Functional Variants Research Articles

Identification of Noncoding Functional Regulatory Variants of STIM1 Gene in Idiopathic Pulmonary Arterial Hypertension.

STIM1 (stromal interaction molecule 1) regulates store-operated calcium entry and is involved in pulmonary artery vasoconstriction and pulmonary artery smooth muscle cell proliferation, leading to pulmonary arterial hypertension (PAH). Bioinformatics analysis and a 2-stage matched case-control study were conducted to screen for noncoding variants that may potentially affect STIM1 transcriptional regulation in 242 patients with idiopathic PAH and 414 healthy controls. Luciferase reporter assay, real-time quantitative polymerase chain reaction, western blot, 5-ethynyl-2'-deoxyuridine (EdU) assay, and intracellular Ca2+ measurement were performed to study the mechanistic roles of those STIM1 noncoding variants in PAH. Five noncoding variants (rs3794050, rs7934581, rs3750996, rs1561876, and rs3750994) were identified and genotyped using Sanger sequencing. Rs3794050, rs7934581, and rs1561876 were associated with idiopathic PAH (recessive model, all P<0.05). Bioinformatics analysis showed that these 3 noncoding variants possibly affect the enhancer function of STIM1 or the microRNA (miRNA) binding to STIM1. Functional validation performed in HEK293 and pulmonary artery smooth muscle cells demonstrated that the noncoding variant rs1561876-G (STIM1 mutant) had significantly stronger transcriptional activity than the wild-type counterpart, rs1561876-A, by affecting the transcriptional regulatory function of both hsa-miRNA-3140-5p and hsa-miRNA-4766-5p. rs1561876-G enhanced intracellular Ca2+ signaling in human pulmonary artery smooth muscle cells secondary to calcium-sensing receptor activation and promoted proliferation of pulmonary artery smooth muscle cells under both normoxia and hypoxia conditions, suggesting a possible contribution to PAH development. The potential clinical implications of the 3 noncoding variants of STIM1, rs3794050, rs7934581, and rs1561876, are 2-fold, as they may help predict the risk and prognosis of idiopathic PAH and guide investigations on novel therapeutic pathway(s).

The singular French PCSK9-p.Ser127Arg gain-of-function variant: a significant player in cholesterol levels from a 775-year-old common ancestor

Background and aimsPCSK9 is a key regulator of LDL-cholesterol levels. PCSK9 gain of function variants (GOFVs) cause autosomal dominant hypercholesterolemia (ADH). The first described PCSK9-GOFV, p.Ser127Arg, almost exclusively reported in France, represents 67% of the PCSK9 French GOFVs due to a founder effect. Few other carriers are reported in South Africa and Norway. This study aims to estimate when the common ancestor lived and to describe a cohort of p.Ser127Arg carriers. MethodsEight families and 14 p.Ser127Arg carriers were genotyped and phenotyped. Haplotypes were constructed using 11 microsatellites around PCSK9 and 6 intragenic single nucleotide polymorphisms (SNPs). To add to the biological analysis, eight additional p.Ser127Arg carriers, 12 carriers of other PCSK9-GOFVs, 93 LDLR loss of function variant (LOFV) carriers and 49 non-carriers subjects were phenotyped. ResultsThe most common ancestor of p.Ser127Arg was estimated to have lived 775 years ago [95% CI: 575-1,075]. French Protestants exiled after the revocation of the Edict of Nantes in 1,685 AD likely brought the variant to South Africa and Norway. As expected for ADH subjects, carriers of LDLR-LOFV, the p.Ser127Arg, or other PCSK9-GOFVs showed significantly higher LDL-C levels than that of the non-carriers. Interestingly, LDL-C levels are higher for LDLR-LOFVs and for the reduced secreted p.Ser127Arg than for secreted PCSK9-GOFVs, suggesting a greater effect of the p.Ser127Arg. Conversely, HDL-C was significantly lower for LDLR-LOFV and p.Ser127Arg carriers. ConclusionsThis first report from a large cohort of PCSK9-p.Ser127Arg carriers provides observations suggesting a stronger hypercholesterolemic potential of the mutated pro-PCSK9 compared with the secreted mature protein. This work also provides additional data to support the association between PCSK9 and HDL metabolism, and molecular evidence that this variant appeared in France around 1248 AD.

