●AIM:To investigate the molecular diagnosis of a threegeneration Chinese family affected with aniridia,and further to identify clinically a PAX6 missense mutation in members with atypical aniridia.●METHODS:Eleven fa...●AIM:To investigate the molecular diagnosis of a threegeneration Chinese family affected with aniridia,and further to identify clinically a PAX6 missense mutation in members with atypical aniridia.●METHODS:Eleven family members with and without atypical aniridia were recruited.All family members underwent comprehensive ophthalmic examinations.A combination of whole exome sequencing(WES)and direct Sanger sequencing were performed to uncover the causative mutation.●RESULTS:Among the 11 family members,8 were clinically diagnosed with congenital aniridia(atypical aniridia phenotype).A rare heterozygous mutation c.622C>T(p.Arg208Trp)in exon 8 of PAX6 was identified in all affected family members but not in the unaffected members or in healthy control subjects.●CONCLUSION:A rare missense mutation in the PAX6 gene is found in members of a three-generation Chinese family with congenital atypical aniridia.This result contributes to an increase in the phenotypic spectrum caused by PAX6 missense heterozygous variants and provides useful information for the clinical diagnosis of atypical aniridia,which may also contribute to genetic counselling and family planning.展开更多
BACKGROUND Peutz-Jeghers syndrome(PJS)is a rare hereditary neoplastic disorder mainly associated with serine/threonine kinase 11(STK11/LKB1)gene mutations.Preimplantation genetic testing can protect a patient’s offsp...BACKGROUND Peutz-Jeghers syndrome(PJS)is a rare hereditary neoplastic disorder mainly associated with serine/threonine kinase 11(STK11/LKB1)gene mutations.Preimplantation genetic testing can protect a patient’s offspring from mutated genes;however,some variations in this gene have been interpreted as variants of uncertain significance(VUS),which complicate reproductive decision-making in genetic counseling.AIM To identify the pathogenicity of two missense variants and provide clinical guidance.METHODS Whole exome gene sequencing and Sanger sequencing were performed on the peripheral blood of patients with PJS treated at the Reproductive and Genetic Hospital of Citic-Xiangya.Software was employed to predict the protein structure,conservation,and pathogenicity of the two missense variation sites in patients with PJS.Additionally,plasmids were constructed and transfected into HeLa cells to observe cell growth.The differences in signal pathway expression between the variant group and the wild-type group were compared using western blot and immunohistochemistry.Statistical analysis was performed using one-way analysis of variance.P<0.05 was considered statistically significant.RESULTS We identified two missense STK11 gene VUS[c.889A>G(p.Arg297Gly)and c.733C>T(p.Leu245Phe)]in 9 unrelated PJS families who were seeking reproductive assistance.The two missense VUS were located in the catalytic domain of serine/threonine kinase,which is a key structure of the liver kinase B1(LKB1)protein.In vitro experiments showed that the phosphorylation levels of adenosine monophosphate-activated protein kinase(AMPK)at Thr172 and LKB1 at Ser428 were significantly higher in transfected variation-type cells than in wild-type cells.In addition,the two missense STK11 variants promoted the proliferation of HeLa cells.Subsequent immunohistochemical analysis showed that phosphorylated-AMPK(Thr172)expression was significantly lower in gastric,colonic,and uterine polyps from PJS patients with missense variations than in non-PJS patients.Our findings indicate that these two missense STK11 variants are likely pathogenic and inactivate the STK11 gene,causing it to lose its function of regulating downstream phosphorylated-AMPK(Thr172),which may lead to the development of PJS.The identification of the pathogenic mutations in these two clinically characterized PJS patients has been helpful in guiding them toward the most appropriate mode of pregnancy assistance.CONCLUSION These two missense variants can be interpreted as likely pathogenic variants that mediated the onset of PJS in the two patients.These findings not only offer insights for clinical decision-making,but also serve as a foundation for further research and reanalysis of missense VUS in rare diseases.展开更多
AIM: To explore the phenotype and genotype of WeillMarchesani syndrome(WMS) in a Chinese family and review related literature.METHODS: Three WMS patients and other unaffected individuals in this family with a history ...AIM: To explore the phenotype and genotype of WeillMarchesani syndrome(WMS) in a Chinese family and review related literature.METHODS: Three WMS patients and other unaffected individuals in this family with a history of consanguineous marriage were included in this study. Medical history, comprehensive ophthalmic examinations, and systemic evaluation, as well as whole exome and Sanger sequencing of specific genomic regions, were performed. RESULTS: The three affected siblings presented with short stature, brachydactyly and ocular disorders, including very shallow anterior chamber, high myopia, microspherophakia lens subluxation with stretched zonules and glaucoma. Genetic analysis verified a homozygous missense mutation(c.2983C>T: p. Arg995Trp) in ADAMTS17,which was correlated with the diseases in this family, indicating an autosomal recessive inherited manner of WMS. This review aims to summarize the mutation sites of WMS genes, so as to prevent the disease and better guide clinical diagnosis and treatment.CONCLUSION: A novel homozygous missense variant of ADAMTS17 is identified in a WMS family with a history of consanguineous marriage. Our study expands the range of mutations associated with WMS and deepens our understanding of pathology in disease associated with ADAMTS17 variants.展开更多
A novel mutation of vascular endothelial growth factor receptor gene （VEGFR-3）, was identified in a four-generation Chinese family with hereditary lymphedema type I （HL-I）. Genetic linkage analysis was performed o...A novel mutation of vascular endothelial growth factor receptor gene （VEGFR-3）, was identified in a four-generation Chinese family with hereditary lymphedema type I （HL-I）. Genetic linkage analysis was performed on the known genetic locus for HL-I with a panel of polymorphic markers, and then mutations were screened out by direct sequencing. By genotyping, the family showed the linkage to HL-I locus on 5q35.3. Mutation screening analysis of the exons encoding the intracellular kinase domains of VEGFR-3, revealed a novel missense mutation D1055V. This mutation cosegregated with the disease phenotype in the family and was not found in 100 normal controls. This finding has expanded the spectrum of the VEGFR-3 gene mutations causing HL-I, and will be useful for further genetic consultation and genetic diagnosis.展开更多
