Deep molecular phenotypes link complex disorders and physiological insult to CpG methylation.

Epigenetic regulation of cellular function provides a mechanism for rapid organismal adaptation to changes in health, lifestyle and environment. Associations of cytosine-guanine di-nucleotide (CpG) methylation with clinical endpoints that overlap with metabolic phenotypes suggest a regulatory role for these CpG sites in the body's response to disease or environmental stress. We previously identified 20 CpG sites in an epigenome-wide association study (EWAS) with metabolomics that were also associated in recent EWASs with diabetes-, obesity-, and smoking-related endpoints. To elucidate the molecular pathways that connect these potentially regulatory CpG sites to the associated disease or lifestyle factors, we conducted a multi-omics association study including 2474 mass-spectrometry-based metabolites in plasma, urine and saliva, 225 NMR-based lipid and metabolite measures in blood, 1124 blood-circulating proteins using aptamer technology, 113 plasma protein N-glycans and 60 IgG-glyans, using 359 samples from the multi-ethnic Qatar Metabolomics Study on Diabetes (QMDiab). We report 138 multi-omics associations at these CpG sites, including diabetes biomarkers at the diabetes-associated TXNIP locus, and smoking-specific metabolites and proteins at multiple smoking-associated loci, including AHRR. Mendelian randomization suggests a causal effect of metabolite levels on methylation of obesity-associated CpG sites, i.e. of glycerophospholipid PC(O-36: 5), glycine and a very low-density lipoprotein (VLDL-A) on the methylation of the obesity-associated CpG loci DHCR24, MYO5C and CPT1A, respectively. Taken together, our study suggests that multi-omics-associated CpG methylation can provide functional read-outs for the underlying regulatory response mechanisms to disease or environmental insults.

Preferential Allele Expression Analysis Identifies Shared Germline and Somatic Driver Genes in Advanced Ovarian Cancer.

Identifying genes where a variant allele is preferentially expressed in tumors could lead to a better understanding of cancer biology and optimization of targeted therapy. However, tumor sample heterogeneity complicates standard approaches for detecting preferential allele expression. We therefore developed a novel approach combining genome and transcriptome sequencing data from the same sample that corrects for sample heterogeneity and identifies significant preferentially expressed alleles. We applied this analysis to epithelial ovarian cancer samples consisting of matched primary ovary and peritoneum and lymph node metastasis. We find that preferentially expressed variant alleles include germline and somatic variants, are shared at a relatively high frequency between patients, and are in gene networks known to be involved in cancer processes. Analysis at a patient level identifies patient-specific preferentially expressed alleles in genes that are targets for known drugs. Analysis at a site level identifies patterns of site specific preferential allele expression with similar pathways being impacted in the primary and metastasis sites. We conclude that genes with preferentially expressed variant alleles can act as cancer drivers and that targeting those genes could lead to new therapeutic strategies.

Mutation in noncoding RNA RNU12 causes early onset cerebellar ataxia.

OBJECTIVE: Exome sequences account for only 2% of the genome and may overlook mutations causing disease. To obtain a more complete view, whole genome sequencing (WGS) was analyzed in a large consanguineous family in which members displayed autosomal recessively inherited cerebellar ataxia manifesting before 2 years of age. METHODS: WGS from blood-derived genomic DNA was used for homozygosity mapping and a rare variant search. RNA from isolated blood leukocytes was used for quantitative polymerase chain reaction (PCR), RNA sequencing, and comparison of the transcriptomes of affected and unaffected family members. RESULTS: WGS revealed a point mutation in noncoding RNA RNU12 that was associated with early onset cerebellar ataxia. The U12-dependent minor spliceosome edits 879 known transcripts. Reverse transcriptase PCR demonstrated minor intron retention in all of 9 randomly selected RNAs from this group, and RNAseq showed splicing disruption specific to all U12-type introns detected in blood monocytes from affected individuals. Moreover, 144 minor intron-containing RNAs were differentially expressed, including transcripts for 3 genes previously associated with cerebellar neurodegeneration. INTERPRETATION: Interference with particular spliceosome components, including small nuclear RNAs, cause reproducible uniquely distributed phenotypic and transcript-specific effects, making this an important category of disease-associated mutation. Our approach to differential expression analysis of minor intron-containing genes is applicable to other diseases involving altered transcriptome processing. ANN NEUROL 2017;81:68-78.