Further characteristics of Noonan Syndrome type 13 caused by MAPK1 loss of function variant: Case report and literature review

Further characteristics of Noonan Syndrome type 13 caused by MAPK1 loss of function variant: Case report and literature review

LARP1 haploinsufficiency is associated with an autosomal dominant neurodevelopmental disorder

Autism spectrum disorder (ASD) is a neurodevelopmental disorder (NDD) that affects approximately 4% of males and 1% of females in the United States. While causes of ASD are multi-factorial, single rare genetic variants contribute to around 20% of cases. Here we report a case series of seven unrelated probands (6 males, 1 female) with ASD or another variable NDD phenotype attributed to de novo heterozygous loss of function or missense variants in the gene LARP1. LARP1 encodes an RNA binding protein that post-transcriptionally regulates the stability and translation of thousands of mRNAs, including those regulating cellular metabolism and metabolic plasticity. Using lymphocytes collected and immortalized from an index proband who carries a truncating variant in one allele of LARP1, we demonstrated lower cellular levels of LARP1 protein causing reduced rates of aerobic respiration and glycolysis. As expression of LARP1 increases during neurodevelopment, with higher levels in neurons and astrocytes, we propose that LARP1 haploinsufficiency contributes to ASD or related NDDs through attenuated metabolic activity in the developing fetal brain.

Variability in Disease Severity Among Cystic Fibrosis Patients carrying Residual Function Variants: Data from the European Cystic Fibrosis Society Patient Registry

BackgroundCystic Fibrosis (CF) variants that exhibit residual function (RF) of the CF transmembrane conductance regulator (CFTR) are considered to have a milder disease, however, the spectrum of CF phenotype within the different RF variants has not been extensively investigated.AimTo characterise the spectrum of CF disease severity in people with CF (pwCF) carrying different RF variants, using the European CF Society Patient Registry (ECFSPR) data.MethodsA retrospective cross-sectional and longitudinal cohort study included data from the ECFSPR during 2008–2016. Demographic and clinical characteristics of pwCF carrying different RF variants were compared to pwCF homozygous for F508del. Among RF, a distinction was made between pwCF carrying class IV or class V variants and pwCF carrying specific RF variants.ResultsOut of 56 701 pwCF in the ECFSPR, 6192 carried RF variants and 22 766 were homozygous for F508del. Class IV/F508del variants were associated with a milder course compared to class V/F508del, both were milder than pwCF homozygous for F508del. FEV1pp declined already in childhood in all groups. For adults, the hazard ratio of death for class V/F508delversusClass IV/F508del was 2.14 (0.99–4.63, p=0.052). PwCF carrying 3849+10kbC&gt;T/F508del and pwCF carrying R334W/F508del had age-specific FEV1pp and chronic bacterial colonisation similar to pwCF homozygous for F508del.ConclusionThere is a wide spectrum of disease severity between the different RF variants, while some such as 3849+10kbC&gt;T have a a severe disease, which is similar to pwCF homozygous for F508del.

Leveraging Functional Genomics for Understanding Beef Quality Complexities and Breeding Beef Cattle for Improved Meat Quality.

Consumer perception of beef is heavily influenced by overall meat quality, a critical factor in the cattle industry. Genomics has the potential to improve important beef quality traits and identify genetic markers and causal variants associated with these traits through genomic selection (GS) and genome-wide association studies (GWAS) approaches. Transcriptomics, proteomics, and metabolomics provide insights into underlying genetic mechanisms by identifying differentially expressed genes, proteins, and metabolic pathways linked to quality traits, complementing GWAS data. Leveraging these functional genomics techniques can optimize beef cattle breeding for enhanced quality traits to meet high-quality beef demand. This paper provides a comprehensive overview of the current state of applications of omics technologies in uncovering functional variants underlying beef quality complexities. By highlighting the latest findings from GWAS, GS, transcriptomics, proteomics, and metabolomics studies, this work seeks to serve as a valuable resource for fostering a deeper understanding of the complex relationships between genetics, gene expression, protein dynamics, and metabolic pathways in shaping beef quality.