AIM: To investigate the germline mutations of MSH6 gene in probands of Chinese hereditary non-polyposis colorectal cancer (HNPCC) families fulfilling different clinical criteria. METHODS: Germline mutations of MSH6 ge...AIM: To investigate the germline mutations of MSH6 gene in probands of Chinese hereditary non-polyposis colorectal cancer (HNPCC) families fulfilling different clinical criteria. METHODS: Germline mutations of MSH6 gene were detected by PCR-based DNA sequencing in 39 unrelated HNPCC probands fulfilling different clinical criteria in which MSH2 and MLH1 mutations were excluded. To further investigate the pathological effects of detected missense mutations, we analyzed the above related MSH6 exons using PCR-based sequencing in 137 healthy persons with no family history. The clinicopathological features were collected from the Archive Library of Cancer Hospital, Fudan University and analyzed. RESULTS: Four germline missense mutations distributed in the 4th, 6th and 9th exons were observed. Of them, three were not found in international HNPCC databases and did not occur in 137 healthy controls, indicating that they were novel missense mutations. The remaining mutation which is consistent with the case H14 at c.3488A>T of exon 6 of MSH6 gene was also found in the controls, the rate was approximately 3.65% (5/137) and the type of mutation was not found in the international HNPCC mutational and SNP databases, suggesting that this missense mutation was a new SNP unreported up to date. CONCLUSION: Three novel missense mutations and a new SNP observed in the probands of Chinese HNPCC families, may play an important role in the development of HNPCC.展开更多
AIMTo identify the genetic defects in a Chinese family with achromatopsia.METHODSA 2.5-year-old boy, who displayed nystagmus, photophobia, and hyperopia since early infancy, was clinically evaluated. To further confir...AIMTo identify the genetic defects in a Chinese family with achromatopsia.METHODSA 2.5-year-old boy, who displayed nystagmus, photophobia, and hyperopia since early infancy, was clinically evaluated. To further confirm and localize the causative mutations in this family, targeted region capture and next-generation sequencing of candidate genes, such as CNGA3, CNGB3, GNAT2, PDE6C, and PDE6H were performed using a custom-made capture array.RESULTSSlit-lamp examination showed no specific findings in the anterior segments. The optic discs and maculae were normal on fundoscopy. The unaffected family members reported no ocular complaints. Clinical signs and symptoms were consistent with a clinical impression of autosomal recessive achromatopsia. The results of sequence analysis revealed two novel missense mutations in CNGA3, c.633T>A (p.D211E) and c.1006G>T (p.V336F), with an autosomal recessive mode of inheritance.CONCLUSIONGenetic analysis of a Chinese family confirmed the clinical diagnosis of achromatopsia. Two novel mutations were identified in CNGA3, which extended the mutation spectrum of this disorder.展开更多
Tooth development is a complex process that involves precise and time-dependent orchestration of multiple genetic, molecular,and cellular interactions. Ameloblastin(AMBN, also named "amelin" or "sheathl...Tooth development is a complex process that involves precise and time-dependent orchestration of multiple genetic, molecular,and cellular interactions. Ameloblastin(AMBN, also named "amelin" or "sheathlin") is the second most abundant enamel matrix protein known to have a key role in amelogenesis. Amelogenesis imperfecta(AI [MIM: 104500]) refers to a genetically and phenotypically heterogeneous group of conditions characterized by inherited developmental enamel defects. The hereditary dentin disorders comprise a variety of autosomal-dominant genetic symptoms characterized by abnormal dentin structure affecting either the primary or both the primary and secondary teeth. The vital role of Ambn in amelogenesis has been confirmed experimentally using mouse models. Only two cases have been reported of mutations of AMBN associated with non-syndromic human AI. However, no AMBN missense mutations have been reported to be associated with both human AI and dentin disorders.We recruited one kindred with autosomal-dominant amelogenesis imperfecta(ADAI) and dentinogenesis imperfecta/dysplasia characterized by generalized severe enamel and dentin defects. Whole exome sequencing of the proband identified a novel heterozygous C-T point mutation at nucleotide position 1069 of the AMBN gene, causing a Pro to Ser mutation at the conserved amino acid position 357 of the protein. Exfoliated third molar teeth from the affected family members were found to have enamel and dentin of lower mineral density than control teeth, with thinner and easily fractured enamel, short and thick roots, and pulp obliteration. This study demonstrates, for the first time, that an AMBN missense mutation causes non-syndromic human AI and dentin disorders.展开更多
PIK3R5 is the regulatory subunit of Phosphoinositide 3-kinase γ (PI3Kγ) that is responsible for phosphory-lating membrane lipids to activate the AKT pathway. PIK3R5 binds Gβγ and facilitates the interaction with p...PIK3R5 is the regulatory subunit of Phosphoinositide 3-kinase γ (PI3Kγ) that is responsible for phosphory-lating membrane lipids to activate the AKT pathway. PIK3R5 binds Gβγ and facilitates the interaction with p110γ catalytic subunit (PIK3CG) during PI3Kγ activation. The identification of PIK3R5 P629S mutation in AOA2 patients indicated a potential defect in the AKT pathway resulting from impaired PIK3R5 interaction with Gβγ and PIK3CG, defective AKT pathway can result in cerebellar cell death causing neurological symptoms. Our in silico macromolecular docking of the wild type and mutant PIK3R5 protein models with ligand revealed an energy requirement to maintain the mutant complexes compared to no energy required to maintain the wild type complexes, in addition, the mutant structures were loose compared to rigid wild type structures, such structural changes may impair the molecular function of the PIK3R5 and hence affect the AKT pathway.展开更多
