High frequency of MEFV disease-causing variants in children with very-early-onset inflammatory bowel disease.

BACKGROUND: Biological similarities between inflammatory bowel disease (IBD) and familial Mediterranean fever (FMF) have been described in humans and animal models suggesting a possible common genetic basis. FMF is caused by variants in the MEFV gene which encodes pyrin, an immune regulator. This study aimed to investigate the carrier rate of disease-causing MEFV variants in children of different ethnicities diagnosed with very-early-onset IBD (VEO-IBD). METHODS: The study included 23 children diagnosed with VEO-IBD who had undergone whole exome sequencing. The exomes were evaluated for MEFV monoallelic and biallelic disease-causing variants and compared to exome sequencing data of 250 probands with suspected monogenic diseases other than IBD. RESULTS: Of the 23 children diagnosed with VEO-IBD, 12 (52%) were carriers of at least one MEFV disease-causing variant, which was threefold higher than in individuals without IBD. The most frequent variants identified were p.M694V and p.E148Q (42% each). The allelic frequency of MEFV variants was found to be higher across the VEO-IBD group in 13 of 14 ethnicities compared to the control group. CONCLUSION: The study suggests that disease-causing variants in the MEFV gene should be sought in cases of VEO-IBD. However, the clinical importance of this finding is yet to be defined. IMPACT: There are biological similarities between inflammatory bowel disease and familial Mediterranean fever, suggesting a possible genetic relationship. Children less than 6 years old clinically diagnosed with inflammatory bowel disease have a threefold higher rate of disease-causing variants in the MEFV gene than controls. Monogenic testing in children with very-early-onset inflammatory bowel disease should include a search for MEFV variants.

Discovery of a Novel Missense Variant in NLRP3 Causing Atypical Cryopyrin-Associated Periodic Syndromes With Hearing Loss as the Primary Presentation, Responsive to Anti-Interleukin-1 Therapy.

OBJECTIVE: Cryopyrin-associated periodic syndromes (CAPS), also known as NLRP3-associated autoinflammatory diseases, are a spectrum of rare autoinflammatory diseases caused by gain-of-function variants in the NLRP3 gene, resulting in inflammasome hyperactivation and dysregulated release of interleukin-1ß (IL-1ß). Many patients with CAPS develop progressive sensorineural hearing loss (SNHL) because of cochlear autoinflammation, which may be the sole manifestation in rare cases. This study was undertaken to establish the suspected diagnosis of CAPS in a family presenting with autosomal-dominant progressive/acute SNHL and a novel missense variant in the NLRP3 gene of unknown significance (NM_001079821.3:c.1784G>A p.Ser595Asn). METHODS: We conducted an ex vivo functional assessment of the NLRP3 inflammasome in heterozygous individuals (n = 10) and healthy family members (n = 5). RESULTS: The assay revealed hyperactivation of the inflammasome among heterozygous individuals, supporting the hypothesis that this missense variant is a pathogenic gain-of-function variant. Administration of IL-1 receptor antagonist resulted in a substantial clinical improvement among pediatric patients, who exhibited near resolution of hearing impairment within 1 to 3 months of treatment. CONCLUSION: Our findings highlight the crucial role of early diagnosis and treatment with an anti-IL-1 agent in reversing cochlear damage. Furthermore, our results suggest that high- and ultrahigh-frequency ranges need to be included in the auditory assessment to enable early detection of subclinical SNHL. Finally, incorporating functional inflammasome assessment as part of the clinical evaluation could establish the diagnosis in inconclusive cases.

A novel SLC25A13 gene splice site variant causes Citrin deficiency in an infant.

Citrin deficiency is an autosomal recessive disorder associated with SLC25A13 gene pathogenic variants, with more than a hundred known at present. It manifests in neonates as failure to thrive and acute liver insufficiency. We herein describe a case of a 4-week-old infant who presented with insufficient weight gain and liver failure accompanied by hyperammonemia. She was diagnosed with Citrin deficiency after a thorough biochemical and molecular analysis including amino acid profile, DNA sequencing of genes of interest and RNA splice site evaluation, to reveal a yet unknown damaging variant of the SLC25A13 gene.

