Compound Heterozygosity of Low-Frequency Promoter Deletions and Rare Loss-of-Function Mutations in TXNL4A Causes Burn-McKeown Syndrome

Abstract We identified and characterized 14 extragenic mutations that suppressed the dominant egg-laying defect of certain lin-12 gain-of-function mutations. These suppressors defined seven genes: sup-l7, lag-2, sel-4, sel-5, sel-6, sel-7 and sel-8. Mutations in six of the genes are recessive suppressors, whereas the two mutations that define the seventh gene, lag-2, are semi-dominant suppressors. These suppressor mutations were able to suppress other lin-12 gain-of-function mutations. The suppressor mutations arose at a very low frequency per gene, 10-50 times below the typical loss-of-function mutation frequency. The suppressor mutations in sup1 7 and lag-2 were shown to be rare non-null alleles, and we present evidence that null mutations in these two genes cause lethality. Temperature-shift studies for two suppressor genes, sup1 7and lag-2, suggest that both genes act at approximately the same time as lin-12in specifying a cell fate. Suppressor alleles of six of these genes enhanced a temperature-sensitive loss-of-function allele of glp-1, a gene related to lin-12 in structure and function. Our analysis of these suppressors suggests that the majority of these genes are part of a shared lin-12/glp-1 signal transduction pathway, or act to regulate the expression or stability of lin-12 and glp-1.

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Identification and characterization of a natural polymorphism in FT-A2 associated with increased number of grains per spike in wheat

Theoretical and Applied Genetics ◽

10.1007/s00122-021-03992-y ◽

2021 ◽

Author(s):

Priscilla Glenn ◽

Junli Zhang ◽

Gina Brown-Guedira ◽

Noah DeWitt ◽

Jason P. Cook ◽

...

Keyword(s):

Grain Yield ◽

Durum Wheat ◽

Tetraploid Wheat ◽

Low Frequency ◽

Yield Component ◽

Loss Of Function ◽

Grain Number ◽

Wheat Varieties ◽

Positive Effects ◽

Grain Number Per Spike

Abstract Key message We discovered a natural FT-A2 allele that increases grain number per spike in both pasta and bread wheat with limited effect on heading time. Abstract Increases in wheat grain yield are necessary to meet future global food demands. A previous study showed that loss-of-function mutations in FLOWERING LOCUS T2 (FT2) increase spikelet number per spike (SNS), an important grain yield component. However, these mutations were also associated with reduced fertility, offsetting the beneficial effect of the increases in SNS on grain number. Here, we report a natural mutation resulting in an aspartic acid to alanine change at position 10 (D10A) associated with significant increases in SNS and no negative effects on fertility. Using a high-density genetic map, we delimited the SNS candidate region to a 5.2-Mb region on chromosome 3AS including 28 genes. Among them, only FT-A2 showed a non-synonymous polymorphism (D10A) present in two different populations segregating for the SNS QTL on chromosome arm 3AS. These results, together with the known effect of the ft-A2 mutations on SNS, suggest that variation in FT-A2 is the most likely cause of the observed differences in SNS. We validated the positive effects of the A10 allele on SNS, grain number, and grain yield per spike in near-isogenic tetraploid wheat lines and in an hexaploid winter wheat population. The A10 allele is present at very low frequency in durum wheat and at much higher frequency in hexaploid wheat, particularly in winter and fall-planted spring varieties. These results suggest that the FT-A2 A10 allele may be particularly useful for improving grain yield in durum wheat and fall-planted common wheat varieties.

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SUN-564 ATG7 Overexpression Results in Reduction of Hepatocellular Lipid Content in Vitro

Journal of the Endocrine Society ◽

10.1210/jendso/bvaa046.272 ◽

2020 ◽

Vol 4 (Supplement_1) ◽

Author(s):

Federica Tavaglione ◽

Guido Baselli ◽

Ester Ciociola ◽

Umberto Vespasiani Gentilucci ◽

Luca Valenti ◽

...