Male infertility is a worldwide health issue,affecting 8%–12%of the global population.Oligoasthenoteratozoospermia(OAT)represents a severe type of male infertility,characterized by reduced sperm count and motility an...Male infertility is a worldwide health issue,affecting 8%–12%of the global population.Oligoasthenoteratozoospermia(OAT)represents a severe type of male infertility,characterized by reduced sperm count and motility and an increased frequency of sperm with aberrant morphology.Using whole-exome sequencing,this study identified a novel missense mutation(c.848C>A,p.A283E)in the coiled-coil domain-containing 34 gene(CCDC34)in a consanguineous Pakistani family.This rare mutation was predicted to be deleterious and to affect the protein stability.Hematoxylin and eosin staining of spermatozoa from the patient with OAT revealed multiple morphological abnormalities of the flagella and transmission electron microscopy indicated axonemal ultrastructural defects with a lack of outer dynein arms.These findings indicated that CCDC34 plays a role in maintaining the axonemal ultrastructure and the assembly or stability of the outer dynein arms,thus expanding the phenotypic spectrum of CCDC34 missense mutations.展开更多
Objective:In this study,we aimed to explore the biological functions of 10 rare case-specific missense mutations in GLI2 and 4 in GLI3,which were previously screened in a cohort of 412 patients with congenital heart d...Objective:In this study,we aimed to explore the biological functions of 10 rare case-specific missense mutations in GLI2 and 4 in GLI3,which were previously screened in a cohort of 412 patients with congenital heart disease(CHD)and 213 normal controls from Shandong Province,China.Methods:A dual-luciferase reporter assay was used to assess the effects of these mutations in GLI2 and GLI3 on the activity of the sonic Hedgehog signaling pathway in HEK293T cells.Differences in protein levels between mutant and wild-type GLI2 and GLI3 were detected in HEK293T cells using Western blotting.Results:The dual-luciferase reporter assay showed that compared to the wild-type GLI2 protein,p.A1113V significantly increased activation of the sonic Hedgehog signaling pathway,whereas p.H78P and p.I1451S did not have a significant effect.The other mutations largely reduced the activation effect.Compared with the wild-type GLI3 protein,only p.A286V,among the four mutations,significantly reduced the activation effect on the SHH signaling pathway.Western blotting data showed reduced expression of GLI2 p.G716V,GLI2 p.K736N,GLI2 p.I1451S,and GLI3 p.A286V,whereas the remaining mutations had no significant effects.Conclusion:The mutations GLI2 c.2147G>T(p.G716V),GLI2 c.2208G>C(p.K736N),and GLI3 c.857C>T(p.A286V)involved in CHD affect the regulation of the sonic Hedgehog signaling pathway;thus,these rare missense mutations in GLI2 and GLI3 might increase the risk of CHD.展开更多
●AIM:To identify disease-causative mutations in families with congenital cataract.●METHODS:Two Chinese families with autosomaldominant congenital cataract(ADCC)were recruited and underwent comprehensive eye examinat...●AIM:To identify disease-causative mutations in families with congenital cataract.●METHODS:Two Chinese families with autosomaldominant congenital cataract(ADCC)were recruited and underwent comprehensive eye examinations.Gene panel next-generation sequencing of common pathogenic genes of congenital cataract was performed in the proband of each family.Sanger sequencing was used to valid the candidate gene mutations and sequence the other family members for co-segregation analysis.The effect of sequence changes on protein structure and function was predicted through bioinformatics analysis.Major intrinsic protein(MIP)-wildtype and MIP-G29R plasmids were constructed and microinjected into zebrafish single-cell stage embryos.Zebrafish embryonic lens phenotypes were screened using confocal microscopy.●RESULTS:A novel heterozygous mutation(c.85G>A;p.G29R)in the MIP gene was identified in the proband of one family.A known heterozygous mutation(c.97C>T;p.R33C;rs864309693)in MIP was found in the proband of another family.In-silico prediction indicated that the novel mutation might affect the MIP protein function.Zebrafish embryonic lens was uniformly transparent in both wild-type PCS2+MIP and mutant PCS2+MIP.●CONCLUSION:Two missense mutations in the MIP gene in Chinese cataract families are identified,and one of which is novel.These findings expand the genetic spectrum of MIP mutations associated with cataracts.The functional studies suggest that the novel MIP mutation might not be a gain-of-function but a loss-of-function mutation.展开更多
AIM:To identify genetic defects in a Chinese family with congenital posterior polar cataracts and assess the pathogenicity.METHODS:A four-generation Chinese family affected with autosomal dominant congenital cataract ...AIM:To identify genetic defects in a Chinese family with congenital posterior polar cataracts and assess the pathogenicity.METHODS:A four-generation Chinese family affected with autosomal dominant congenital cataract was recruited.Nineteen individuals took part in this study including 5 affected and 14 unaffected individuals.Sanger sequencing targeted hot-spot regions of 27 congenital cataract-causing genes for variant discovery.The pathogenicity of the variant was evaluated by the guidelines of American College of Medical Genetics and InterVar software.Confocal microscopy was applied to detect the subcellular localization of fluorescence-labeled ephrin type-A receptor 2(EPHA2).Co-immunoprecipitation assay was implemented to estimate the interaction between EphA2 and other lens membrane proteins.The mRNA and protein expression were analyzed by reverse transcription-polymerase chain reaction(qRT-PCR)and Western blotting assay,respectively.The cell migration was analyzed by wound healing assay.Zebrafish model was generated by ectopic expression of human EPHA2/p.R957P mutant to demonstrate whether the mutant could cause lens opacity in vivo.RESULTS:A novel missense and pathogenic variant c.2870G>C was identified in the sterile alpha motif(SAM)domain of EPHA2.Functional studies demonstrated the variant’s impact:reduced EPHA2 protein expression,altered subcellular localization,and disrupted interactions with other lens membrane proteins.This mutant notably enhanced human lens epithelial cell migration,and induced a central cloudy region and roughness in zebrafish lenses with ectopic expression of human EPHA2/p.R957P mutant under differential interference contrast(DIC)optics.CONCLUSION:Novel pathogenic c.2870G>C variant of EPHA2 in a Chinese congenital cataract family contributes to disease pathogenesis.展开更多
