scholarly journals A Nonsense Variant in CCDC65 Gene Causes Respiratory Failure Associated with Increased Lamb Mortality in French Lacaune Dairy Sheep

Genes ◽  
2021 ◽  
Vol 13 (1) ◽  
pp. 45
Author(s):  
Ben Braiek ◽  
Carole Moreno-Romieux ◽  
Charlotte Allain ◽  
Philippe Bardou ◽  
Arnaud Bordes ◽  
...  
Keyword(s):  
Stop Codon ◽  
Coiled Coil ◽  
Premature Stop Codon ◽  
Variant Calling ◽  
Dairy Sheep ◽  
Single Nucleotide ◽  
Respiratory Problems ◽  
Regulatory Complex ◽  
Heterozygous Carriers ◽  
Lung Lobes

We recently demonstrated that the Lacaune deficient homozygous haplotype 6 (LDHH6) potentially hosts a recessive perinatal lethal mutation in Lacaune dairy sheep mapped on OAR3. In the present study, we have analyzed the whole-genome sequences of two Lacaune ram heterozygous carriers of LDHH6. After variant calling and filtering against the variants of 86 non-carrier rams, we have identified a single nucleotide variant (SNV) in the two LDHH6 carriers whose variant allele induced a premature stop codon (p.Glu111*) in the Coiled-Coil Domain Containing 65 (CCDC65) gene. CCDC65 is involved in the assembly of the nexin-dynein regulatory complex for the formation of microtubules in ciliated cells. In order to identify the phenotype in homozygous sheep, we generated at-risk matings (n = 17) between rams and ewes heterozygous for the candidate variant in CCDC65. A total of 16 lambs were born alive with five genotyped as homozygous carriers. The homozygous lambs suffered from respiratory problems, and four of them died within the first month of life. At necropsy, we observed a broad hepatization of lung lobes possibly induced by infectious pneumonia. The management of this lethal recessive allele (frequency of 0.06) through reasoned mating in the Lacaune sheep selection schemes could reduce lamb mortality by 2%.

scholarly journals A Novel Truncating Mutation in HOMER2 Causes Nonsyndromic Progressive DFNA68 Hearing Loss in a Spanish Family

Genes ◽  
2021 ◽  
Vol 12 (3) ◽  
pp. 411
Author(s):  
María Lachgar ◽  
Matías Morín ◽  
Manuela Villamar ◽  
Ignacio del Castillo ◽  
Miguel Ángel Moreno-Pelayo
Keyword(s):  
Hearing Loss ◽  
Stop Codon ◽  
Coiled Coil ◽  
Premature Stop Codon ◽  
Scaffolding Protein ◽  
Common Mechanism ◽  
Chinese Family ◽  
Truncating Mutation ◽  
Spanish Family ◽  
Sensory Defect

Nonsyndromic hereditary hearing loss is a common sensory defect in humans that is clinically and genetically highly heterogeneous. So far, 122 genes have been associated with this disorder and 50 of them have been linked to autosomal dominant (DFNA) forms like DFNA68, a rare subtype of hearing impairment caused by disruption of a stereociliary scaffolding protein (HOMER2) that is essential for normal hearing in humans and mice. In this study, we report a novel HOMER2 variant (c.832_836delCCTCA) identified in a Spanish family by using a custom NGS targeted gene panel (OTO-NGS-v2). This frameshift mutation produces a premature stop codon that may lead in the absence of NMD to a shorter variant (p.Pro278Alafs*10) that truncates HOMER2 at the CDC42 binding domain (CBD) of the coiled-coil structure, a region that is essential for protein multimerization and HOMER2-CDC42 interaction. c.832_836delCCTCA mutation is placed close to the previously identified c.840_840dup mutation found in a Chinese family that truncates the protein (p.Met281Hisfs*9) at the CBD. Functional assessment of the Chinese mutant revealed decreased protein stability, reduced ability to multimerize, and altered distribution pattern in transfected cells when compared with wild-type HOMER2. Interestingly, the Spanish and Chinese frameshift mutations might exert a similar effect at the protein level, leading to truncated mutants with the same Ct aberrant protein tail, thus suggesting that they can share a common mechanism of pathogenesis. Indeed, age-matched patients in both families display quite similar hearing loss phenotypes consisting of early-onset, moderate-to-profound progressive hearing loss. In summary, we have identified the third variant in HOMER2, which is the first one identified in the Spanish population, thus contributing to expanding the mutational spectrum of this gene in other populations, and also to clarifying the genotype–phenotype correlations of DFNA68 hearing loss.

