemblies will also facilitate genetic mapping approaches to narrow the large CMD2 region to a few candidate genes for better informed strategies to develop robust geminivirus resistance in susceptible cassava cultivars.

Integrated Transcriptomic and Proteomic Analysis of Primary Human Umbilical Vein Endothelial Cells.

Understanding the molecular profile of every human cell type is essential for understanding its role in normal physiology and disease. Technological advancements in DNA sequencing, mass spectrometry, and computational methods allow us to carry out multiomics analyses although such approaches are not routine yet. Human umbilical vein endothelial cells (HUVECs) are a widely used model system to study pathological and physiological processes associated with the cardiovascular system. In this study, next-generation sequencing and high-resolution mass spectrometry to profile the transcriptome and proteome of primary HUVECs is employed. Analysis of 145 million paired-end reads from next-generation sequencing confirmed expression of 12 186 protein-coding genes (FPKM ≥0.1), 439 novel long non-coding RNAs, and revealed 6089 novel isoforms that were not annotated in GENCODE. Proteomics analysis identifies 6477 proteins including confirmation of N-termini for 1091 proteins, isoforms for 149 proteins, and 1034 phosphosites. A database search to specifically identify other post-translational modifications provide evidence for a number of modification sites on 117 proteins which include ubiquitylation, lysine acetylation, and mono-, di- and tri-methylation events. Evidence for 11 "missing proteins," which are proteins for which there was insufficient or no protein level evidence, is provided. Peptides supporting missing protein and novel events are validated by comparison of MS/MS fragmentation patterns with synthetic peptides. Finally, 245 variant peptides derived from 207 expressed proteins in addition to alternate translational start sites for seven proteins and evidence for novel proteoforms for five proteins resulting from alternative splicing are identified. Overall, it is believed that the integrated approach employed in this study is widely applicable to study any primary cell type for deeper molecular characterization.

Differential expression networks and inheritance patterns of long non-coding RNAs in castor bean seeds.

Long non-coding RNAs (lncRNAs) serve as versatile regulators of plant growth and development. The potential functions and inheritance patterns of lncRNAs, as well as the epigenetic regulation of lncRNA itself, remain largely uncharacterized in plant seeds, especially in the persistent endosperm of the dicotyledons. In this study, we investigated diverse RNA-seq data and catalogued 5356 lncRNAs in castor bean seeds. A small fraction of lncRNAs were transcribed from the same direction as the promoters of protein-coding genes (PCgenes) and exhibited strongly coordinated expression with the nearby PCgene. Co-expression analysis with weighted gene co-expression network analysis (WGCNA) showed these lncRNAs to be involved in differential transcription networks between the embryo and endosperm in the early developing seed. Genomic DNA methylation analyses revealed that the expression level of lncRNAs was tightly linked to DNA methylation and that endosperm hypomethylation could promote the expression of linked lncRNAs. Intriguingly, upon hybridization, most lncRNAs with divergent genome sequences between two parents could be reconciled and were expressed according to their parental genome contribution; however, some deviation in the expression of allelic lncRNAs was observed and found to be partially dependent on parental effects. In triploid endosperm, the expression of most lncRNAs was not dosage sensitive, as only 20 lncRNAs had balanced dosage. Our findings not only demonstrate that lncRNAs play potential roles in regulating the development of castor bean endosperm and embryo, but also provide novel insights into the parental effects, allelic expression and epigenetic regulation of lncRNAs in dicotyledonous seeds.

Effects of parental genetic divergence on gene expression patterns in interspecific hybrids of Camellia.