Background:Isolated methylmalonic acidemia is a rare autosomal recessive metabolic disorder mostly caused by mutations in the methylmalonyl coenzyme A mutase (MCM) gene (MUT).This study aimed to verify whether missens...Background:Isolated methylmalonic acidemia is a rare autosomal recessive metabolic disorder mostly caused by mutations in the methylmalonyl coenzyme A mutase (MCM) gene (MUT).This study aimed to verify whether missense mutations in MUT in Chinese patients affect the stability and enzymatic activity of MCM.Methods:Eight Chinese patients were identified with novel mutations.Plasmids carrying the wild-type and mutated MUT cDNA were constructed and transfected into HEK293T cells for functional analyses.The expression and activity of MCM were determined by western blot and ultra-performance liquid chromatography,respectively.Results:All patients had high levels of blood propionylcarnitine and urinary methylmalonyl acid.By the end of the study,two patients were lost to follow-up,three died,and three survived with mental retardation.Compared to the wild-type protein,the expression levels of all missense mutations of in vitro MCM protein were decreased (P<0.05) except those for I597R,and the MCM activity of the mutations was reduced in a permissive assay.Conclusion:The missense mutations L140P,A141T,G161V,W309G,I505T,Q514K,I597R and G723D affected the stability and enzymatic activity of MCM,indicating that they had a disease-causing capacity.展开更多
Background：Charcot-Marie-Tooth disease （CMT） is the most common inherited peripheral neuropathy.A great number of causative genes have been described in CMT,and among them,the heterozygous duplication of peripheral...Background：Charcot-Marie-Tooth disease （CMT） is the most common inherited peripheral neuropathy.A great number of causative genes have been described in CMT,and among them,the heterozygous duplication of peripheral myelin protein-22 （PMP22） is the major cause.Although the missense mutation in PMP22 is rarely reported,it has been demonstrated to be associated with CMT.This study described a novel missense mutation of PMP22 in a Chinese family with CMT phenotype.Methods：Targeted next-generation sequencing （NGS） was used to screen the causative genes in a family featured with an autosomal dominant demyelinating form of CMT.The potential variants identified by targeted NGS were verified by Sanger sequencing and classified according to the American College of Medical Genetics and Genomics standards and guidelines.Further cell transfection studies were performed to characterize the function of the novel variant.Results：Using targeted NGS,a novel heterozygous missense variant in PMP22 （c.320G〉A,p.G107D） was identified.In vitro cell functional studies revealed that mutant PMP22 protein carrying p.G 107D mutation lost the ability to reach the plasma membrane,was mainly retained in the endoplasmic reticulum,and induced cell apoptosis.Conclusions：This study supported the notion that missense mutations in PMP22 give rise to a CMT phenotype,possibly through a toxic gain-of-function mechanism.展开更多
To the Editor:Adams-Oliver syndrome (AOS,including 6 types) was initially reported in a three-generation family by Adams and Oliver in 1945,[1] with an estimated incidence of 1 in 225,000 live births.[2] Approximately...To the Editor:Adams-Oliver syndrome (AOS,including 6 types) was initially reported in a three-generation family by Adams and Oliver in 1945,[1] with an estimated incidence of 1 in 225,000 live births.[2] Approximately 84% of AOS patients have terminal transverse limb defects,including amputations,syndactyly,brachydactyly,or oligodactyly.展开更多
The Psychiatric Genomics Consortium(PGC)has recently identified 10 potential functional coding variants for schizophrenia.However,how these coding variants confer schizophrenia risk remains largely unknown.Here,we inv...The Psychiatric Genomics Consortium(PGC)has recently identified 10 potential functional coding variants for schizophrenia.However,how these coding variants confer schizophrenia risk remains largely unknown.Here,we investigate the associations between eight potential functional coding variants identified by PGC and schizophrenia in a large Han Chinese sample(n=4022 cases and 9270 controls).Among the eight tested single nucelotide polymorphisms(SNPs),rs3617(a missense variant,p.K315 Q in the ITIH3 gene)showed genome-wide significant association with schizophrenia in the Han Chinese population(P=8.36×10-16),with the same risk allele as in PGC.Interestingly,rs3617 is located in a genomic region that is highly evolutionarily conserved,and its schizophrenia risk allele(C allele)was associated with lower ITIH3 mRNA and protein expression.Intriguingly,mouse neural stem cells stably overexpressing ITIH3 with different alleles of rs3617 exhibited significant differences in proliferation,migration,and differentiation,suggesting the impact of rs3617 on neurodevelopment.Subsequent transcriptome analysis found that the differentially expressed genes in neural stem cells stably overexpressing different alleles of rs3617 were significantly enriched in schizophrenia-related pathways,including cell adhesion,synapse assembly,MAPK and PI3 K-AKT pathways.Our study provides convergent lines of evidence suggesting that rs3617 in ITIH3 likely affects protein function and neurodevelopment and thereby confers risk of schizophrenia.展开更多
Excess de novo likely gene-disruptive and missense variants within dozens of genes have been identified in autism spectrum disorder(ASD)and other neurodevelopmental disorders.However,many rare inherited missense varia...Excess de novo likely gene-disruptive and missense variants within dozens of genes have been identified in autism spectrum disorder(ASD)and other neurodevelopmental disorders.However,many rare inherited missense variants of these high-risk genes have not been thoroughly evaluated.In this study,we analyzed the rare missense variant burden of POGZ in a large cohort of ASD patients from the Autism Clinical and Genetic Resources in China(ACGC)and further dissected the functional effect of diseaseassociated missense variants on neuronal development.Our results showed a significant burden of rare missense variants in ASD patients compared to the control population(P=4.6×10-5,OR=3.96),and missense variants in ASD patients showed more severe predicted functional outcomes than those in controls.Furthermore,by leveraging published large-scale sequencing data of neurodevelopmental disorders(NDDs)and sporadic case reports,we identified 8 de novo missense variants of POGZ in NDD patients.Functional analysis revealed that two inherited,but not de novo,missense variants influenced the cellular localization of POGZ and failed to rescue the defects in neurite and dendritic spine development caused by Pogz knockdown in cultured mouse primary cortical neurons.Significantly,L1CAM,an autism candidate risk gene,is differentially expressed in POGZ deficient cell lines.Reduced expression of L1cam was able to partially rescue the neurite length defects caused by Pogz knockdown.Our study showed the important roles of rare inherited missense variants of POGZ in ASD risk and neuronal development and identified the potential downstream targets of POGZ,which are important for further molecular mechanism studies.展开更多
