scholarly journals Germline Variants of CYBA and TRPM4 Predispose to Familial Colorectal Cancer

2021 ◽  
Author(s):  
Lizhen Zhu ◽  
Beiping Miao ◽  
Dagmara Dymerska ◽  
Magdalena Kuświk ◽  
Elena Bueno-Martínez ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Stop Codon ◽  
Bacterial Colonization ◽  
Functional Characterization ◽  
Premature Stop Codon ◽  
Familial Colorectal Cancer ◽  
Intestinal Mucus ◽  
Healthy Elderly ◽  
Predisposing Genes ◽  
Mucus Barrier

Familial colorectal cancer (CRC) is only partially explained by known germline predisposing genes. We performed whole genome sequencing in 15 Polish families of many affected individuals, without mutations in known CRC predisposing genes. We focused on loss-of-function variants and functionally characterized them. We identified a frameshift variant in the CYBA gene (c.246delC) in one family and a splice site variant in the TRPM4 gene (c.25-1 G>T) in another family. While both variants were absent or extremely rare in gene variant databases, we identified four additional Polish familial CRC cases and two healthy elderly individuals with the CYBA variant (odds ratio 2.46, 95% confidence interval 0.48-12.69). Both variants led to a premature stop codon and to a truncated protein. Functional characterization of the variants showed that knockdown of CYBA or TRPM4 depressed generation of reactive oxygen species (ROS) in LS174T and HT-29 cell lines. Knockdown of TRPM4 resulted in decreased MUC2 protein production. CYBA encodes a component in the NADPH oxidase system which generates ROS and controls, e.g., bacterial colonization in the gut. Germline CYBA variants are associated with early onset inflammatory bowel disease, supported with experimental evidence on loss of intestinal mucus barrier function due to ROS deficiency. TRPM4 encodes a calcium-activated ion channel, which in a human colonic cancer cell line controls calcium-mediated secretion of MUC2, a major component of intestinal mucus barrier. We suggest that the gene defects in CYBA and TRPM4 mechanistically involve intestinal barrier integrity through ROS and mucus biology, which converges in chronic bowel inflammation.

scholarly journals POS0352 ALLELIC VARIANTS IN THE ABCG2 GENE CAN LEAD TO HYPERURICEMIA AND GOUT

2021 ◽  
Vol 80 (Suppl 1) ◽  
pp. 406.2-407
Author(s):  
K. Pavelcova ◽  
J. Bohata ◽  
B. Stiburkova
Keyword(s):  
Uric Acid ◽  
Rare Variants ◽  
Stop Codon ◽  
Functional Characterization ◽  
Premature Stop Codon ◽  
European Population ◽  
Allelic Variants ◽  
Abcg2 Protein ◽  
The Impact ◽  
Abcg2 Gene

Background:The level of uric acid is largely determined by the functions of urate transporters, which are located in the kidney and intestine. The ABCG2 protein is the major excretor of uric acid and its dysfunction may lead to the development of hyperuricemia and gout.Objectives:The aim of our study was to detect the occurrence and frequency of allelic variants in the ABCG2 gene that can lead to impaired function of the ABCG2 protein and to the development of hyperuricemia and gout.Methods:We examined allelic variants of ABCG2 using PCR amplification and Sanger sequencing of all coding regions and exon-intron boundaries in 359 patients with primary hyperuricemia and gout.Results:We found a rare in-frame deletion p.K360del and 15 missense variants, two of which were common (p.V12M, p.Q141K) and 13 were very rare (p.M71V, p.G74D, p.M131I, p.R147W, p.T153M, p.I242T, p.R236X, p.F373C, p.T421A, p.T434M, p.S476P, p.S572R, p.D620N). The p.R236X variant leads to a premature stop codon. The p.V12M variant probably has a protective effect against gout (minor allele frequency – MAF – in our cohort = 0,025 / MAF in the European population = 0,061), while the p.Q141K variant increases the risk of gout (MAF in our cohort = 0,213 / MAF in the European population = 0,094) (1). As for the rare variants, the p.R147W, p.T153M, p.F373C, p.T434M, p.S476P and p.S572R according to functional analyzes reduce the function of the ABCG2 protein (2). Based on in silico prediction, the impact on reduced function is expected for variants p.M71V, p.G74D, p.M131I, p.R147W, p.I242T, p.F373C, p.T434M, p.S476P and p.S572R.Conclusion:Our data suggest that the common variant p.Q141K and most of the rare variants in the ABCG2 gene affect the function of the ABCG2 urate transporter and are a genetic risk factor for hyperuricemia and gout.References:[1]Stiburkova B, et al. Functional non-synonymous variants of ABCG2 and gout risk. Rheumatology (Oxford). 2017 Nov 1; 56(11):1982-1992.[2]Toyoda Y, et al. Functional characterization of clinically-relevant rare variants in ABCG2 identified in a gout and hyperuricemia cohort. Cells. 2019 Apr 18;8(4).Acknowledgements:This study was supported by the project for conceptual development of research organization 00023728 (Institute of Rheumatology) and RVO VFN64165.Disclosure of Interests:None declared