Possible biallelic inheritance in TIE1 in a family with congenital lymphedema, intestinal lymphangiectasia and cutis aplasia.

A short report of two male siblings born with cutis aplasia, lymphedema and intestinal lymphangiectasia, one found to carry bi-allelic variants in the TIE1 gene known to be associated with congenital lymphedema.

Microdeletion of 16q24.1-q24.2-A unique etiology of Lymphedema-Distichiasis syndrome and neurodevelopmental disorder.

Interstitial deletions of 16q24.1-q24.2 are associated with alveolar capillary dysplasia, congenital renal malformations, neurodevelopmental disorders, and congenital abnormalities. Lymphedema-Distichiasis syndrome (LDS; OMIM # 153400) is a dominant condition caused by heterozygous pathogenic variants in FOXC2. Usually, lymphedema and distichiasis occur in puberty or later on, and affected individuals typically achieve normal developmental milestones. Here, we describe a boy with congenital lymphedema, distichiasis, bilateral hydronephrosis, and global developmental delay, with a de novo microdeletion of 894 kb at 16q24.1-q24.2. This report extends the phenotype of both 16q24.1-q24.2 microdeletion syndrome and of LDS. Interestingly, the deletion involves only the 3'-UTR part of FOXC2.

The prevalence of prenatal sonographic findings in postnatal diagnostic exome sequencing performed for neurocognitive phenotypes: A cohort study.

OBJECTIVE: Prenatal exome sequencing (ES) is currently indicated for fetal malformations. Some neurocognitive genetic disorders may not have a prenatal phenotype. We assessed the prevalence of prenatally detectable phenotypes among patients with neurocognitive syndromes diagnosed postnatally by ES. METHODS: The medical files of a cohort of 138 patients diagnosed postnatally with a neurocognitive disorder using ES were reviewed for prenatal sonographic data. The Online Mendelian Inheritance in Man (OMIM) database was searched for prenatally detectable phenotypes for all genes identified. RESULTS: Prenatal imaging data were available for 122 cases. Of these, 29 (23.75%) had fetal structural abnormalities and another 29 had other ultrasound abnormalities (fetal growth restriction, polyhydramnios, elevated nuchal translucency). In 30 patients, structural aberrations that were not diagnosed prenatally were detected at birth; in 21 (17.2%), the abnormalities could theoretically be detected prenatally by third-trimester/targeted scans. According to OMIM, 55.9% of the diagnosed genes were not associated with structural anomalies. CONCLUSIONS: Most patients (52.5%) with postnatally diagnosed neurocognitive disorders did not have prenatal sonographic findings indicating prenatal ES should be considered. The prevalence of specific prenatal phenotypes such as fetal growth restriction and polyhydramnios in our cohort suggests that additional prenatal findings should be assessed as possible indications for prenatal ES.

Di-genic inheritance of germline POLE and PMS2 pathogenic variants causes a unique condition associated with pediatric cancer predisposition.

Polymerase proofreading-associated polyposis (PPAP) and Lynch syndrome, caused by mutated POLE and mismatch repair (MMR) genes, respectively, are associated with adult-onset cancer. PPAP and MMR-deficient tumors are both hypermutated, and each has a unique mutational signature. We describe a 4.5-year-old boy with multiple café au lait spots who presented with metastatic Sonic Hedgehog-activated medulloblastoma, with partial response to intensive chemotherapy and immunotherapy. The tumor showed microsatellite stability, loss of PMS2 nuclear expression, and an exceptionally high tumor mutational burden of 276 Mut/Mb. Germline molecular analysis revealed an inherited heterozygous pathogenic POLE variant and a de novo heterozygous PMS2 pathogenic variant. The tumor featured the MMR, POLE, and POLE+MMR mutational signatures. This is the first description of a di-genic condition, which we named "POL-LYNCH syndrome," manifested by an aggressive ultra-mutant pediatric medulloblastoma with a unique genomic signature.