Keyword(s):

Liver Disease ◽

Lipid Content ◽

Lipid Droplets ◽

Low Frequency ◽

Underlying Mechanism ◽

Loss Of Function ◽

Wild Type ◽

Intracellular Lipid ◽

Heparg Cells

Abstract Abstract: Non-alcoholic fatty liver disease (NAFLD) is currently the most common liver disease worldwide, paralleling the epidemic of obesity and type 2 diabetes. Despite the high prevalence of NAFLD, only a minority of patients progress to NASH and advanced fibrosis. The reasons for this inter-individual variability are not completely understood but can be partially accounted for by genetic risk factors (1). Although several common genetic variants associated with liver disease have been identified, there is still a proportion of NAFLD heritability that remains unknown. The rare rs143545741 C>T variant in the autophagy related 7 (ATG7) gene (P426L) has been associated with a higher risk of progressive NAFLD (2). Interestingly, ATG7 encodes a E1-like ubiquitin activating enzyme which is involved in hepatic lipophagy (3). We hypothesized that the unknown heritability of NAFLD might be partially explained by rare genetic variants, therefore not identified in the GWAS studies. Moreover, we assumed that loss-of-function variants in ATG7 might confer an increased susceptibility to NAFLD by reducing autophagic catabolism of lipid droplets in the liver. To examine the underlying mechanism of the low-frequency V471A variant and the rare T86I, L127I, Q170E, and P426L variants in ATG7, we performed in vitro experiments of HepaRG cells overexpressing the human V5-tagged ATG7. We observed a reduction in intracellular lipid content in HepaRG cells overexpressing the ATG7 wild type and the 86I mutant protein (p=0.029, n=4) but not the 127I, 170E, 426L, and 471A mutant proteins. Cells with the ATG7 127I, 170E, 426L, and 471A mutants had higher intracellular lipid content compared to cells overexpressing the wild type protein (p=0.029, n=4). Our data suggested that the low-frequency V471A variant and the rare L127I, Q170E, and P426L variants in ATG7 are loss-of-function, resulting in defective lipophagy, reduced hepatocellular lipid droplets turnover, and excessive lipid accumulation. More experiments are needed to clarify the underlying mechanism of the T86I variant. In conclusion, we highlighted a role for ATG7 in reducing hepatocellular lipid content. Furthermore, we provided evidence showing non-synonymous variants in ATG7 increase the risk of NAFLD and that these variants are loss-of-function. We speculate that ATG7 might be a new susceptibility risk genetic locus for liver disease development and progression. References: (1) Eslam et al. J Hepatol. 2018;68(2):268–279. (2) Baselli et al. The Liver Meeting 2018 - AASLD. Hepatology. October 2018. Volume 68, Issue S1. (3) Martinez-Lopez and Singh. Annu Rev Nutr. 2015;35:215–37.

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Metastatic and recurrent adrenocortical cancer is not defined by its genomic landscape

BMC Medical Genomics ◽

10.1186/s12920-020-00809-7 ◽

2020 ◽

Vol 13 (1) ◽

Author(s):

Tito Fojo ◽

Lyn Huff ◽

Thomas Litman ◽

Kate Im ◽

Maureen Edgerly ◽

...

Keyword(s):

Copy Number ◽

Expression Profiles ◽

Low Frequency ◽

Beta Catenin ◽

Cancer Genes ◽

Loss Of Function ◽

Adrenocortical Cancer ◽

Genomic Aberration ◽

Transcript Length ◽

Metastatic Sites

Abstract Background Adrenocortical carcinoma (ACC) is a rare, often-aggressive neoplasm of the adrenal cortex, with a 14–17 month median overall survival. We asked whether tumors from patients with advanced or metastatic ACC would offer clues as to putative genes that might have critical roles in disease progression or in more aggressive disease biology. Methods We conducted comprehensive genomic and expression analyses of ACCs from 43 patients, 30 female, and 42 from metastatic sites, including deep sequencing, copy number analysis, mRNA expression and microRNA arrays. Results Copy number gains and losses were similar to that previously reported for ACC. We identified a median mutation rate of 3.38 per megabase (Mb). The mutational signature was characterized by a predominance of C > T, C > A and T > C transitions. Only cancer genes TP53 (26%) and beta-catenin (CTNNB1, 14%) were mutated in more than 10% of samples. The TCGA-identified putative cancer genes MEN1 and PRKAR1A were found in low frequency—4.7 and 2.3%, respectively. The majority of the mutations were in genes not implicated in the etiology or maintenance of cancer. Specifically, amongst the 38 genes that were mutated in more than 9% of samples, only four were represented in Tier 1 of the 576 COSMIC Cancer Gene Census (CCGC). Thus, 82% of genes found to have mutations likely have no role in the etiology or biology of ACC; while the role of the other 18%, if any, remains to be proven. Finally, the transcript length for the 38 most frequently mutated genes in ACC is statistically longer than the average of all coding genes, raising the question of whether transcript length in part determined mutation probability. Conclusions We conclude that the mutational and expression profiles of advanced and metastatic tumors are very similar to those from newly diagnosed patients—with very little in the way of genomic aberration to explain differences in biology. With relatively low mutation rates, few major oncogenic drivers, and loss of function mutations in several epigenetic regulators, an epigenetic basis for ACC may be postulated and serve as the basis for future studies.