Expression of Autophagy-Related Proteins in Microlissencephaly Associated with a Novel Variant in the WDR81 Gene

Introduction: Microlissencephaly is a subtype of congenital microcephaly characterized by extreme microcephaly with simplified gyral pattern. Other brain malformations may accompany it. WDR81 encodes a multi-domain transmembrane protein that is predominantly expressed in the brain and is thought to play a role in endolysosomal trafficking and autophagy. Methods: We reported two siblings with microlissencephaly born to consanguineous Turkish parents and reviewed all previously reported patients with WDR81 variants presenting with severe microcephaly accompanied by congenital brain malformations. Whole-exome sequencing was performed on both siblings. Sanger DNA sequencing was performed on the patients’ parents. We also examined LC3, p62, and Beclin 1 mRNA and protein expression levels from blood samples of both siblings using real-time PCR and Western blotting, respectively. Results: Whole-exome sequencing revealed a novel biallelic c.4157+5G>A splice variant in the WDR81 gene in both siblings. In our study, LC3, p62, and Beclin 1 mRNA expression levels were high, LC3 protein expression level was also high and p62 and Beclin 1 protein expression levels were low. Conclusion: High LC3 and low p62 protein expression levels supported studies concluding that WDR81 inhibits autophagy through PI3KC3 inhibition, while low Beclin 1 protein expression level supported studies concluding that autophagy is suppressed in case of loss of function variants in the WDR81 gene. We suggest that due to its role in endolysosomal trafficking, WDR81-related diseases should be included in vesicular trafficking disorders.

Liver eQTL meta-analysis illuminates potential molecular mechanisms of cardiometabolic traits

Understanding the molecular mechanisms of complex traits is essential for developing targeted interventions. We analyzed liver expression quantitative-trait locus (eQTL) meta-analysis data on 1,183 participants to identify conditionally distinct signals. We found 9,013 eQTL signals for 6,564 genes; 23% of eGenes had two signals, and 6% had three or more signals. We then integrated the eQTL results with data from 29 cardiometabolic genome-wide association study (GWAS) traits and identified 1,582 GWAS-eQTL colocalizations for 747 eGenes. Non-primary eQTL signals accounted for 17% of all colocalizations. Isolating signals by conditional analysis prior to coloc resulted in 37% more colocalizations than using marginal eQTL and GWAS data, highlighting the importance of signal isolation. Isolating signals also led to stronger evidence of colocalization: among 343 eQTL-GWAS signal pairs in multi-signal regions, analyses that isolated the signals of interest resulted in higher posterior probability of colocalization for 41% of tests. Leveraging allelic heterogeneity, we predicted causal effects of gene expression on liver traits for four genes. To predict functional variants and regulatory elements, we colocalized eQTL with liver chromatin accessibility QTL (caQTL) and found 391 colocalizations, including 73 with non-primary eQTL signals and 60 eQTL signals that colocalized with both a caQTL and a GWAS signal. Finally, we used publicly available massively parallel reporter assays in HepG2 to highlight 14 eQTL signals that include at least one expression-modulating variant. This multi-faceted approach to unraveling the genetic underpinnings of liver-related traits could lead to therapeutic development.

Expanding the phenotypes of ABL1 deficiency syndromes: When mutations in different isoforms Lead to different diseases.

All reported ABL1 gain of function and loss of function (LOF) variants, impact both isoforms 1a and 1b. Our findings suggest that LOF variants affecting solely ABL1 isoform 1b may lead to a distinct autosomal recessive new phenotype.

Heterozygous loss of function variants in IFT140 are associated with polycystic kidney disease.