A first genetic map of date palm (Phoenix dactylifera) reveals long-range genome structure conservation in the palms.

BACKGROUND: The date palm is one of the oldest cultivated fruit trees. It is critical in many ways to cultures in arid lands by providing highly nutritious fruit while surviving extreme heat and environmental conditions. Despite its importance from antiquity, few genetic resources are available for improving the productivity and development of the dioecious date palm. To date there has been no genetic map and no sex chromosome has been identified. RESULTS: Here we present the first genetic map for date palm and identify the putative date palm sex chromosome. We placed ~4000 markers on the map using nearly 1200 framework markers spanning a total of 1293 cM. We have integrated the genetic map, derived from the Khalas cultivar, with the draft genome and placed up to 19% of the draft genome sequence scaffolds onto linkage groups for the first time. This analysis revealed approximately ~1.9 cM/Mb on the map. Comparison of the date palm linkage groups revealed significant long-range synteny to oil palm. Analysis of the date palm sex-determination region suggests it is telomeric on linkage group 12 and recombination is not suppressed in the full chromosome. CONCLUSIONS: Based on a modified genotyping-by-sequencing approach we have overcome challenges due to lack of genetic resources and provide the first genetic map for date palm. Combined with the recent draft genome sequence of the same cultivar, this resource offers a critical new tool for date palm biotechnology, palm comparative genomics and a better understanding of sex chromosome development in the palms.

The genetic landscape of autism spectrum disorder in the Middle Eastern population.

Introduction: Autism spectrum disorder (ASD) is characterized by aberrations in social interaction and communication associated with repetitive behaviors and interests, with strong clinical heterogeneity. Genetic factors play an important role in ASD, but about 75% of ASD cases have an undetermined genetic risk. Methods: We extensively investigated an ASD cohort made of 102 families from the Middle Eastern population of Qatar. First, we investigated the copy number variations (CNV) contribution using genome-wide SNP arrays. Next, we employed Next Generation Sequencing (NGS) to identify de novo or inherited variants contributing to the ASD etiology and its associated comorbid conditions in families with complete trios (affected child and the parents). Results: Our analysis revealed 16 CNV regions located in genomic regions implicated in ASD. The analysis of the 88 ASD cases identified 41 genes in 39 ASD subjects with de novo (n = 24) or inherited variants (n = 22). We identified three novel de novo variants in new candidate genes for ASD (DTX4, ARMC6, and B3GNT3). Also, we have identified 15 de novo variants in genes that were previously implicated in ASD or related neurodevelopmental disorders (PHF21A, WASF1, TCF20, DEAF1, MED13, CREBBP, KDM6B, SMURF1, ADNP, CACNA1G, MYT1L, KIF13B, GRIA2, CHM, and KCNK9). Additionally, we defined eight novel recessive variants (RYR2, DNAH3, TSPYL2, UPF3B KDM5C, LYST, and WNK3), four of which were X-linked. Conclusion: Despite the ASD multifactorial etiology that hinders ASD genetic risk discovery, the number of identified novel or known putative ASD genetic variants was appreciable. Nevertheless, this study represents the first comprehensive characterization of ASD genetic risk in Qatar's Middle Eastern population.

Characterization of exercise-induced hemolysis in endurance horses.