In this work, the most detrimental missense mutations of aspartoacylase that cause Canavan's disease were identified computationally and the substrate binding efficiencies of those missense mutations were analyzed...In this work, the most detrimental missense mutations of aspartoacylase that cause Canavan's disease were identified computationally and the substrate binding efficiencies of those missense mutations were analyzed. Out of 30 missense mutations, I-Mutant 2.0, SIFT and PolyPhen programs identified 22 variants that were less stable, deleterious and damaging respectively. Subsequently, modeling of these 22 variants was performed to understand the change in their conformations with respect to the native aspartoacylase by computing their root mean squared deviation (RMSD). Furthermore, the native protein and the 22 mutants were docked with the substrate NAA (N-Acetyl-Aspartic acid) to explain the substrate binding efficiencies of those detrimental missense mutations. Among the 22 mutants, the docking studies identified that 15 mutants caused lower binding affinity for NAA than the native protein. Finally, normal mode analysis determined that the loss of binding affinity of these 15 mutants was caused by altered flexibility in the amino acids that bind to NAA compared with the native protein. Thus, the pre- sent study showed that the majority of the substrate-binding amino acids in those 15 mutants displayed loss of flexibility, which could be the theoretical explanation of decreased binding affinity between the mutant aspartoacylases and NAA.展开更多
Background： Gaucher＇s disease （GD） is an autosomal recessive disorder caused by a deficiency of acid β-glucosidase （glucocerebrosidase [GBA]） that results in the accumulation of glucocerebroside within macropha...Background： Gaucher＇s disease （GD） is an autosomal recessive disorder caused by a deficiency of acid β-glucosidase （glucocerebrosidase [GBA]） that results in the accumulation of glucocerebroside within macrophages. Many mutations have been reported to be associated with this disorder. This study aimed to discover more mutations and provide data for the genetic pattern of the gene, which will help the development of quick and accurate genetic diagnostic tools for this disease. Methods： Genomic DNA was obtained from peripheral blood leukocytes of the patient and Sanger sequencing is used to sequence GBA gene. Sequence alignments of mammalian β-GBA （GCase） and three-dimensional protein structure prediction of the mutation were made. A construct of this mutant and its compound heterozygous counterpart were used to measure GCase in vitro. Results： GCase is relatively conserved at p.T219A. This novel mutation differs from its wild-type in structure. Moreover, it also causes a reduction in GCase enzyme activity. Conclusion： This novel mutation （c.655A〉G, p.T219A） is a pathogenic missense mutation, which contributes to GD.展开更多
In this work, the most detrimental missense mutations of Madl protein that cause various types of cancer were identified computationally and the substrate binding efficiencies of those missense mutations were analyzed...In this work, the most detrimental missense mutations of Madl protein that cause various types of cancer were identified computationally and the substrate binding efficiencies of those missense mutations were analyzed. Out of 13 missense mutations, I Mutant 2.0, SIFT and PolyPhen programs identified 3 variants that were less stable, deleterious and damaging respectively. Subsequently, modeling of these 3 variants was performed to understand the change in their conformations with respect to the native Madl by computing their root mean squared deviation （RMSD）. Furthermore, the native protein and the 3 mutants were docked with the binding partner Mad2 to explain the substrate binding efficiencies of those detrimental missense mutations. The docking studies identified that all the 3 mutants caused lower binding affinity for Mad2 than the native protein. Finally, normal mode analysis determined that the loss of binding affinity of these 3 mutants was caused by altered flexibility in the amino acids that bind to Mad2 compared with the native protein. Thus, the present study showed that majority of the substrate binding amino acids in those 3 mutants displayed loss of flexibility, which could be the theoretical explanation of decreased binding affinity between the mutant Madl and Mad2.展开更多
文摘●AIM:To investigate the molecular diagnosis of a threegeneration Chinese family affected with aniridia,and further to identify clinically a PAX6 missense mutation in members with atypical aniridia.●METHODS:Eleven family members with and without atypical aniridia were recruited.All family members underwent comprehensive ophthalmic examinations.A combination of whole exome sequencing(WES)and direct Sanger sequencing were performed to uncover the causative mutation.●RESULTS:Among the 11 family members,8 were clinically diagnosed with congenital aniridia(atypical aniridia phenotype).A rare heterozygous mutation c.622C>T(p.Arg208Trp)in exon 8 of PAX6 was identified in all affected family members but not in the unaffected members or in healthy control subjects.●CONCLUSION:A rare missense mutation in the PAX6 gene is found in members of a three-generation Chinese family with congenital atypical aniridia.This result contributes to an increase in the phenotypic spectrum caused by PAX6 missense heterozygous variants and provides useful information for the clinical diagnosis of atypical aniridia,which may also contribute to genetic counselling and family planning.