BACKGROUND: The merging of two divergent genomes during hybridization can result in the remodeling of parental gene expression in hybrids. A molecular basis underling expression change in hybrid is regulatory divergence, which may change with the parental genetic divergence. However, there still no unanimous conclusion for this hypothesis. RESULTS: Three species of Camellia with a range of genetic divergence and their F1 hybrids were used to study the effect of parental genetic divergence on gene expression and regulatory patterns in hybrids by RNA-sequencing and allelic expression analysis. We found that though the proportion of differentially expressed genes (DEGs) between the hybrids and their parents did not increase, a greater proportion of DEGs would be non-additively (especially transgressively) expressed in the hybrids as genomes between the parents become more divergent. In addition, the proportion of genes with significant evidence of cis-regulatory divergence increased, whereas with trans-regulatory divergence decreased with parental genetic divergence. CONCLUSIONS: The discordance within hybrid would intensify as the parents become more divergent, manifesting as more DEGs would be non-additively expressed. Trans-regulatory divergence contributed more to the additively inherited genes than cis, however, its contribution to expression difference would be weakened as cis mutations accumulated over time; and this might be an important reason for that the more divergent the parents are, the greater proportion of DEGs would be non-additively expressed in hybrid.

Erasure and reestablishment of random allelic expression imbalance after epigenetic reprogramming.

Clonal level random allelic expression imbalance and random monoallelic expression provides cellular heterogeneity within tissues by modulating allelic dosage. Although such expression patterns have been observed in multiple cell types, little is known about when in development these stochastic allelic choices are made. We examine allelic expression patterns in human neural progenitor cells before and after epigenetic reprogramming to induced pluripotency, observing that loci previously characterized by random allelic expression imbalance (0.63% of expressed genes) are generally reset to a biallelic state in induced pluripotent stem cells (iPSCs). We subsequently neuralized the iPSCs and profiled isolated clonal neural stem cells, observing that significant random allelic expression imbalance is reestablished at 0.65% of expressed genes, including novel loci not found to show allelic expression imbalance in the original parental neural progenitor cells. Allelic expression imbalance was associated with altered DNA methylation across promoter regulatory regions, with clones characterized by skewed allelic expression being hypermethylated compared to their biallelic sister clones. Our results suggest that random allelic expression imbalance is established during lineage commitment and is associated with increased DNA methylation at the gene promoter.

Genome analysis of Diploscapter coronatus: insights into molecular peculiarities of a nematode with parthenogenetic reproduction.

BACKGROUND: Sexual reproduction involving the fusion of egg and sperm is prevailing among eukaryotes. In contrast, the nematode Diploscapter coronatus, a close relative of the model Caenorhabditis elegans, reproduces parthenogenetically. Neither males nor sperm have been observed and some steps of meiosis are apparently skipped in this species. To uncover the genomic changes associated with the evolution of parthenogenesis in this nematode, we carried out a genome analysis. RESULTS: We obtained a 170 Mbp draft genome in only 511 scaffolds with a N50 length of 1 Mbp. Nearly 90% of these scaffolds constitute homologous pairs with a 5.7% heterozygosity on average and inversions and translocations, meaning that the 170 Mbp sequences correspond to the diploid genome. Fluorescent staining shows that the D. coronatus genome consists of two chromosomes (2n = 2). In our genome annotation, we found orthologs of 59% of the C. elegans genes. However, a number of genes were missing or very divergent. These include genes involved in sex determination (e.g. xol-1, tra-2) and meiosis (e.g. the kleisins rec-8 and coh-3/4) giving a possible explanation for the absence of males and the second meiotic division. The high degree of heterozygosity allowed us to analyze the expression level of individual alleles. Most of the homologous pairs show very similar expression levels but others exhibit a 2-5-fold difference. CONCLUSIONS: Our high-quality draft genome of D. coronatus reveals the peculiarities of the genome of parthenogenesis and provides some clues to the genetic basis for parthenogenetic reproduction. This draft genome should be the basis to elucidate fundamental questions related to parthenogenesis such as its origin and mechanisms through comparative analyses with other nematodes. Furthermore, being the closest outgroup to the genus Caenorhabditis, the draft genome will help to disclose many idiosyncrasies of the model C. elegans and its congeners in future studies.