Exercise-induced hemolysis occurs as the result of intense physical exercise and is caused by metabolic and mechanical factors including repeated muscle contractions leading to capillary vessels compression, vasoconstriction of internal organs and foot strike among others. We hypothesized that exercise-induced hemolysis occurred in endurance racehorses and its severity was associated with the intensity of exercise. To provide further insight into the hemolysis of endurance horses, the aim of the study was to deployed a strategy for small molecules (metabolites) profiling, beyond standard molecular methods. The study included 47 Arabian endurance horses competing for either 80, 100, or 120 km distances. Blood plasma was collected before and after the competition and analyzed macroscopically, by ELISA and non-targeted metabolomics with liquid chromatography-mass spectrometry. A significant increase in all hemolysis parameters was observed after the race, and an association was found between the measured parameters, average speed, and distance completed. Levels of hemolysis markers were highest in horses eliminated for metabolic reasons in comparison to finishers and horses eliminated for lameness (gait abnormality), which may suggest a connection between the intensity of exercise, metabolic challenges, and hemolysis. Utilization of omics methods alongside conventional methods revealed a broader insight into the exercise-induced hemolysis process by displaying, apart from commonly measured hemoglobin and haptoglobin, levels of hemoglobin degradation metabolites. Obtained results emphasized the importance of respecting horse limitations in regard to speed and distance which, if underestimated, may lead to severe damages.

DNA-based assays to distinguish date palm (Arecaceae) gender.

PREMISE OF THE STUDY: Date palm (Phoenix dactylifera) is one of the oldest cultivated trees and is critical to the development of arid land. The date palm is a dioecious monocot with separate male and female trees. This presents a challenge in development as it is impossible to distinguish trees until they flower approximately five to eight years after planting. METHODS AND RESULTS: We have developed PCR-based assays capable of sex differentiation in multiple date palm cultivars. The primers are designed across gender-specific polymorphisms and demonstrated greater than 90% accuracy in distinguishing date palm gender across multiple varieties. CONCLUSIONS: These results indicate that the primers should be helpful in rapidly distinguishing date palm gender from the earliest stages that DNA can safely be collected. This is a vast savings in time over present approaches.

Family-Based Genome-Wide Association Study of Autism Spectrum Disorder in Middle Eastern Families.

Autism spectrum disorder (ASD) is a neurodevelopmental disease characterized by abnormalities in language and social communication with substantial clinical heterogeneity. Genetic factors play an important role in ASD with heritability estimated between 70% to 80%. Genome-wide association studies (GWAS) have identified multiple loci associated with ASD. However, most studies were performed on European populations and little is known about the genetic architecture of ASD in Middle Eastern populations. Here, we report the first GWAS of ASD in the Middle eastern population of Qatar. We analyzed 171 families with ASD, using linear mixed models adjusting for relatedness and other confounders. Results showed that common single nucleotide polymorphisms (SNP) in seven loci are associated with ASD (p < 1 × 10-5). Although the identified loci did not reach genome-wide significance, many of the top associated SNPs are located within or near genes that have been implicated in ASD or related neurodevelopmental disorders. These include GORASP2, GABBR2, ANKS6, THSD4, ERCC6L, ARHGEF6, and HDAC8. Additionally, three of the top associated SNPs were significantly associated with gene expression. We also found evidence of association signals in two previously reported ASD-susceptibility loci (rs10099100 and rs4299400). Our results warrant further functional studies and replication to provide further insights into the genetic architecture of ASD.

Epigenetics meets proteomics in an epigenome-wide association study with circulating blood plasma protein traits.

DNA methylation and blood circulating proteins have been associated with many complex disorders, but the underlying disease-causing mechanisms often remain unclear. Here, we report an epigenome-wide association study of 1123 proteins from 944 participants of the KORA population study and replication in a multi-ethnic cohort of 344 individuals. We identify 98 CpG-protein associations (pQTMs) at a stringent Bonferroni level of significance. Overlapping associations with transcriptomics, metabolomics, and clinical endpoints suggest implication of processes related to chronic low-grade inflammation, including a network involving methylation of NLRC5, a regulator of the inflammasome, and associated pQTMs implicating key proteins of the immune system, such as CD48, CD163, CXCL10, CXCL11, LAG3, FCGR3B, and B2M. Our study links DNA methylation to disease endpoints via intermediate proteomics phenotypes and identifies correlative networks that may eventually be targeted in a personalized approach of chronic low-grade inflammation.