基金Supported by the Natural Science Foundation of Hunan Province,China,No.2023JJ30422.
文摘BACKGROUND Peutz-Jeghers syndrome(PJS)is a rare hereditary neoplastic disorder mainly associated with serine/threonine kinase 11(STK11/LKB1)gene mutations.Preimplantation genetic testing can protect a patient’s offspring from mutated genes;however,some variations in this gene have been interpreted as variants of uncertain significance(VUS),which complicate reproductive decision-making in genetic counseling.AIM To identify the pathogenicity of two missense variants and provide clinical guidance.METHODS Whole exome gene sequencing and Sanger sequencing were performed on the peripheral blood of patients with PJS treated at the Reproductive and Genetic Hospital of Citic-Xiangya.Software was employed to predict the protein structure,conservation,and pathogenicity of the two missense variation sites in patients with PJS.Additionally,plasmids were constructed and transfected into HeLa cells to observe cell growth.The differences in signal pathway expression between the variant group and the wild-type group were compared using western blot and immunohistochemistry.Statistical analysis was performed using one-way analysis of variance.P<0.05 was considered statistically significant.RESULTS We identified two missense STK11 gene VUS[c.889A>G(p.Arg297Gly)and c.733C>T(p.Leu245Phe)]in 9 unrelated PJS families who were seeking reproductive assistance.The two missense VUS were located in the catalytic domain of serine/threonine kinase,which is a key structure of the liver kinase B1(LKB1)protein.In vitro experiments showed that the phosphorylation levels of adenosine monophosphate-activated protein kinase(AMPK)at Thr172 and LKB1 at Ser428 were significantly higher in transfected variation-type cells than in wild-type cells.In addition,the two missense STK11 variants promoted the proliferation of HeLa cells.Subsequent immunohistochemical analysis showed that phosphorylated-AMPK(Thr172)expression was significantly lower in gastric,colonic,and uterine polyps from PJS patients with missense variations than in non-PJS patients.Our findings indicate that these two missense STK11 variants are likely pathogenic and inactivate the STK11 gene,causing it to lose its function of regulating downstream phosphorylated-AMPK(Thr172),which may lead to the development of PJS.The identification of the pathogenic mutations in these two clinically characterized PJS patients has been helpful in guiding them toward the most appropriate mode of pregnancy assistance.CONCLUSION These two missense variants can be interpreted as likely pathogenic variants that mediated the onset of PJS in the two patients.These findings not only offer insights for clinical decision-making,but also serve as a foundation for further research and reanalysis of missense VUS in rare diseases.
基金Supported by The Cadre Health Research Program of the Sichuan Province (No.2023-119)Sichuan Science and Technology Program (No.2021YFS0213)。
文摘AIM: To explore the phenotype and genotype of WeillMarchesani syndrome(WMS) in a Chinese family and review related literature.METHODS: Three WMS patients and other unaffected individuals in this family with a history of consanguineous marriage were included in this study. Medical history, comprehensive ophthalmic examinations, and systemic evaluation, as well as whole exome and Sanger sequencing of specific genomic regions, were performed. RESULTS: The three affected siblings presented with short stature, brachydactyly and ocular disorders, including very shallow anterior chamber, high myopia, microspherophakia lens subluxation with stretched zonules and glaucoma. Genetic analysis verified a homozygous missense mutation(c.2983C>T: p. Arg995Trp) in ADAMTS17,which was correlated with the diseases in this family, indicating an autosomal recessive inherited manner of WMS. This review aims to summarize the mutation sites of WMS genes, so as to prevent the disease and better guide clinical diagnosis and treatment.CONCLUSION: A novel homozygous missense variant of ADAMTS17 is identified in a WMS family with a history of consanguineous marriage. Our study expands the range of mutations associated with WMS and deepens our understanding of pathology in disease associated with ADAMTS17 variants.
文摘A novel mutation of vascular endothelial growth factor receptor gene （VEGFR-3）, was identified in a four-generation Chinese family with hereditary lymphedema type I （HL-I）. Genetic linkage analysis was performed on the known genetic locus for HL-I with a panel of polymorphic markers, and then mutations were screened out by direct sequencing. By genotyping, the family showed the linkage to HL-I locus on 5q35.3. Mutation screening analysis of the exons encoding the intracellular kinase domains of VEGFR-3, revealed a novel missense mutation D1055V. This mutation cosegregated with the disease phenotype in the family and was not found in 100 normal controls. This finding has expanded the spectrum of the VEGFR-3 gene mutations causing HL-I, and will be useful for further genetic consultation and genetic diagnosis.