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Abstract 116: A Targeted Sequencing Approach in Subjects with Extremely High HDL-C Identifies individuals with Potentially Causal Mutations in Both Alleles of 53 Genes

Arteriosclerosis Thrombosis and Vascular Biology ◽

10.1161/atvb.33.suppl_1.a116 ◽

2013 ◽

Vol 33 (suppl_1) ◽

Author(s):

Daniel B Larach ◽

Aniruddh P Patel ◽

Gina M Peloso ◽

Paolo Zanoni ◽

Stephanie L DerOhannessian ◽

...

Keyword(s):

Association Studies ◽

Low Frequency ◽

Targeted Sequencing ◽

Phenotypic Characterization ◽

Specific Gene ◽

Compound Heterozygous ◽

Genome Wide Association Studies ◽

Loss Of Function ◽

Low Hdl ◽

Hdl Metabolism

Introduction The molecular etiology of extremely elevated HDL-C is poorly understood. Genome-wide association studies (GWAS) have identified many genetic loci associated with HDL-C, but very few of these harbor genes in which rare mutations are known to cause extremely elevated HDL-C levels. We performed targeted sequencing of exons in genes at GWAS lipid loci on subjects with extremely high HDL-C compared with those with low HDL-C in order to identify individuals with nonsynonymous mutations that could be causal for the extreme high HDL-C phenotype. Methods 389 cases with HDL-C >95 th percentile (102±19 mg/dL), and 387 controls with HDL-C <25 th percentile (32±5 mg/dL) underwent targeted sequencing of exons in genes ±300 kb around each of 95 recently identified GWAS loci associated with lipid parameters. Candidate variants were identified by the following criteria: (1) missense, nonsense, and frameshift variants; (2) predicted to be deleterious by at least 1 of 3 in silico prediction methods; (3) a ratio of high HDL-C heterozygotes to low HDL-C heterozygotes >1.5:1; (4) absence of any low HDL-C homozygotes; (5) MAF <0.01. Individuals with a potentially causal low-frequency variant in both alleles of a specific gene were identified for further phenotypic characterization. Results 46 individuals with extremely high HDL-C were identified to have potentially causal low-frequency variants in both alleles of the same gene. These individuals were homozygous or compound heterozygous for mutations in 53 distinct genes. Most of these genes were not known to play defined roles in HDL metabolism, and include ARHGAP1, BTN2A2, CAMK2B, LILRA3, and OSCAR. These individuals are currently being called back for detailed phenotyping. Conclusions A targeted sequencing approach focused on genes at GWAS HDL-C loci in subjects with extremely high HDL-C identified 46 individuals with potentially causal low-frequency variants in both alleles of the same gene, including genes not known to play a role in HDL metabolism. More detailed phenotyping and family studies are underway, and are likely to provide new information regarding the phenotypic consequences of loss-of-function mutations in both alleles of several genes that influence HDL metabolism.

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Compound heterozygosity for loss-of-function FARSB variants in a patient with classic features of recessive aminoacyl-tRNA synthetase-related disease

Human Mutation ◽

10.1002/humu.23424 ◽

2018 ◽

Vol 39 (6) ◽

pp. 834-840 ◽

Cited By ~ 15

Author(s):

Anthony Antonellis ◽

Stephanie N. Oprescu ◽

Laurie B. Griffin ◽

Amer Heider ◽

Andrea Amalfitano ◽

...