Autosomal dominant polycystic kidney disease (ADPKD) affects 1 in 1000 adults. Most cases result from causative PKD1 or PKD2 variants. HNF1B, GANAB and ALG9 variants are also associated with ADPKD. Recent evidence indicates that monoallelic loss-of-function (LoF) IFT140 variants are a cause for non-syndromic ADPKD. We describe 368 patients with IFT140 LoF variants and a spectrum of phenotypic findings that support the association of IFT140 with PKD. We reviewed patients with an unknown cause for their cystic disease and those with heterozygous LoF IFT140 variants classified as pathogenic or likely pathogenic from a cohort that received genetic testing using a panel of 385 renal disease-associated genes. IFT140 LoF variants were significantly enriched in patients with cystic disease when compared with those without cystic disease. A cystic phenotype was reported in 223 of the 368 (60.6%) individuals harboring an IFT140 LoF variant, 98% of which had no other identified cause for their cystic disease. Of 122 unique LoF IFT140 variants identified, 56 (46%) were frameshift, 38 (31%) nonsense, 22 (18%) splice site and 6 (5%) exon-level deletions. Only six IFT140 individuals were reported with end-stage kidney disease, consistent with observed milder clinical presentations in IFT140-related PKD. This study offers further evidence for the involvement ofLoF IFT140 variants in PKD, particularly when no additional molecular etiology has been identified.

Addressing epistasis in the design of protein function

Mutations in protein active sites can dramatically improve function. The active site, however, is densely packed and extremely sensitive to mutations. Therefore, some mutations may only be tolerated in combination with others in a phenomenon known as epistasis. Epistasis reduces the likelihood of obtaining improved functional variants and dramatically slows natural and lab evolutionary processes. Research has shed light on the molecular origins of epistasis and its role in shaping evolutionary trajectories and outcomes. In addition, sequence- and AI-based strategies that infer epistatic relationships from mutational patterns in natural or experimental evolution data have been used to design functional protein variants. In recent years, combinations of such approaches and atomistic design calculations have successfully predicted highly functional combinatorial mutations in active sites. These were used to design thousands of functional active-site variants, demonstrating that, while our understanding of epistasis remains incomplete, some of the determinants that are critical for accurate design are now sufficiently understood. We conclude that the space of active-site variants that has been explored by evolution may be expanded dramatically to enhance natural activities or discover new ones. Furthermore, design opens the way to systematically exploring sequence and structure space and mutational impacts on function, deepening our understanding and control over protein activity.

High-throughput identification of functional regulatory SNPs in systemic lupus erythematosus

Genome-wide association studies implicate multiple loci in risk for systemic lupus erythematosus (SLE), but few contain exonic variants, rendering systematic identification of non-coding variants essential to decoding SLE genetics. We utilized SNP-seq and bioinformatic enrichment to interrogate 2180 single-nucleotide polymorphisms (SNPs) from 87 SLE risk loci for potential binding of transcription factors and related proteins from B cells. 52 SNPs that passed initial screening were tested by electrophoretic mobility shift and luciferase reporter assays. To validate the approach, we studied rs2297550 in detail, finding that the risk allele enhanced binding to the transcription factor Ikaros (encoded by IKZF1), thereby modulating expression of IKBKE. Correspondingly, primary cells from genotyped healthy donors bearing the risk allele expressed higher levels of the interferon / NF-κB regulator IKKε. Together, these findings define a set of likely functional non-coding lupus risk variants and identify a regulatory pathway involving rs2297550, Ikaros, and IKKε implicated by human genetics in risk for SLE.

Genotype-phenotype spectrum and correlation of PHARC Syndrome due to pathogenic ABHD12 variants