Conclusive evidence for hexasomic inheritance in chrysanthemum based on analysis of a 183 k SNP array.

BACKGROUND: Cultivated chrysanthemum is an outcrossing hexaploid (2n = 6× = 54) with a disputed mode of inheritance. In this paper, we present a single nucleotide polymorphism (SNP) selection pipeline that was used to design an Affymetrix Axiom array with 183 k SNPs from RNA sequencing data (1). With this array, we genotyped four bi-parental populations (with sizes of 405, 53, 76 and 37 offspring plants respectively), and a cultivar panel of 63 genotypes. Further, we present a method for dosage scoring in hexaploids from signal intensities of the array based on mixture models (2) and validation of selection steps in the SNP selection pipeline (3). The resulting genotypic data is used to draw conclusions on the mode of inheritance in chrysanthemum (4), and to make an inference on allelic expression bias (5). RESULTS: With use of the mixture model approach, we successfully called the dosage of 73,936 out of 183,130 SNPs (40.4%) that segregated in any of the bi-parental populations. To investigate the mode of inheritance, we analysed markers that segregated in the large bi-parental population (n = 405). Analysis of segregation of duplex x nulliplex SNPs resulted in evidence for genome-wide hexasomic inheritance. This evidence was substantiated by the absence of strong linkage between markers in repulsion, which indicated absence of full disomic inheritance. We present the success rate of SNP discovery out of RNA sequencing data as affected by different selection steps, among which SNP coverage over genotypes and use of different types of sequence read mapping software. Genomic dosage highly correlated with relative allele coverage from the RNA sequencing data, indicating that most alleles are expressed according to their genomic dosage. CONCLUSIONS: The large population, genotyped with a very large number of markers, is a unique framework for extensive genetic analyses in hexaploid chrysanthemum. As starting point, we show conclusive evidence for genome-wide hexasomic inheritance.

Association of breast cancer risk in BRCA1 and BRCA2 mutation carriers with genetic variants showing differential allelic expression: identification of a modifier of breast cancer risk at locus 11q22.3.

PURPOSE: Cis-acting regulatory SNPs resulting in differential allelic expression (DAE) may, in part, explain the underlying phenotypic variation associated with many complex diseases. To investigate whether common variants associated with DAE were involved in breast cancer susceptibility among BRCA1 and BRCA2 mutation carriers, a list of 175 genes was developed based of their involvement in cancer-related pathways. METHODS: Using data from a genome-wide map of SNPs associated with allelic expression, we assessed the association of ~320 SNPs located in the vicinity of these genes with breast and ovarian cancer risks in 15,252 BRCA1 and 8211 BRCA2 mutation carriers ascertained from 54 studies participating in the Consortium of Investigators of Modifiers of BRCA1/2. RESULTS: We identified a region on 11q22.3 that is significantly associated with breast cancer risk in BRCA1 mutation carriers (most significant SNP rs228595 p = 7 × 10-6). This association was absent in BRCA2 carriers (p = 0.57). The 11q22.3 region notably encompasses genes such as ACAT1, NPAT, and ATM. Expression quantitative trait loci associations were observed in both normal breast and tumors across this region, namely for ACAT1, ATM, and other genes. In silico analysis revealed some overlap between top risk-associated SNPs and relevant biological features in mammary cell data, which suggests potential functional significance. CONCLUSION: We identified 11q22.3 as a new modifier locus in BRCA1 carriers. Replication in larger studies using estrogen receptor (ER)-negative or triple-negative (i.e., ER-, progesterone receptor-, and HER2-negative) cases could therefore be helpful to confirm the association of this locus with breast cancer risk.

Allelic expression imbalance polymorphisms in susceptibility chromosome regions and the risk and survival of breast cancer.