scholarly journals A commensal adhesin enhances E. coli retention by mucin, while mucin desulfation by mucin-foraging bacteria enhances its transmigration through the mucus barrier

2017 ◽  
Author(s):  
Fitua Al-Saedi ◽  
Diana Pereira Vaz ◽  
Daniel H Stones ◽  
Anne Marie Krachler
Keyword(s):  
Bacterial Adhesion ◽  
Bacterial Colonization ◽  
Cell Entry ◽  
Dependent Manner ◽  
Epithelial Surface ◽  
E Coli ◽  
Intestinal Mucus ◽  
Receptor Interactions ◽  
Protein Receptors ◽  
Mucus Barrier

ABSTRACTBacterial adhesion to host receptors is an early and essential step in bacterial colonization, and the nature of adhesion-receptor interactions determines bacterial localization and thus the outcome of these interactions. Here, we determine the host receptors for the Multivalent Adhesion Molecule from the gut commensal E. coli HS (MAMHS), which contains an array of seven mammalian cell entry (MCE) domains. The MAMHS adhesin interacts with a range of host receptors, through recognition of a shared 3-O-sulfo-galactosyl moiety. This functional group is also found in mucin, a component of the intestinal mucus layer and thus one of the prime adherence targets for commensal E. coli. Mucin gels impede the motility of E. coli by acting as a physical barrier, and the barrier effect is enhanced by specific interactions between mucin and MAMHS in a sulfation-dependent manner. Desulfation of mucin by pure sulfatase or the sulfatase-producing commensal Bacteroides thetaiotaomicron decreases binding of E. coli to mucin and increases attachment of bacteria to the epithelial surface, through interactions with surface-localized sulfated lipid and protein receptors. Together, our results demonstrate that the E. coli adhesin MAMHS facilitates retention of a gut commensal by mucin, and suggest that the amount of sulfatase secreted by mucin-foraging bacteria inhabiting the same niche, such as B. thetaiotaomicron, may affect the capacity of the mucus barrier to retain commensal E. coli.

A New Germline Stop Codon Mutation in Exon 15 of the APC Gene Predisposing to Familial Adenomatous Polyposis

2013 ◽  
Vol 28 (4) ◽  
pp. 405-408 ◽  
Author(s):  
Laura Schirosi ◽  
Marcello Pellegrino ◽  
Paolo Tarantino ◽  
Salvatore Mauro ◽  
Andrea Tinelli ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Familial Adenomatous Polyposis ◽  
Germline Mutation ◽  
Stop Codon ◽  
Premature Stop Codon ◽  
Colorectal Tumor ◽  
Apc Gene ◽  
Sequencing Analysis ◽  
Adenomatous Polyposis ◽  
Truncated Protein

Familial adenomatous polyposis (FAP) is an autosomal dominant disorder related to germline mutations of the adenomatous polyposis coli ( APC) gene. It is characterized by the detection of numerous adenomatous polyps that, if untreated, develop into colorectal cancer. We studied an Italian family with FAP history and the related colorectal tumor sample of the proband. Sequencing analysis of blood samples revealed the presence of a never-reported germline mutation in the APC gene (exon 15): an heterozygous G deletion at position c.2126 resulting in a premature stop codon (p.Gly721GlufsX6) and in a truncated protein. This mutation was also identified in the colorectal tumor tissue, together with a second known pathogenic heterozygotic somatic mutation, c.4348C>T (p.Arg1450X), which generates a premature truncated protein. The novel identified germline mutation is therefore related to FAP and, in accordance with Knudson's “two hit” hypothesis, can be considered the first event predisposing to the insurgence of colorectal cancer in these patients. The somatic hit inactivating the second allele of the APC gene is located in the mutation cluster region of the gene; this is not a random event since it depends on the position of the germline mutation. The inactivation of APC generates the neoplastic growth advantage to the cell.