BackgroundA comprehensive understanding of the genetic basis of rare diseases and their regulatory mechanisms is essential for human molecular genetics. However, the genetic mutant spectrum of pathogenic genes within the Chinese population remains underrepresented. Here, we reported previously unreported functional ABHD12 variants in two Chinese families and explored the correlation between genetic polymorphisms and phenotypes linked to PHARC syndrome.MethodsParticipants with biallelic pathogenic ABHD12 variants were recruited from the Chinese Deafness Genetics Cohort. These participants underwent whole-genome sequencing. Subsequently, a comprehensive literature review was conducted.ResultsTwo Han Chinese families were identified, one with a compound heterozygous variant and the other with a novel homozygous variant in ABHD12. Among 65 PHARC patients, including 62 from the literature and 3 from this study, approximately 90% (57 out of 63) exhibited hearing loss, 82% (50 out of 61) had cataracts, 82% (46 out of 56) presented with retinitis pigmentosa, 79% (42 out of 53) experienced polyneuropathy, and 63% (36 out of 57) displayed ataxia. Seventeen different patterns were observed in the five main phenotypes of PHARC syndrome. A total of 33 pathogenic variants were identified in the ABHD12. Compared with other genotypes, individuals with biallelic truncating variants showed a higher incidence of polyneuropathy (p = 0.006), but no statistically significant differences were observed in the incidence of hearing loss, ataxia, retinitis pigmentosa and cataracts.ConclusionsThe diagnosis of PHARC syndrome is challenging because of its genetic heterogeneity. Therefore, exploring novel variants and establishing genotype-phenotype correlations can significantly enhance gene diagnosis and genetic counseling for this complex disease.

DNA-binding factor footprints and enhancer RNAs identify functional non-coding genetic variants

BackgroundGenome-wide association studies (GWAS) have revealed a multitude of candidate genetic variants affecting the risk of developing complex traits and diseases. However, the highlighted regions are typically in the non-coding genome, and uncovering the functional causative single nucleotide variants (SNVs) is challenging. Prioritization of variants is commonly based on genomic annotation with markers of active regulatory elements, but current approaches still poorly predict functional variants. To address this, we systematically analyze six markers of active regulatory elements for their ability to identify functional variants.ResultsWe benchmark against molecular quantitative trait loci (molQTL) from assays of regulatory element activity that identify allelic effects on DNA-binding factor occupancy, reporter assay expression, and chromatin accessibility. We identify the combination of DNase footprints and divergent enhancer RNA (eRNA) as markers for functional variants. This signature provides high precision, but with a trade-off of low recall, thus substantially reducing candidate variant sets to prioritize variants for functional validation. We present this as a framework called FINDER—Functional SNV IdeNtification using DNase footprints and eRNA.ConclusionsWe demonstrate the utility to prioritize variants using leukocyte count trait and analyze variants in linkage disequilibrium with a lead variant to predict a functional variant in asthma. Our findings have implications for prioritizing variants from GWAS, in development of predictive scoring algorithms, and for functionally informed fine mapping approaches.

Functional genetic variants and susceptibility and prediction of gestational diabetes mellitus

The aetiological mechanism of gestational diabetes mellitus (GDM) has still not been fully understood. The aim of this study was to explore the associations between functional genetic variants screened from a genome-wide association study (GWAS) and GDM risk among 554 GDM patients and 641 healthy controls in China. Functional analysis of single nucleotide polymorphisms (SNPs) positively associated with GDM was further performed. Univariate regression and multivariate logistic regression analyses were used to screen clinical risk factors, and a predictive nomogram model was established. After adjusting for age and prepregnancy BMI, rs9283638 was significantly associated with GDM susceptibility (P < 0.05). Moreover, an obvious interaction between rs9283638 and clinical variables was detected (Pinteraction < 0.05). Functional analysis confirmed that rs9283638 can regulate not only target gene transcription factor binding, but it also regulates the mRNA levels of SAMD7 (P < 0.05). The nomogram model constructed with the factors of age, FPG, 1hPG, 2hPG, HbA1c, TG and rs9283638 revealed an area under the ROC curve of 0.920 (95% CI 0.902–0.939). Decision curve analysis (DCA) suggested that the model had greater net clinical benefit. Conclusively, genetic variants can alter women's susceptibility to GDM by affecting the transcription of target genes. The predictive nomogram model constructed based on genetic and clinical variables can effectively distinguish individuals with different GDM risk factors.