The diagnostic efficacy of exome data analysis using fixed neurodevelopmental gene lists: Implications for prenatal setting.

OBJECTIVE: Laboratories performing prenatal exome sequencing (ES) frequently limit analysis to predetermined gene lists. We used a diagnostic postnatal ES cohort to assess how many of the genes diagnosed are not included in a number of select fixed lists used for prenatal diagnosis. METHODS: Of 601 postnatal ES tests, pathogenic variants related to neurodevelopmental disorders were detected in 138 probands. We evaluated if causative genes were present in the following: (1) Developmental Disorders Genotype-Phenotype database list, (2) a commercial laboratory list for prenatal ES, (3) the PanelApp fetal anomalies panel, and (4) a published list used for prenatal diagnosis by ES (Prenatal Assessment of Genomes and Exomes study). RESULTS: The percentages of cases where the diagnosed gene was not included in the selected four lists were; 11.6%, 17.24%, 23.2%, and 10.9%, respectively. In 13/138 (9.4%) cases, the causative gene was not included in any of the lists; in 4/13 (∼30%) cases noninclusion was explained by a relatively recent discovery of gene-phenotype association. CONCLUSIONS: A significant number of genes related to neurocognitive phenotypes are not included in some of the lists used for prenatal ES data interpretation. These are not only genes related to recently discovered disorders, but also genes with well-established gene-phenotype.

Re-evaluating the pathogenicity of the c.783+2T>C BAP1 germline variant.

BAP1 germline pathogenic sequence variants (PSVs) underlie a unique tumor predisposition syndrome (BAP1-TPDS) associated with an increased lifetime risk for developing primarily pleural and peritoneal mesothelioma and uveal and cutaneous melanoma. Overwhelmingly, BAP1 PSVs are unique, family-specific inactivating variants. We identified seven families, six of Jewish Iraqi origin, harboring an identical BAP1 splice variant (c.783+2T>C), currently assigned a "likely pathogenic" status. Given a nonclassical BAP1-TPDS tumor type clustering and low penetrance in these families, the pathogenicity of this variant was re-evaluated by a combined approach including literature analysis, revised bioinformatics analysis, allelic loss, effect on the transcript, and tumor protein expression patterns. None of the three available tumors showed an allelic loss, there was no discernable effect on alternative splicing based on reverse-transcription polymerase chain reaction, and there was no decrease or loss of somatic protein expression in 2/3 analyzed tumors. This led to assigning a Benign Strong (BS) criteria, BS4, supporting BS3 criteria, and weakening the Pathogenic Supporting (PP) criteria PP5. Combined, these data suggest that this sequence variant should be reclassified as a variant of unknown significance by American College of Medical Genetics (ACMG) criteria.

Epilepsy and electroencephalogram evolution in YWHAG gene mutation: A new phenotype and review of the literature.

A male patient with a de novo mutation in the YWHAG gene and mild phenotype is presented. He had normal delivery and normal development, with normal speech and social milestones. At the age of 9 months, myoclonic seizures started, with generalized epileptiform discharges. The child responded well to levetiracetam monotherapy with complete seizure resolution. Levetiracetam was stopped and he remained seizure-free for 10 months. His development was appropriate for age according to psychological evaluation and he attended a regular kindergarten. At the age of approximately 4 years, the seizures reappeared with different semiology of staring with eye blinking. Electroencephalogram (EEG) showed multifocal spikes. Brain magnetic resonance imaging did not reveal any structural abnormality. Genetic analysis revealed a de novo likely pathogenic missense variant in the YWHAG gene (c.619G>A p.Glu207Lys). We compared our case to the other cases published in the literature. Our case is unique in its seizure semiology and evolution of EEG. Moreover, in contrast to our case, the majority of cases described in the literature have dysmorphism and intellectual disability or autistic spectrum disorder. This report emphasizes the phenotypic heterogeneity of YWHAG mutation as is the case in other developmental encephalopathies.