scholarly journals Genetic analysis and functional characterization of prothrombins Corpus Christi (Arg382-Cys), Dhahran (Arg271-His), and hypoprothrombinemia

Blood ◽  
1996 ◽  
Vol 88 (7) ◽  
pp. 2611-2618 ◽  
Author(s):  
AS O'Marcaigh ◽  
WL Nichols ◽  
NL Hassinger ◽  
JD Mullins ◽  
AA Mallouh ◽  
...  
Keyword(s):  
Stop Codon ◽  
Functional Characterization ◽  
Family Members ◽  
Premature Stop Codon ◽  
Factor Xa ◽  
Nucleotide Position ◽  
Cpg Dinucleotides ◽  
Corpus Christi ◽  
Normal Platelet ◽  
Fibrinogen Binding

The structural abnormalities and functional characteristics of dysfunctional prothrombin variants in two new kindreds have been determined. Prothrombin Corpus Christi (family 1) was purified and found to have markedly reduced fibrinogen clotting activity, yet normal amidolytic and near-normal platelet aggregating activity. A transition (C to T) at nucleotide position 8885, present in the heterozygous form in affected family members, resulted in the substitution of Cys for Arg 382. This substitution results in the loss of a positive charge within the fibrinogen-binding exosite of thrombin, thus accounting for the observed functional defect. A heterozygous C to T transition was also present at position 19994 in other family members with a hypoprothrombinemic phenotype. This mutation results in the replacement of Gln 541 (CAA) by a premature stop codon (TAA). Prothrombin Dhahran (family 2) was found to have markedly reduced fibrinogen clotting activity, but normal amidolytic activity. Affected family members were found to have a G to A transition at nucleotide position 7312 resulting in the substitution of His for Arg 271. This substitution results in the abolition of a factor Xa cleavage site, yielding meizothrombin rather than thrombin, on activation of prothrombin Dhahran by factor Xa. All but one of the above mutations occur at CpG dinucleotides, thus further supporting the observation of a high incidence of CpG transitions in hereditary dysprothrombinemia. The significant bleeding tendencies of individuals homozygous for prothrombin Dhahran (prothrombin clotting activity 5% to 7%) contrast sharply with the absence of significant chronic bleeding in the proband expressing prothrombin Corpus Christi (prothrombin clotting activity 2%). Our findings underscore the capacity of thrombin to contribute to clinical hemostasis by mechanisms other than its fibrinogen clotting activity.

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

10 THE ROLE OF DIETARY L-SERINE IN THE REGULATION OF INTESTINAL MUCUS BARRIER DURING INFLAMMATION

2020 ◽  
Vol 26 (Supplement_1) ◽  
pp. S42-S42
Author(s):  
Kohei Sugihara ◽  
Nobuhiko Kamada
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Gut Microbiota ◽  
Control Diet ◽  
Bacterial Strains ◽  
Germ Free ◽  
Intestinal Mucus ◽  
Gnotobiotic Mice ◽  
Mucus Barrier

Abstract Background Recent accumulating evidence suggests that amino acids have crucial roles in the maintenance of intestinal homeostasis. In inflammatory bowel disease (IBD), amino acid metabolism is changed in both host and the gut microbiota. Among amino acids, L-serine plays a central role in several metabolic processes that are essential for the growth and survival of both mammalian and bacterial cells. However, the role of L-serine in intestinal homeostasis and IBD remains incompletely understood. In this study, we investigated the effect of dietary L-serine on intestinal inflammation in a murine model of colitis. Methods Specific pathogen-free (SPF) mice were fed either a control diet (amino acid-based diet) or an L-serine-deficient diet (SDD). Colitis was induced by the treatment of dextran sodium sulfate (DSS). The gut microbiome was analyzed by 16S rRNA sequencing. We also evaluate the effect of dietary L-serine in germ-free mice and gnotobiotic mice that were colonized by a consortium of non-mucolytic bacterial strains or the consortium plus mucolytic bacterial strains. Results We found that the SDD exacerbated experimental colitis in SPF mice. However, the severity of colitis in SDD-fed mice was comparable to control diet-fed mice in germ-free condition, suggesting that the gut microbiota is required for exacerbation of colitis caused by the restriction of dietary L-serine. The gut microbiome analysis revealed that dietary L-serine restriction fosters the blooms of a mucus-degrading bacterium Akkermansia muciniphila and adherent-invasive Escherichia coli in the inflamed gut. Consistent with the expansion of mucolytic bacteria, SDD-fed mice showed a loss of the intestinal mucus layer. Dysfunction of the mucus barrier resulted in increased intestinal permeability, thereby leading to bacterial translocation to the intestinal mucosa, which subsequently increased the severity of colitis. The increased intestinal permeability and subsequent bacterial translocation were observed in SDD-fed gnotobiotic mice that colonized by mucolytic bacteria. In contrast, dietary L-serine restriction did not alter intestinal barrier integrity in gnotobiotic mice that colonized only by non-mucolytic bacteria. Conclusion Our results suggest that dietary L-serine regulates the integrity of the intestinal mucus barrier during inflammation by limiting the expansion of mucus degrading bacteria.