Abstract Tu069: Development of a Novel Biallelic, Haploinsufficient Mouse Model of MYBPC3 Related Hypertrophic Cardiomyopathy

Background: No current animal models re-capitulate MyBP-c haploinsufficiency and hypertrophic cardiomyopathy (HCM) observed in patients with heterozygous MYBPC3 truncating variants, the most common genetic cause of HCM. We recently identified a hypomorphic (partial) loss of function MYBPC3 variant that increases disease severity in HCM patients when combined with an MYBPC3 truncating variant. Hypothesis: Additive effects from a hypomorphic loss of function MYBPC3 variant observed in human HCM will result in MyBP-c protein haploinsufficiency and HCM in a biallelic murine model. Methods: To test MYBPC3 dose effects, induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) were first derived from an HCM patient with biallelic hypomorphic and complete loss of function variants in MYBPC3 . We then generated Mybpc3 knock out (Mybpc3 KO ) and hypomorphic knock in (Mybpc3 KI ) alleles in mice. Control (Mybpc3 +/+ ), monoallelic, and biallelic mice and iPSC-CMs were characterized with morphometric measurements, echocardiography, mass spectroscopy, and RNA sequencing. Results: Biallelic iPSC-CMs demonstrated haploinsufficiency not present in heterozygous models through dose reduction in MYBPC3 protein (Fig 1A). Similarly, biallelic Mybpc3 KI/KO mice had a reduction in Mybpc3 transcripts (62±3%, p&lt;0.001) with 43±11%% reduction in MyBP-c protein (Fig 1B) and cardiac hypertrophy (10±5% increase in heart mass, p=0.002, Fig 1C). Conclusions: Graded reduction in Mybpc3 transcripts in a biallelic mouse model results in MyBP-c haploinsufficiency similar to that observed in HCM patients. Corresponding hypertrophy establishes this approach as a clinically relevant, novel model of MYBPC3 HCM.

Interference of default mode on attention networks in adults with attention-deficit/hyperactivity disorder and its association with genetic variants and treatment outcomes.

Altered brain functional connectivity has been proposed as the neurobiological underpinnings of attention-deficit/hyperactivity disorder (ADHD), and the default mode interference hypothesis is one of the most popular neuropsychological models. Here, we explored whether this hypothesis is supported in adults with ADHD and the association with high-risk genetic variants and treatment outcomes. Voxel-based whole-brain connectome analysis was conducted on resting-state functional MRI data from 84 adults with ADHD and 89 healthy controls to identify functional connectivity substrates corresponding to ADHD-related alterations. The candidate genetic variants and 12-week cognitive behavioral therapy data were leveraged from the same population to assess these associations. We detected breakdowns of functional connectivity in the precuneus and left middle temporal gyrus in adults with ADHD, with exact contributions from decreased connectivity within the default mode, dorsal and ventral attention networks, as well as increased connectivity among them with the middle temporal gyrus serving as a crucial 'bridge'. Additionally, significant associations between the altered functional connectivity and genetic variants in both MAOA and MAOB were detected. Treatment restored brain function, with the amelioration of connectivity of the middle temporal gyrus, accompanied by improvements in ADHD core symptoms. These findings support the interference of default mode on attention in adults with ADHD and its association with genetic risk variants and clinical management, providing insights into the underlying pathogenesis of ADHD and potential biomarkers for treatment evaluation.

Functional variants of the pentraxin 3 gene are associated with the metastasis and progression of prostate cancer.

Age, ethnic background and genetic components have been identified as the established risks for prostate cancer (PCa). Pentraxin 3 (PTX3), originally identified as a pattern-recognition molecule for defence against infectious agents, has multiple functions in tissue repair and in the regulation of cancer-associated inflammation. In this study, we sought to investigate the impact of PTX3 gene variants on the development of PCa. Genotypes of four common single-nucleotide polymorphisms (SNPs) of PTX3 gene, including rs1840680, rs2305619, rs3816527 and rs2120243, were profiled among 705 PCa patients and 705 ethnicity-matched controls. In this study, we found that patients who carry at least one minor allele (C) of rs3816527 (AC and CC) tended to develop advanced forms of diseases (clinical large T stage, OR, 1.593, p = 0.032; pathologically-confirmed nodal spread, OR, 1.987, p = 0.011; metastatic tumour, OR, 3.896, p = 0.032) as compared with those homologous for the major allele (AA). Further stratification analysis showed that such association of rs3816527 with lymphatic and distal metastasis of PCa was accentuated in the younger age group (≤65 at diagnosis) but not seen in the older age group (>65 at diagnosis), suggesting an age-specific effect of PTX3 variants. Prediction of PTX3 protein structure implied that polymorphism may alter the quaternary organization and oligomerization of PTX3 protein. Moreover, our gene silencing experiments and survey of public datasets revealed that elevation of PTX3 levels in PCa was required for cell migration and associated with tumour metastasis. Our results highlight an association of PTX3 rs3816527 with the progression of PCa.