Two intronic cis-acting variants in both alleles of the POLR3A gene cause progressive spastic ataxia with hypodontia.

POLR3A encodes the largest subunit of the DNA-dependent RNA polymerase III. Pathogenic variants in this gene are associated with dysregulation of tRNA production and other non-coding RNAs. POLR3A-related disorders include variable phenotypes. The genotype-phenotype correlation is still unclear. Phenotypic analysis and exome sequencing were performed in four affected siblings diagnosed clinically with hereditary spastic ataxia, two healthy siblings and their unaffected mother. All four affected siblings (ages 46-55) had similar clinical features of early childhood-onset hypodontia and adolescent-onset progressive spastic ataxia. None had progeria, gonadal dysfunction or dysmorphism. All affected individuals had biallelic POLR3A pathogenic variants composed by two cis-acting intronic splicing-altering variants, c.1909 + 22G > A and c.3337-11 T > C. The two healthy siblings had wild-type alleles. The mother and another unaffected sibling were heterozygous for the allele containing both variants. This is the first report addressing the clinical consequence associated with homozygosity for a unique pathogenic intronic allele in the POLR3A gene. This allele was previously reported in compound heterozygous combinations in patients with Wiedemann-Rautenstrauch syndrome, a severe progeroid POLR3A-associated phenotype. We show that homozygosity for this allele is associated with spastic ataxia with hypodontia, and not with progeroid features. These findings contribute to the characterization of genotype-phenotype correlation in POLR3A-related disorders.

The role of phenotype-based search approaches using public online databases in diagnostics of Mendelian disorders.

PURPOSE: To investigate the effectiveness of phenotype-based search approaches using publicly available online databases. METHODS: We included consecutively solved cases from our exome database. For each case, the combination of Human Phenotype Ontology terms reported by the referring clinician was used to perform a search in three commonly used databases: OMIM (first 300 results), Phenolyzer (first 300 results), and Mendelian (all 100 results). RESULTS: One hundred cases were included (43 females; mean age: 10 years). The actual molecular diagnosis identified through exome sequencing was not included in the search results of any of the queried databases in 33% of cases. In 85% of cases it was not found within the top five search results. When included, its median rank was 61 (range: 1-295), 21 (1-270), and 29 (1-92) in OMIM, Phenolyzer and Mendelian, respectively. CONCLUSION: This study demonstrates that, in most cases, phenotype-based search approaches using public online databases is ineffective in providing a probable diagnosis for Mendelian conditions. Genotype-first approach through molecular-guided diagnostics with backward phenotyping may be a more appropriate approach for these disorders, unless a specific diagnosis is considered a priori based on highly unique phenotypic features or a specific facial gestalt.

When phenotype does not match genotype: importance of "real-time" refining of phenotypic information for exome data interpretation.

PURPOSE: Clinical data provided to genetic testing laboratories are frequently scarce. Our purpose was to evaluate clinical scenarios where phenotypic refinement in proband's family members might impact exome data interpretation. METHODS: Of 614 exomes, 209 were diagnostic and included in this study. Phenotypic information was gathered by the variant interpretation team from genetic counseling letters and images. If a discrepancy between reported clinical findings and presumably disease-causing variant segregation was observed, referring clinicians were contacted for phenotypic clarification. RESULTS: In 16/209 (7.7%) cases, phenotypic refinement was important due to (1) lack of cosegregation of disease-causing variant with the reported phenotype; (2) identification of different disorders with overlapping symptoms in the same family; (3) similar features in proband and family members, but molecular cause identified in proband only; and (4) previously unrecognized maternal condition causative of child's phenotype. As a result of phenotypic clarification, in 12/16 (75%) cases definition of affected versus unaffected status in one of the family members has changed, and in one case variant classification has changed. CONCLUSION: Detailed description of phenotypes in family members including differences in clinical presentations, even if subtle, are important in exome interpretation and should be communicated to the variant interpretation team.