scholarly journals Protection against Fatal Sindbis Virus Encephalitis by Beclin, a Novel Bcl-2-Interacting Protein

1998 ◽  
Vol 72 (11) ◽  
pp. 8586-8596 ◽  
Author(s):  
Xiao Huan Liang ◽  
Linda K. Kleeman ◽  
Hui Hui Jiang ◽  
Gerald Gordon ◽  
James E. Goldman ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Sindbis Virus ◽  
Stop Codon ◽  
Coiled Coil ◽  
Resonance Energy Transfer ◽  
Premature Stop Codon ◽  
Resonance Energy ◽  
Cellular Protein ◽  
Interacting Protein ◽  
Induced Apoptosis

ABSTRACT bcl-2, the prototypic cellular antiapoptotic gene, decreases Sindbis virus replication and Sindbis virus-induced apoptosis in mouse brains, resulting in protection against lethal encephalitis. To investigate potential mechanisms by which Bcl-2 protects against central nervous system Sindbis virus infection, we performed a yeast two-hybrid screen to identify Bcl-2-interacting gene products in an adult mouse brain library. We identified a novel 60-kDa coiled-coil protein, Beclin, which we confirmed interacts with Bcl-2 in mammalian cells, using fluorescence resonance energy transfer microscopy. To examine the role of Beclin in Sindbis virus pathogenesis, we constructed recombinant Sindbis virus chimeras that express full-length human Beclin (SIN/beclin), Beclin lacking the putative Bcl-2-binding domain (SIN/beclinΔBcl-2BD), or Beclin containing a premature stop codon near the 5′ terminus (SIN/beclinstop). The survival of mice infected with SIN/beclin was significantly higher (71%) than the survival of mice infected with SIN/beclinΔBcl-2BD (9%) or SIN/beclinstop (7%) (P < 0.001). The brains of mice infected with SIN/beclin had fewer Sindbis virus RNA-positive cells, fewer apoptotic cells, and lower viral titers than the brains of mice infected with SIN/beclinΔBcl-2BD or SIN/beclinstop. These findings demonstrate that Beclin is a novel Bcl-2-interacting cellular protein that may play a role in antiviral host defense.

A Frameshift Mutation in the Human Fibrinogen Aα-chain Gene (Aα [499] Ala Frameshift Stop) Leading to Dysfibrinogen San Giovanni Rotondo

2001 ◽  
Vol 86 (12) ◽  
pp. 1483-1488 ◽  
Author(s):  
Gennaro Vecchione ◽  
Rosa Santacroce ◽  
Francesca D’Angelo ◽  
Bruno Casetta ◽  
Maria Luisa Papa ◽  
...  
Keyword(s):  
Mass Spectrum ◽  
Dna Analysis ◽  
Stop Codon ◽  
Premature Stop Codon ◽  
Disulfide Bridge ◽  
Chain Gene ◽  
Human Fibrinogen ◽  
Single Nucleotide ◽  
Sds Page

SummaryWe have investigated a 53-yr-old asymptomatic white man with decreased functional, but not immunologic, fibrinogen plasma levels together with prolonged thrombin and reptilase times, detected through routine coagulation studies prior to a surgical procedure. A new heterozygous single nucleotide deletion (C) at position Ala499 within the Aα-chain gene was identified, which predicted changes of the corresponding aminoacids encoded by the subsequent portion of the exon V and the appearance of a premature stop codon at position 518 (A [499]Ala frameshift stop). The new dysfunctional fibrinogen, San Giovanni Rotondo variant, was confirmed in vivo by SDS-PAGE analysis of HPLC-purified fibrinogen chains. Mass spectrum examination of the abnormal HPLC-purified peak gave an estimated mass (56,088 Da) similar to that predicted by DNA analysis of the mutated Aα-chain gene (56,088 Da) and, after tryptic digestion, the truncated Aα-chain was shown only in the propositus, who also carried normal Aα-chain. In addition, mass spectrum analysis of the tryptic digest of the abnormal chain confirmed the presence of a new and unpaired cysteine at the last position that was predicted to form a disulfide bridge with human serum albumin. Immuno-blot analysis confirmed that fibrinogen San Giovanni Rotondo variant, but not normal fibrinogen, contained substantial amounts of albumin. Present findings confirm that truncated Aα-chain lacking part of the terminal domain may be incorporated into mature fibrinogen molecules and normally secreted in the bloodstream.