基金Supported by Shanghai Medical Development Fund for Major Projects, No. 05Ⅲ004 and Shanghai Pu Jiang Projects for Talented-Men, 06PJ14019
文摘AIM: To investigate the germline mutations of MSH6 gene in probands of Chinese hereditary non-polyposis colorectal cancer (HNPCC) families fulfilling different clinical criteria. METHODS: Germline mutations of MSH6 gene were detected by PCR-based DNA sequencing in 39 unrelated HNPCC probands fulfilling different clinical criteria in which MSH2 and MLH1 mutations were excluded. To further investigate the pathological effects of detected missense mutations, we analyzed the above related MSH6 exons using PCR-based sequencing in 137 healthy persons with no family history. The clinicopathological features were collected from the Archive Library of Cancer Hospital, Fudan University and analyzed. RESULTS: Four germline missense mutations distributed in the 4th, 6th and 9th exons were observed. Of them, three were not found in international HNPCC databases and did not occur in 137 healthy controls, indicating that they were novel missense mutations. The remaining mutation which is consistent with the case H14 at c.3488A>T of exon 6 of MSH6 gene was also found in the controls, the rate was approximately 3.65% (5/137) and the type of mutation was not found in the international HNPCC mutational and SNP databases, suggesting that this missense mutation was a new SNP unreported up to date. CONCLUSION: Three novel missense mutations and a new SNP observed in the probands of Chinese HNPCC families, may play an important role in the development of HNPCC.
基金Supported by the National Natural Science Foundation of China(No.81371005No.31100991)
文摘AIMTo identify the genetic defects in a Chinese family with achromatopsia.METHODSA 2.5-year-old boy, who displayed nystagmus, photophobia, and hyperopia since early infancy, was clinically evaluated. To further confirm and localize the causative mutations in this family, targeted region capture and next-generation sequencing of candidate genes, such as CNGA3, CNGB3, GNAT2, PDE6C, and PDE6H were performed using a custom-made capture array.RESULTSSlit-lamp examination showed no specific findings in the anterior segments. The optic discs and maculae were normal on fundoscopy. The unaffected family members reported no ocular complaints. Clinical signs and symptoms were consistent with a clinical impression of autosomal recessive achromatopsia. The results of sequence analysis revealed two novel missense mutations in CNGA3, c.633T>A (p.D211E) and c.1006G>T (p.V336F), with an autosomal recessive mode of inheritance.CONCLUSIONGenetic analysis of a Chinese family confirmed the clinical diagnosis of achromatopsia. Two novel mutations were identified in CNGA3, which extended the mutation spectrum of this disorder.
基金partially supported by a grant from the National Natural Science Foundation of China 31371279 (to Fu Xiong)the National Natural Science Foundation of China 81371137 (to Bu-Ling Wu)the Science and Technology Program of Guangzhou 201707010301 (to Fu Xiong)
文摘Tooth development is a complex process that involves precise and time-dependent orchestration of multiple genetic, molecular,and cellular interactions. Ameloblastin(AMBN, also named "amelin" or "sheathlin") is the second most abundant enamel matrix protein known to have a key role in amelogenesis. Amelogenesis imperfecta(AI [MIM: 104500]) refers to a genetically and phenotypically heterogeneous group of conditions characterized by inherited developmental enamel defects. The hereditary dentin disorders comprise a variety of autosomal-dominant genetic symptoms characterized by abnormal dentin structure affecting either the primary or both the primary and secondary teeth. The vital role of Ambn in amelogenesis has been confirmed experimentally using mouse models. Only two cases have been reported of mutations of AMBN associated with non-syndromic human AI. However, no AMBN missense mutations have been reported to be associated with both human AI and dentin disorders.We recruited one kindred with autosomal-dominant amelogenesis imperfecta(ADAI) and dentinogenesis imperfecta/dysplasia characterized by generalized severe enamel and dentin defects. Whole exome sequencing of the proband identified a novel heterozygous C-T point mutation at nucleotide position 1069 of the AMBN gene, causing a Pro to Ser mutation at the conserved amino acid position 357 of the protein. Exfoliated third molar teeth from the affected family members were found to have enamel and dentin of lower mineral density than control teeth, with thinner and easily fractured enamel, short and thick roots, and pulp obliteration. This study demonstrates, for the first time, that an AMBN missense mutation causes non-syndromic human AI and dentin disorders.
文摘PIK3R5 is the regulatory subunit of Phosphoinositide 3-kinase γ (PI3Kγ) that is responsible for phosphory-lating membrane lipids to activate the AKT pathway. PIK3R5 binds Gβγ and facilitates the interaction with p110γ catalytic subunit (PIK3CG) during PI3Kγ activation. The identification of PIK3R5 P629S mutation in AOA2 patients indicated a potential defect in the AKT pathway resulting from impaired PIK3R5 interaction with Gβγ and PIK3CG, defective AKT pathway can result in cerebellar cell death causing neurological symptoms. Our in silico macromolecular docking of the wild type and mutant PIK3R5 protein models with ligand revealed an energy requirement to maintain the mutant complexes compared to no energy required to maintain the wild type complexes, in addition, the mutant structures were loose compared to rigid wild type structures, such structural changes may impair the molecular function of the PIK3R5 and hence affect the AKT pathway.
基金supported by the National Natural Science Foundation of China(No.82071709,No.32070850,and No.82171601)the National Key Research and Developmental Program of China(2022YFC2702601 and 2022YFA0806303)the Joint Fund for New Medicine of USTC(YD9100002034).
文摘Male infertility is a worldwide health issue,affecting 8%–12%of the global population.Oligoasthenoteratozoospermia(OAT)represents a severe type of male infertility,characterized by reduced sperm count and motility and an increased frequency of sperm with aberrant morphology.Using whole-exome sequencing,this study identified a novel missense mutation(c.848C>A,p.A283E)in the coiled-coil domain-containing 34 gene(CCDC34)in a consanguineous Pakistani family.This rare mutation was predicted to be deleterious and to affect the protein stability.Hematoxylin and eosin staining of spermatozoa from the patient with OAT revealed multiple morphological abnormalities of the flagella and transmission electron microscopy indicated axonemal ultrastructural defects with a lack of outer dynein arms.These findings indicated that CCDC34 plays a role in maintaining the axonemal ultrastructure and the assembly or stability of the outer dynein arms,thus expanding the phenotypic spectrum of CCDC34 missense mutations.