Keyword(s):

Trna Synthetase ◽

Compound Heterozygosity ◽

Loss Of Function ◽

Aminoacyl Trna Synthetase ◽

Related Disease

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Somatic Mutations in Vascular Malformations of Hereditary Hemorrhagic Telangiectasia Result in Biallelic Loss of ENG or ACVRL1

10.1101/731588 ◽

2019 ◽

Cited By ~ 2

Author(s):

Daniel Snellings ◽

Carol Gallione ◽

Dewi Clark ◽

Nicholas Vozoris ◽

Marie Faughnan ◽

...

Keyword(s):

Hereditary Hemorrhagic Telangiectasia ◽

Somatic Mutations ◽

Vascular Malformations ◽

Low Frequency ◽

The Body ◽

Mendelian Disease ◽

Loss Of Function ◽

Long Read ◽

In Trans ◽

Vascular Defect

AbstractHereditary Hemorrhagic Telangiectasia (HHT) is a Mendelian disease characterized by vascular malformations including visceral arteriovenous malformations and mucosal telangiectasia. HHT is caused by loss-of-function mutations in one of 3 genes; ENG, ACVRL1 or SMAD4 and is inherited as an autosomal dominant condition. Intriguingly, the constitutional mutation causing HHT is present throughout the body, yet the multiple vascular malformations in HHT patients occur focally, rather than manifesting as a systemic vascular defect. This disconnect between genotype and phenotype suggests that a local event is necessary for the development of vascular malformations. We investigated the hypothesis that local somatic mutations seed the formation HHT-related telangiectasia in a genetic two-hit mechanism. We identified low-frequency somatic mutations in 9/19 telangiectasia using high depth next-generation sequencing. We established phase for 7 of 9 samples using long-read sequencing, which confirm that the germline and somatic mutations in all 7 samples exist in trans configuration; consistent with a genetic two-hit mechanism. These combined data suggest that biallelic loss of ENG or ACVRL1 may be a required event in the development of telangiectasia, and that rather than haploinsufficiency, vascular malformations in HHT are caused by a Knudsonian two-hit mechanism.

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mRNA processing in mutant zebrafish lines generated by chemical and CRISPR-mediated mutagenesis produces potentially functional transcripts

10.1101/154856 ◽

2017 ◽

Cited By ~ 1

Author(s):

Jennifer L Anderson ◽

Timothy S Mulligan ◽

Meng-Chieh Shen ◽

Hui Wang ◽

Catherine M Scahill ◽

...

Keyword(s):

Gene Function ◽

Reverse Genetics ◽

Model Organism ◽

Low Frequency ◽

Mrna Processing ◽

Null Alleles ◽

Reverse Genetic ◽

Loss Of Function ◽

Genomic Change ◽

Frame Shift

AbstractAs model organism-based research shifts from forward to reverse genetics approaches, largely due to the ease of genome editing technology, allow frequency of abnormal phenotypes is being observed in lines with mutations predicted to lead to deleterious effects on the encoded protein. In zebrafish, this low frequency is in part explained by compensation by genes of redundant or similar function, often resulting from the additional round of teleost-specific whole genome duplication within vertebrates. Here we offer additional explanations for the low frequency of mutant phenotypes. We analyzed mRNA processing in seven zebrafish lines with mutations expected to disrupt gene function, generated by CRISPR/Cas9 or ENU mutagenesis methods. Five of the seven lines showed evidence of genomic compensation by means of altered mRNA processing: one through a skipped exon that did not lead to a frame shift, one through nonsense-associated splicing that did not lead to a frame shift, and three through the use of cryptic splice sites. These results highlight the need for a methodical analysis of the mRNA produced in mutant lines before making conclusions or embarking on studies that assume loss of function as a result of a given genomic change. Furthermore, recognition of the types of genomic adaptations that can occur may inform the strategies of mutant generation.Author summaryThe recent rise of reverse genetic, gene targeting methods has allowed researchers to readily generate mutations in any gene of interest with relative ease. Should these mutations have the predicted effect on the mRNA and encoded protein, we would expect many more abnormal phenotypes than are typically being seen in reverse genetic screens. Here we set out to explore some of the reasons for this discrepancy by studying seven separate mutations in zebrafish. We present evidence that thorough cDNA sequence analysis is a key step in assessing the likelihood that a given mutation will produce hypomorphic or null alleles. This study reveals that alternative mRNA processing in the mutant background often produces transcripts that escape nonsense-mediated decay, thereby potentially preserving gene function. By understanding the ways that cells avoid the deleterious consequences of mutations, researchers can better design reverse genetic strategies to increase the likelihood of gene disruption.