scholarly journals Fine Mapping of a Grain Shape Gene from a Rice Landrace Longliheinuo-Dwarf (Oryza sativa L. ssp. japonica)

Agronomy ◽  
2020 ◽  
Vol 10 (3) ◽  
pp. 380 ◽  
Author(s):  
Fei Shang ◽  
Xu Chao ◽  
Kaiwen Meng ◽  
Xianghe Meng ◽  
Qin Li ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Grain Shape ◽  
Agronomic Traits ◽  
Stop Codon ◽  
Oryza Sativa L ◽  
Premature Stop Codon ◽  
Nucleotide Polymorphisms ◽  
Genetic Population ◽  
Single Nucleotide

Identification of grain shape genes can facilitate breeding of rice cultivars with optimal grain shape and appearance quality. In this study, we selected two rice germplasms, namely Longliheinuo-dwarf (LH) and N643, with different grain shape, to construct a genetic population for quantitative trait locus (QTL) analysis. A major QTL (qGS7), controlling the ratio of grain length to grain width, was mapped on the chromosome 7 in a BC1F4 line. By high-resolution linkage analysis, qGS7 was delimited to a 52.8 kb region including eight predicted genes. Through sequence alignment and real-time PCR expression analysis of these ORFs, ORF3 (LOC_Os07g42410) was selected as the candidate gene for further analysis. Single nucleotide polymorphisms (SNP) diversity analysis of ORF3 revealed that a single nucleotide deletion in the 7th exon resulted in a frameshift in parent LH and the parent in which a premature stop codon was identified. It was a rare mutation that caused grain shape difference. Real-time PCR analyses showed that the expression characteristics of ORF3 was in accordance with the development of spikelets. Of the 18 agronomic traits investigation in qGS7 near isogenic lines (NILs) showed that qGS7 not only changed grain shape but also affected plant height, panicle curvature, panicle length, the length of second leaf from the top, and chalkiness.

scholarly journals Revelation by Single-Nucleotide Polymorphism Genotyping That Mutations Leading to a Premature Stop Codon in inlA Are Common among Listeria monocytogenes Isolates from Ready-To-Eat Foods but Not Human Listeriosis Cases

2010 ◽  
Vol 76 (9) ◽  
pp. 2783-2790 ◽  
Author(s):  
A. Van Stelten ◽  
J. M. Simpson ◽  
T. J. Ward ◽  
K. K. Nightingale
Keyword(s):  
United States ◽  
Single Nucleotide Polymorphism ◽  
Listeria Monocytogenes ◽  
Stop Codon ◽  
Human Health Risk ◽  
Premature Stop Codon ◽  
The United States ◽  
Nucleotide Polymorphism ◽  
Pathogenic Potential ◽  
Single Nucleotide

ABSTRACT Listeria monocytogenes utilizes internalin A (InlA; encoded by inlA) to cross the intestinal barrier to establish a systemic infection. Multiple naturally occurring mutations leading to a premature stop codon (PMSC) in inlA have been reported worldwide, and these mutations are causally associated with attenuated virulence. Five inlA PMSC mutations recently discovered among isolates from France and the United States were included as additional markers in our previously described inlA single-nucleotide polymorphism (SNP) genotyping assay. This assay was used to screen >1,000 L. monocytogenes isolates from ready-to-eat (RTE) foods (n = 502) and human listeriosis cases (n = 507) for 18 inlA PMSC mutations. A significantly (P < 0.0001) greater proportion of RTE food isolates (45.0%) carried a PMSC mutation in inlA compared to human clinical isolates (5.1%). The proportion of L. monocytogenes with or without PMSC mutations in inlA was similar among isolates from different RTE food categories except for deli meats, which included a marginally higher proportion (P = 0.12) of isolates carrying a PMSC in inlA. We also analyzed the distribution of epidemic clone (EC) strains, which have been linked to the majority of listeriosis outbreaks worldwide and are overrepresented among sporadic cases in the United States. We observed a significant (P < 0.05) overrepresentation of EC strains in deli and seafood salads and a significant (P < 0.05) underrepresentation of EC strains in smoked seafood. These results provide important data to predict the human health risk of exposure to L. monocytogenes strains that differ in pathogenic potential through consumption of contaminated RTE foods.