Environmental genome-wide association studies across precipitation regimes reveal that the E3 ubiquitin ligase MBR1 regulates plant adaptation to rainy environments

In an era characterized by rapidly changing and less-predictable weather conditions fueled by the climate crisis, understanding the mechanisms underlying local adaptation in plants is of paramount importance for the conservation of species. As the frequency and intensity of extreme precipitation events increase, so are the flooding events resulting from soil water saturation. The subsequent onset of hypoxic stress is one of the leading causes of crop damage and yield loss. By combining genomics and remote sensing data, it is now possible to probe natural plant populations that have evolved in different rainfall regimes and look for molecular adaptation to hypoxia. Here, using an environmental genome-wide association study (eGWAS) of 934 non-redundant georeferenced Arabidopsis ecotypes, we have identified functional variants of the gene MED25 BINDING RING-H2 PROTEIN 1 (MBR1). This gene encodes a ubiquitin-protein ligase that regulates MEDIATOR25 (MED25), part of a multiprotein complex that interacts with transcription factors that act as key drivers of the hypoxic response in Arabidopsis, namely the RELATED TO AP2 proteins RAP2.2 and RAP2.12. Through experimental validation, we show that natural variants of MBR1 have different effects on the stability of MED25 and, in turn, on hypoxia tolerance. This study also highlights the pivotal role of the MBR1/MED25 module in establishing a comprehensive hypoxic response. Our findings show that molecular candidates for plant environmental adaptation can be effectively mined from large datasets. This thus supports the need for integration of forward and reverse genetics with robust molecular physiology validation of outcomes.

Potentially functional variants of ERRFI1 in hypoxia-related genes predict survival of non-small cell lung cancer patients.

Hypoxia is often involved in tumor microenvironment, and the hypoxia-induced signaling pathways play a key role in aggressive cancer phenotypes, including angiogenesis, immune evasion, and therapy resistance. However, it is unknown what role genetic variants in the hypoxia-related genes play in survival of patients with non-small cell lung cancer (NSCLC). We evaluated the associations between 16,092 single-nucleotide polymorphisms (SNPs) in 182 hypoxia-related genes and survival outcomes of NSCLC patients. Data from the Prostate, Lung, Colorectal, and Ovarian (PLCO) Cancer Screening Trial were used as the discovery dataset, and the Harvard Lung Cancer Susceptibility (HLCS) Study served as the replication dataset. We also performed additional linkage disequilibrium analysis and a stepwise multivariable Cox proportional hazards regression analysis in the PLCO dataset. An independent SNP, ERRFI1 rs28624 A > C, was identified with an adjusted hazards ratio (HR) of 1.31 (95% CI = 1.14-1.51, p = 0.0001) for overall survival (OS). In further analyses, unfavorable genotypes AC and CC, compared with the AA genotype, were associated a worse OS (HR = 1.20, 95% CI = 1.03-1.39, p = 0.014) and disease-specific survival (HR = 1.21, 95% CI = 1.04-1.42, p = 0.016). Further expression quantitative trait loci analysis indicated that ERRFI1 rs28624C genotypes were significantly associated with higher ERRFI1 mRNA expression levels in the whole blood. Additional analysis showed that high ERRFI1 mRNA expression levels were associated with a worse OS in patients with lung adenocarcinoma. Our findings suggest that genetic variants in the hypoxia-related gene ERRFI1 may modulate NSCLC survival, potentially through their effect on the gene expression.