Connecting genetic risk to disease end points through the human blood plasma proteome.

Genome-wide association studies (GWAS) with intermediate phenotypes, like changes in metabolite and protein levels, provide functional evidence to map disease associations and translate them into clinical applications. However, although hundreds of genetic variants have been associated with complex disorders, the underlying molecular pathways often remain elusive. Associations with intermediate traits are key in establishing functional links between GWAS-identified risk-variants and disease end points. Here we describe a GWAS using a highly multiplexed aptamer-based affinity proteomics platform. We quantify 539 associations between protein levels and gene variants (pQTLs) in a German cohort and replicate over half of them in an Arab and Asian cohort. Fifty-five of the replicated pQTLs are located in trans. Our associations overlap with 57 genetic risk loci for 42 unique disease end points. We integrate this information into a genome-proteome network and provide an interactive web-tool for interrogations. Our results provide a basis for novel approaches to pharmaceutical and diagnostic applications.

A Genome-Wide Survey of Date Palm Cultivars Supports Two Major Subpopulations in Phoenix dactylifera.

The date palm (Phoenix dactylifera L.) is one of the oldest cultivated trees and is intimately tied to the history of human civilization. There are hundreds of commercial cultivars with distinct fruit shapes, colors, and sizes growing mainly in arid lands from the west of North Africa to India. The origin of date palm domestication is still uncertain, and few studies have attempted to document genetic diversity across multiple regions. We conducted genotyping-by-sequencing on 70 female cultivar samples from across the date palm-growing regions, including four Phoenix species as the outgroup. Here, for the first time, we generate genome-wide genotyping data for 13,000-65,000 SNPs in a diverse set of date palm fruit and leaf samples. Our analysis provides the first genome-wide evidence confirming recent findings that the date palm cultivars segregate into two main regions of shared genetic background from North Africa and the Arabian Gulf. We identify genomic regions with high densities of geographically segregating SNPs and also observe higher levels of allele fixation on the recently described X-chromosome than on the autosomes. Our results fit a model with two centers of earliest cultivation including date palms autochthonous to North Africa. These results adjust our understanding of human agriculture history and will provide the foundation for more directed functional studies and a better understanding of genetic diversity in date palm.

Exome sequencing of only seven Qataris identifies potentially deleterious variants in the Qatari population.

The Qatari population, located at the Arabian migration crossroads of African and Eurasia, is comprised of Bedouin, Persian and African genetic subgroups. By deep exome sequencing of only 7 Qataris, including individuals in each subgroup, we identified 2,750 nonsynonymous SNPs predicted to be deleterious, many of which are linked to human health, or are in genes linked to human health. Many of these SNPs were at significantly elevated deleterious allele frequency in Qataris compared to other populations worldwide. Despite the small sample size, SNP allele frequency was highly correlated with a larger Qatari sample. Together, the data demonstrate that exome sequencing of only a small number of individuals can reveal genetic variations with potential health consequences in understudied populations.

De novo genome sequencing and comparative genomics of date palm (Phoenix dactylifera).

Date palm is one of the most economically important woody crops cultivated in the Middle East and North Africa and is a good candidate for improving agricultural yields in arid environments. Nonetheless, long generation times (5-8 years) and dioecy (separate male and female trees) have complicated its cultivation and genetic analysis. To address these issues, we assembled a draft genome for a Khalas variety female date palm, the first publicly available resource of its type for a member of the order Arecales. The ∼380 Mb sequence, spanning mainly gene-rich regions, includes >25,000 gene models and is predicted to cover ∼90% of genes and ∼60% of the genome. Sequencing of eight other cultivars, including females of the Deglet Noor and Medjool varieties and their backcrossed males, identified >3.5 million polymorphic sites, including >10,000 genic copy number variations. A small subset of these polymorphisms can distinguish multiple varieties. We identified a region of the genome linked to gender and found evidence that date palm employs an XY system of gender inheritance.