Allelic expression imbalance (AEI) has been applied to indicate potential function of genetic variants. Combining earlier results from global differential allele-specific expression analysis and genome wide association studies (GWASs), we select the single nuclear polymorphisms (SNPs) exhibiting AEI phenomenon located in breast cancer susceptibility chromosome regions, and evaluate their associations with breast cancer risk and survival. We examined the genotypes of 10 AEI SNPs in 1551 incident breast cancer cases and 1605 age-frequency matched controls from Guangzhou, China. In total, 1168 cases were followed up. MUC16 rs2591592 (AT/AA vs. TT) was associated with an increased risk of premenopausal breast cancer (OR [95%CI]: 1.30 [1.07, 1.57]); SLAMF1 rs1061217 (CT/TT vs. CC) decreased the risk of breast cancer among overweight women (OR [95%CI]: 0.74 [0.57, 0.96]) but increased the risk among normal-weight women (OR [95%CI]: 1.15 [1.01, 1.39]); ZNF331 rs8109631 (AG/AA vs. GG) and CHRAC1 rs10216653 (GC/GG vs. CC) were associated with progression free survival among breast cancer patients with negative ER/PR status and higher clinical stage (HRs [95%CIs]: 2.39 [1.14, 5.00], 1.85 [1.03, 3.32], and 0.49 [0.30, 0.80], respectively). ZNF331 rs8109631 and CHRAC1 rs10216653 were further found to represent several functional SNPs through bioinformatic analysis. In conclusion, our findings demonstrated suggestive associations of AEI polymorphisms with breast cancer risk (MUC16 rs2591592 and SLAMF1 rs1061217) and prognosis (ZNF331 rs8109631 and CHRAC1 rs10216653). © 2016 Wiley Periodicals, Inc.

The abundance of cis-acting loci leading to differential allele expression in F1 mice and their relationship to loci harboring genes affecting complex traits.

BACKGROUND: Genome-wide surveys have detected cis-acting quantitative trait loci altering levels of RNA transcripts (RNA-eQTLs) by associating SNV alleles to transcript levels. However, the sensitivity and specificity of detection of cis- expression quantitative trait loci (eQTLs) by genetic approaches, reliant as it is on measurements of transcript levels in recombinant inbred strains or offspring from arranged crosses, is unknown, as is their relationship to QTL's for complex phenotypes. RESULTS: We used transcriptome-wide differential allele expression (DAE) to detect cis-eQTLs in forebrain and kidney from reciprocal crosses between three mouse inbred strains, 129S1/SvlmJ, DBA/2J, and CAST/EiJ and C57BL/6 J. Two of these crosses were previously characterized for cis-eQTLs and QTLs for various complex phenotypes by genetic analysis of recombinant inbred (RI) strains. 5.4 %, 1.9 % and 1.5 % of genes assayed in forebrain of B6/129SF1, B6/DBAF1, and B6/CASTF1 mice, respectively, showed differential allelic expression, indicative of cis-acting alleles at these genes. Moreover, the majority of DAE QTLs were observed to be tissue-specific with only a small fraction showing cis-effects in both tissues. Comparing DAE QTLs in F1 mice to cis-eQTLs previously mapped in RI strains we observed that many of the cis-eQTLs were not confirmed by DAE. Additionally several novel DAE-QTLs not identified as cis-eQTLs were identified suggesting that there are differences in sensitivity and specificity for QTL detection between the two methodologies. Strain specific DAE QTLs in B6/DBAF1 mice were located in excess at candidate genes for alcohol use disorders, seizures, and angiogenesis previously implicated by genetic linkage in C57BL/6J × DBA/2JF2 mice or BXD RI strains. CONCLUSIONS: Via a survey for differential allele expression in F1 mice, a substantial proportion of genes were found to have alleles altering expression in cis-acting fashion. Comparing forebrain and kidney, many or most of these alleles were tissue-specific in action. The identification of strain specific DAE QTLs, can assist in assessment of candidate genes located within the large intervals associated with trait QTLs.