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ATRT-11. PREVALENCE OF GERMLINE VARIANTS IN SMARCB1 INCLUDING SOMATIC MOSAICISM IN AT/RT AND OTHER RHABDOID TUMORS

Neuro-Oncology ◽

10.1093/neuonc/noaa222.011 ◽

2020 ◽

Vol 22 (Supplement_3) ◽

pp. iii277-iii278

Author(s):

Ryota Shirai ◽

Tomoo Osumi ◽

Keita Terashima ◽

Chikako Kiyotani ◽

Meri Uchiyama ◽

...

Keyword(s):

Direct Sequencing ◽

Snp Array ◽

Somatic Mosaicism ◽

Low Frequency ◽

Rhabdoid Tumor ◽

Loss Of Function ◽

Germline Mosaicism ◽

Germline Variants ◽

Predisposition Genes ◽

Rhabdoid Tumors

Abstract BACKGROUND Genetic hallmark of atypical teratoid/rhabdoid tumor (AT/RT) is loss-of-function variants or deletions in SMARCB1 gene on 22q11.2 chromosome, which is common to extracranial malignant rhabdoid tumors (MRT). Previous studies demonstrated that approximately one-thirds of AT/RT and extracranial MRT patients harbored germline SMARCB1 variants as the rhabdoid tumor predisposing syndrome. We studied herein intensive analysis of the SMARCB1 gene in AT/RT and extracranial MRT patients focusing on prevalence of germline genetic variants. PROCEDURE: In total, 16 patients were included. Both tumor-derived DNA and germline DNA were obtained from all patients. First, screening for SMARCB1 alterations in the tumor specimens was done by direct sequencing, ddPCR and SNP array analysis. Then, analysis of germline DNA samples focusing on the genomic abnormalities detected in the paired tumors in each case was performed. RESULTS In eight of 16 cases (50%), genomic alterations observed in the tumor-derived DNA were also detected in the germline DNA. It is worth noting that three patients had germline mosaicism. Two of three patients had mosaic deletion, including SMARCB1 region, and the average copy number of the deleted region in the SMARCB1 gene in the germline was 1.60 and 1.76. For another patient, the fraction of SMARCB1 variants in normal cells was as low as 1.7%. CONCLUSIONS Approximately half the MRT cases in this study had SMARCB1 germline alterations. Considering the presence of low-frequency mosaicisms which conventional methods might overlook, inherited germline variants in predisposition genes are more important than previously assumed for the pathogenesis of pediatric cancers.

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The reversion variant (p.Arg90Leu) at the evolutionarily adaptive p.Arg90 site in CELA3B predisposes to chronic pancreatitis

10.1101/2020.11.28.20240135 ◽

2020 ◽

Author(s):

Emmanuelle Masson ◽

Vinciane Rebours ◽

Louis Buscail ◽

Frédérique Frete ◽

Mael Pagenault ◽

...

Keyword(s):

Chronic Pancreatitis ◽

Amino Acid ◽

Human Genome ◽

Low Frequency ◽

Missense Variant ◽

Loss Of Function ◽

Ancestral Allele ◽

Functional Effect ◽

Gain Of Function ◽

Missense Variants

ABSTRACTA gain-of-function missense variant in the CELA3B gene, p.Arg90Cys (c.268C>T), has recently been reported to cause pancreatitis in an extended pedigree. Herein, we sequenced the CELA3B gene in 644 genetically unexplained French chronic pancreatitis (CP) patients (all unrelated) and 566 controls. No predicted loss-of-function variants were identified. None of the six low frequency or common missense variants detected showed significant association with CP. Nor did the aggregate rare/very rare missense variants (n=14) show any significant association with CP. However, p.Arg90Leu (c.269G>T), which was found in 4 patients but no controls and affects the same amino acid as p.Arg90Cys, serves to revert p.Arg90 to the human elastase ancestral allele. Since p.Arg90Leu has previously been shown to exert a similar functional effect to p.Arg90Cys, our findings not only confirm the involvement of CELA3B in the etiology of CP but also pinpoint a new evolutionarily adaptive site in the human genome.

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