Biallelic variants in ETV2 in a family with congenital heart defects, vertebral abnormalities and preaxial polydactyly.

The combination of congenital heart defects and vertebral anomalies with or without additional abnormalities has been reported in many genetic disorders. We describe a family in which four consecutive pregnancies were characterized by the combination of fetal congenital heart malformations and vertebral anomalies. In addition, preaxial polydactyly was detected in one of the fetuses. Reanalysis of the non-diagnostic clinical exome data revealed compound heterozygous variants c.350del, p.(Gly117AlafsTer90) and c.757G > T, p.(Asp253Tyr) in ETV2 which have previously not been known to be associated with a phenotype in humans. In mice, Etv2 encodes an obligatory transcription factor involved in the generation of hematopoietic and endothelial cells. Its homozygous disruption results in embryonic lethality due to severe blood and vessel defects. The Etv2 promoter may be bound by Nkx2-5, a key transcription factor in heart development. Pathogenic variants in the NKx2-5 homolog in humans (NKX2-5) are related to congenital heart defects. The identification of additional fetuses or live-born individuals with biallelic pathogenic variants in ETV2 will shed further light on this presumably novel gene-phenotype association and on the full phenotypic spectrum.

Similar burden of pathogenic coding variants in exceptionally long-lived individuals and individuals without exceptional longevity.

Centenarians (exceptionally long-lived individuals-ELLI) are a unique segment of the population, exhibiting long human lifespan and healthspan, despite generally practicing similar lifestyle habits as their peers. We tested disease-associated mutation burden in ELLI genomes by determining the burden of pathogenic variants reported in the ClinVar and HGMD databases using data from whole exome sequencing (WES) conducted in a cohort of ELLI, their offspring, and control individuals without antecedents of familial longevity (n = 1879), all descendent from the founder population of Ashkenazi Jews. The burden of pathogenic variants did not differ between the three groups. Additional analyses of variants subtypes and variant effect predictor (VEP) biotype frequencies did not reveal a decrease of pathogenic or loss-of-function (LoF) variants in ELLI and offspring compared to the control group. Case-control pathogenic variants enrichment analyses conducted in ELLI and controls also did not identify significant differences in any of the variants between the groups and polygenic risk scores failed to provide a predictive model. Interestingly, cancer and Alzheimer's disease-associated variants were significantly depleted in ELLI compared to controls, suggesting slower accumulation of mutation. That said, polygenic risk score analysis failed to find any predictive variants among the functional variants tested. The high similarity in the burden of pathogenic variation between ELLI and individuals without familial longevity supports the notion that extension of lifespan and healthspan in ELLI is not a consequence of pathogenic variant depletion but rather a result of other genomic, epigenomic, or potentially nongenomic properties.

Grain protein content and thousand kernel weight QTLs identified in a durum × wild emmer wheat mapping population tested in five environments.

KEY MESSAGE: Genetic dissection of GPC and TKW in tetraploid durum × WEW RIL population, based on high-density SNP genetic map, revealed 12 GPC QTLs and 11 TKW QTLs, with favorable alleles for 11 and 5 QTLs, respectively, derived from WEW. Wild emmer wheat (Triticum turgidum ssp. dicoccoides, WEW) was shown to exhibit high grain protein content (GPC) and therefore possess a great potential for improvement of cultivated wheat nutritional value. Genetic dissection of thousand kernel weight (TKW) and grain protein content (GPC) was performed using a high-density genetic map constructed based on a recombinant inbred line (RIL) population derived from a cross between T. durum var. Svevo and WEW acc. Y12-3. Genotyping of 208 F6 RILs with a 15 K wheat single nucleotide polymorphism (SNP) array yielded 4166 polymorphic SNP markers, of which 1510 were designated as skeleton markers. A total map length of 2169 cM was obtained with an average distance of 1.5 cM between SNPs. A total of 12 GPC QTLs and 11 TKW QTLs were found under five different environments. No significant correlations were found between GPC and TKW across all environments. Four major GPC QTLs with favorable alleles from WEW were found on chromosomes 4BS, 5AS, 6BS and 7BL. The 6BS GPC QTL coincided with the physical position of the NAC transcription factor TtNAM-B1, underlying the cloned QTL, Gpc-B1. Comparisons of the physical intervals of the GPC QTLs described here with the results previously reported in other durum × WEW RIL population led to the discovery of seven novel GPC QTLs. Therefore, our research emphasizes the importance of GPC QTL dissection in diverse WEW accessions as a source of novel alleles for improvement of GPC in cultivated wheat.