scholarly journals Development and Implementation of a Multiplex Single-Nucleotide Polymorphism Genotyping Assay for Detection of Virulence-Attenuating Mutations in the Listeria monocytogenes Virulence-Associated Gene inlA

2008 ◽  
Vol 74 (23) ◽  
pp. 7365-7375 ◽  
Author(s):  
A. Van Stelten ◽  
K. K. Nightingale
Keyword(s):  
Single Nucleotide Polymorphism ◽  
Listeria Monocytogenes ◽  
Stop Codon ◽  
Premature Stop Codon ◽  
Snp Genotyping ◽  
Regulatory Agencies ◽  
Nucleotide Polymorphism ◽  
Single Nucleotide ◽  
Genotyping Assay ◽  
Virulence Attenuation

ABSTRACT The virulence factor internalin A (InlA) facilitates the uptake of Listeria monocytogenes by epithelial cells that express the human isoform of E-cadherin. Previous studies identified naturally occurring premature stop codon (PMSC) mutations in inlA and demonstrated that these mutations are responsible for virulence attenuation. We assembled >1,700 L. monocytogenes isolates from diverse sources representing 90 EcoRI ribotypes. A subset of this isolate collection was selected based on ribotype frequency and characterized by a Caco-2 cell invasion assay. The sequencing of inlA genes from isolates with attenuated invasion capacities revealed three novel inlA PMSCs which had not been identified previously among U.S. isolates. Since ribotypes include isolates with and without inlA PMSCs, we developed a multiplex single-nucleotide polymorphism (SNP) genotyping assay to detect isolates with virulence-attenuating PMSC mutations in inlA. The SNP genotyping assay detects all inlA PMSC mutations that have been reported worldwide and verified in this study to date by the extension of unlabeled primers with fluorescently labeled dideoxynucleoside triphosphates. We implemented the SNP genotyping assay to characterize human clinical and food isolates representing common ribotypes associated with novel inlA PMSC mutations. PMSCs in inlA were significantly (ribotypes DUP-1039C and DUP-1045B; P < 0.001) or marginally (ribotype DUP-1062D; P = 0.11) more common among food isolates than human clinical isolates. SNP genotyping revealed a fourth novel PMSC mutation among U.S. L. monocytogenes isolates, which was observed previously among isolates from France and Portugal. This SNP genotyping assay may be implemented by regulatory agencies and the food industry to differentiate L. monocytogenes isolates carrying virulence-attenuating PMSC mutations in inlA from strains representing the most significant health risk.

scholarly journals A novel LRP6 variant in a Japanese family with oligodontia

2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Hiroki Goto ◽  
Masashi Kimura ◽  
Junichiro Machida ◽  
Akiko Ota ◽  
Mitsuko Nakashima ◽  
...  
Keyword(s):  
Stop Codon ◽  
Premature Stop Codon ◽  
Japanese Patients ◽  
Tooth Agenesis ◽  
Japanese Family ◽  
Single Nucleotide ◽  
Extracellular Region ◽  
Wnt Ligands ◽  
Nucleotide Insertion ◽  
Exome Sequence

AbstractCongenital tooth agenesis is a common anomaly in human development. We performed exome sequence analysis of genomic DNA collected from Japanese patients with tooth agenesis and their relatives. We found a novel single-nucleotide insertion in the LRP6 gene, the product of which is involved in Wnt/β-catenin signaling as a coreceptor for Wnt ligands. The single-nucleotide insertion results in a premature stop codon in the extracellular region of the encoded protein.