Induction of Translational Readthrough across the Thalassemia-Causing Premature Stop Codon in β-Globin-Encoding mRNA

Biochemistry ◽  
2019 ◽  
Vol 59 (1) ◽  
pp. 80-84 ◽  
Author(s):  
Debaleena Kar ◽  
Karthi Sellamuthu ◽  
Sangeetha Devi Kumar ◽  
Sandeep M. Eswarappa
Keyword(s):  
Stop Codon ◽  
Premature Stop Codon ◽  
Translational Readthrough

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 Cis-Segregation of c.1171C>T Stop Codon (p.R391*) in SERPINC1 Gene and c.1691G>A Transition (p.R506Q) in F5 Gene and Selected GWAS Multilocus Approach in Inherited Thrombophilia

Genes ◽  
2021 ◽  
Vol 12 (6) ◽  
pp. 934
Author(s):  
Donato Gemmati ◽  
Giovanna Longo ◽  
Eugenia Franchini ◽  
Juliana Araujo Silva ◽  
Ines Gallo ◽  
...  
Keyword(s):  
At Risk ◽  
Stop Codon ◽  
Association Studies ◽  
Premature Stop Codon ◽  
Large Family ◽  
Genome Wide Association Studies ◽  
Loss Of Function ◽  
Gene Markers ◽  
Inherited Thrombophilia ◽  
Fv Leiden

Inherited thrombophilia (e.g., venous thromboembolism, VTE) is due to rare loss-of-function mutations in anticoagulant factors genes (i.e., SERPINC1, PROC, PROS1), common gain-of-function mutations in procoagulant factors genes (i.e., F5, F2), and acquired risk conditions. Genome Wide Association Studies (GWAS) recently recognized several genes associated with VTE though gene defects may unpredictably remain asymptomatic, so calculating the individual genetic predisposition is a challenging task. We investigated a large family with severe, recurrent, early-onset VTE in which two sisters experienced VTE during pregnancies characterized by a perinatal in-utero thrombosis in the newborn and a life-saving pregnancy-interruption because of massive VTE, respectively. A nonsense mutation (CGA > TGA) generating a premature stop-codon (c.1171C>T; p.R391*) in the exon 6 of SERPINC1 gene (1q25.1) causing Antithrombin (AT) deficiency and the common missense mutation (c.1691G>A; p.R506Q) in the exon 10 of F5 gene (1q24.2) (i.e., FV Leiden; rs6025) were coinherited in all the symptomatic members investigated suspecting a cis-segregation further confirmed by STR-linkage-analyses [i.e., SERPINC1 IVS5 (ATT)5–18, F5 IVS2 (AT)6–33 and F5 IVS11 (GT)12–16] and SERPINC1 intragenic variants (i.e., rs5878 and rs677). A multilocus investigation of blood-coagulation balance genes detected the coexistence of FV Leiden (rs6025) in trans with FV HR2-haplotype (p.H1299R; rs1800595) in the aborted fetus, and F11 rs2289252, F12 rs1801020, F13A1 rs5985, and KNG1 rs710446 in the newborn and other members. Common selected gene variants may strongly synergize with less common mutations tuning potential life-threatening conditions when combined with rare severest mutations. Merging classic and newly GWAS-identified gene markers in at risk families is mandatory for VTE risk estimation in the clinical practice, avoiding partial risk score evaluation in unrecognized at risk patients.

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