Cloning of the wheat Yr15 resistance gene sheds light on the plant tandem kinase-pseudokinase family.

Yellow rust, caused by Puccinia striiformis f. sp. tritici (Pst), is a devastating fungal disease threatening much of global wheat production. Race-specific resistance (R)-genes are used to control rust diseases, but the rapid emergence of virulent Pst races has prompted the search for a more durable resistance. Here, we report the cloning of Yr15, a broad-spectrum R-gene derived from wild emmer wheat, which encodes a putative kinase-pseudokinase protein, designated as wheat tandem kinase 1, comprising a unique R-gene structure in wheat. The existence of a similar gene architecture in 92 putative proteins across the plant kingdom, including the barley RPG1 and a candidate for Ug8, suggests that they are members of a distinct family of plant proteins, termed here tandem kinase-pseudokinases (TKPs). The presence of kinase-pseudokinase structure in both plant TKPs and the animal Janus kinases sheds light on the molecular evolution of immune responses across these two kingdoms.

Genomic Instabilities, Cellular Senescence, and Aging: In Vitro, In Vivo and Aging-Like Human Syndromes.

As average life span and elderly people prevalence in the western world population is gradually increasing, the incidence of age-related diseases such as cancer, heart diseases, diabetes, and dementia is increasing, bearing social and economic consequences worldwide. Understanding the molecular basis of aging-related processes can help extend the organism's health span, i.e., the life period in which the organism is free of chronic diseases or decrease in basic body functions. During the last few decades, immense progress was made in the understanding of major components of aging and healthy aging biology, including genomic instability, telomere attrition, epigenetic changes, proteostasis, nutrient sensing, mitochondrial dysfunction, cellular senescence, stem cell exhaustion, and intracellular communications. This progress has been made by three spear-headed strategies: in vitro (cell and tissue culture from various sources), in vivo (includes diverse model and non-model organisms), both can be manipulated and translated to human biology, and the study of aging-like human syndromes and human populations. Herein, we will focus on current repository of genomic "senescence" stage of aging, which includes health decline, structural changes of the genome, faulty DNA damage response and DNA damage, telomere shortening, and epigenetic alterations. Although aging is a complex process, many of the "hallmarks" of aging are directly related to DNA structure and function. This review will illustrate the variety of these studies, done in in vitro, in vivo and human levels, and highlight the unique potential and contribution of each research level and eventually the link between them.

Evolution and Adaptation of Wild Emmer Wheat Populations to Biotic and Abiotic Stresses.

The genetic bottlenecks associated with plant domestication and subsequent selection in man-made agroecosystems have limited the genetic diversity of modern crops and increased their vulnerability to environmental stresses. Wild emmer wheat, the tetraploid progenitor of domesticated wheat, distributed along a wide range of ecogeographical conditions in the Fertile Crescent, has valuable "left behind" adaptive diversity to multiple diseases and environmental stresses. The biotic and abiotic stress responses are conferred by series of genes and quantitative trait loci (QTLs) that control complex resistance pathways. The study of genetic diversity, genomic organization, expression profiles, protein structure and function of biotic and abiotic stress-resistance genes, and QTLs could shed light on the evolutionary history and adaptation mechanisms of wild emmer populations for their natural habitats. The continuous evolution and adaptation of wild emmer to the changing environment provide novel solutions that can contribute to safeguarding food for the rapidly growing human population.