scholarly journals Single nucleotide replacement in the Atlantic salmon genome using CRISPR/Cas9 and asymmetrical oligonucleotide donors

BMC Genomics ◽  
2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Anne Hege Straume ◽  
Erik Kjærner-Semb ◽  
Kai Ove Skaftnesmo ◽  
Hilal Güralp ◽  
Simon Lillico ◽  
...  
Keyword(s):  
Atlantic Salmon ◽  
Generation Time ◽  
Genetic Improvement ◽  
Selective Breeding ◽  
Stop Codon ◽  
Premature Stop Codon ◽  
Single Amino Acid ◽  
Single Nucleotide ◽  
Homology Directed Repair ◽  
New Breeding Technologies

Abstract Background New breeding technologies (NBT) using CRISPR/Cas9-induced homology directed repair (HDR) has the potential to expedite genetic improvement in aquaculture. The long generation time in Atlantic salmon makes breeding an unattractive solution to obtain homozygous mutants and improving the rates of perfect HDR in founder (F0) fish is thus required. Genome editing can represent small DNA changes down to single nucleotide replacements (SNR). This enables edits such as premature stop codons or single amino acid changes and may be used to obtain fish with traits favorable to aquaculture, e.g. disease resistance. A method for SNR has not yet been demonstrated in salmon. Results Using CRISPR/Cas9 and asymmetrical ODNs, we were able to perform precise SNR and introduce a premature stop codon in dnd in F0 salmon. Deep sequencing demonstrated up to 59.2% efficiency in single embryos. In addition, using the same asymmetrical ODN design, we inserted a FLAG element into slc45a2 and dnd, showing high individual perfect HDR efficiencies (up to 36.7 and 32.7%, respectively). Conclusions In this work, we demonstrate that precise SNR and knock-in (KI) can be performed in F0 salmon embryos using asymmetrical oligonucleotide (ODN) donors. We suggest that HDR-induced SNR can be applied as a powerful NBT, allowing efficient introgression of favorable alleles and bypassing challenges associated with traditional selective breeding.

Ectopic Transcript Analysis Indicates that Allelic Exclusion is an Important Cause of Type I Protein C Deficiency in Patients with Nonsense and Frameshift Mutations in the PROC Gene

1996 ◽  
Vol 75 (06) ◽  
pp. 870-876 ◽  
Author(s):  
José Manuel Soria ◽  
Lutz-Peter Berg ◽  
Jordi Fontcuberta ◽  
Vijay V Kakkar ◽  
Xavier Estivill ◽  
...  
Keyword(s):  
Splice Site ◽  
Protein C ◽  
Stop Codon ◽  
Premature Stop Codon ◽  
Allelic Exclusion ◽  
Polymorphism Analysis ◽  
Type I ◽  
Protein C Deficiency ◽  
Nonsense Mutations ◽  
Stop Codons

SummaryNonsense mutations, deletions and splice site mutations are a common cause of type I protein C deficiency. Either directly or indirectly by altering the reading frame, these' lesions generate or may generate premature stop codons and could therefore be expected to result in premature termination of translation. In this study, the possibility that such mutations could instead exert their pathological effects at an earlier stage in the expression pathway, through “allelic exclusion” at the RNA level, was investigated. Protein C (PROC) mRNA was analysed in seven Spanish type I protein C deficient patients heterozygous for two nonsense mutations, a 7bp deletion, a 2bp insertion and three splice site mutations. Ectopic RNA transcripts from patient and control lymphocytes were analysed by RT-PCR and direct sequencing of amplified PROC cDNA fragments. The nonsense mutations and the deletion were absent from the cDNAs indicating that only mRNA derived from the normal allele had been expressed. Similarly for the splice site mutations, only normal PROC cDNAs were obtained. In one case, exclusion of the mutated allele could be confirmed by polymorphism analysis. In contrast to these six mutations, the 2 bp insertion was not associated with loss of mRNA from the mutated allele. In this case, cDNA analysis revealed the absence of 19 bases from the PROC mRNA consistent with the generation and utilization of a cryptic splice site 3’ to the site of mutation, which would result in a frameshift and a premature stop codon. It is concluded that allelic exclusion is a common causative mechanism in those cases of type I protein C deficiency which result from mutations that introduce premature stop codons

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