基金National Key R&D Program of China(2021YFC2701101)National Natural Science Foundation of China(81930036,82150008,and 81601298)
文摘Objective:In this study,we aimed to explore the biological functions of 10 rare case-specific missense mutations in GLI2 and 4 in GLI3,which were previously screened in a cohort of 412 patients with congenital heart disease(CHD)and 213 normal controls from Shandong Province,China.Methods:A dual-luciferase reporter assay was used to assess the effects of these mutations in GLI2 and GLI3 on the activity of the sonic Hedgehog signaling pathway in HEK293T cells.Differences in protein levels between mutant and wild-type GLI2 and GLI3 were detected in HEK293T cells using Western blotting.Results:The dual-luciferase reporter assay showed that compared to the wild-type GLI2 protein,p.A1113V significantly increased activation of the sonic Hedgehog signaling pathway,whereas p.H78P and p.I1451S did not have a significant effect.The other mutations largely reduced the activation effect.Compared with the wild-type GLI3 protein,only p.A286V,among the four mutations,significantly reduced the activation effect on the SHH signaling pathway.Western blotting data showed reduced expression of GLI2 p.G716V,GLI2 p.K736N,GLI2 p.I1451S,and GLI3 p.A286V,whereas the remaining mutations had no significant effects.Conclusion:The mutations GLI2 c.2147G>T(p.G716V),GLI2 c.2208G>C(p.K736N),and GLI3 c.857C>T(p.A286V)involved in CHD affect the regulation of the sonic Hedgehog signaling pathway;thus,these rare missense mutations in GLI2 and GLI3 might increase the risk of CHD.
基金Supported by the Science,Technology and Innovation Commission of Shenzhen Municipality(No.GJHZ20220913142618036,No.JCYJ20210324113610029).
文摘●AIM:To identify disease-causative mutations in families with congenital cataract.●METHODS:Two Chinese families with autosomaldominant congenital cataract(ADCC)were recruited and underwent comprehensive eye examinations.Gene panel next-generation sequencing of common pathogenic genes of congenital cataract was performed in the proband of each family.Sanger sequencing was used to valid the candidate gene mutations and sequence the other family members for co-segregation analysis.The effect of sequence changes on protein structure and function was predicted through bioinformatics analysis.Major intrinsic protein(MIP)-wildtype and MIP-G29R plasmids were constructed and microinjected into zebrafish single-cell stage embryos.Zebrafish embryonic lens phenotypes were screened using confocal microscopy.●RESULTS:A novel heterozygous mutation(c.85G>A;p.G29R)in the MIP gene was identified in the proband of one family.A known heterozygous mutation(c.97C>T;p.R33C;rs864309693)in MIP was found in the proband of another family.In-silico prediction indicated that the novel mutation might affect the MIP protein function.Zebrafish embryonic lens was uniformly transparent in both wild-type PCS2+MIP and mutant PCS2+MIP.●CONCLUSION:Two missense mutations in the MIP gene in Chinese cataract families are identified,and one of which is novel.These findings expand the genetic spectrum of MIP mutations associated with cataracts.The functional studies suggest that the novel MIP mutation might not be a gain-of-function but a loss-of-function mutation.
基金Supported by the Natural Science Foundation of Fujian Province(No.2021J01229)National Key Research and Development Program of China(No.2016YFC1000307).
文摘AIM:To identify genetic defects in a Chinese family with congenital posterior polar cataracts and assess the pathogenicity.METHODS:A four-generation Chinese family affected with autosomal dominant congenital cataract was recruited.Nineteen individuals took part in this study including 5 affected and 14 unaffected individuals.Sanger sequencing targeted hot-spot regions of 27 congenital cataract-causing genes for variant discovery.The pathogenicity of the variant was evaluated by the guidelines of American College of Medical Genetics and InterVar software.Confocal microscopy was applied to detect the subcellular localization of fluorescence-labeled ephrin type-A receptor 2(EPHA2).Co-immunoprecipitation assay was implemented to estimate the interaction between EphA2 and other lens membrane proteins.The mRNA and protein expression were analyzed by reverse transcription-polymerase chain reaction(qRT-PCR)and Western blotting assay,respectively.The cell migration was analyzed by wound healing assay.Zebrafish model was generated by ectopic expression of human EPHA2/p.R957P mutant to demonstrate whether the mutant could cause lens opacity in vivo.RESULTS:A novel missense and pathogenic variant c.2870G>C was identified in the sterile alpha motif(SAM)domain of EPHA2.Functional studies demonstrated the variant’s impact:reduced EPHA2 protein expression,altered subcellular localization,and disrupted interactions with other lens membrane proteins.This mutant notably enhanced human lens epithelial cell migration,and induced a central cloudy region and roughness in zebrafish lenses with ectopic expression of human EPHA2/p.R957P mutant under differential interference contrast(DIC)optics.CONCLUSION:Novel pathogenic c.2870G>C variant of EPHA2 in a Chinese congenital cataract family contributes to disease pathogenesis.
文摘Background:Isolated methylmalonic acidemia is a rare autosomal recessive metabolic disorder mostly caused by mutations in the methylmalonyl coenzyme A mutase (MCM) gene (MUT).This study aimed to verify whether missense mutations in MUT in Chinese patients affect the stability and enzymatic activity of MCM.Methods:Eight Chinese patients were identified with novel mutations.Plasmids carrying the wild-type and mutated MUT cDNA were constructed and transfected into HEK293T cells for functional analyses.The expression and activity of MCM were determined by western blot and ultra-performance liquid chromatography,respectively.Results:All patients had high levels of blood propionylcarnitine and urinary methylmalonyl acid.By the end of the study,two patients were lost to follow-up,three died,and three survived with mental retardation.Compared to the wild-type protein,the expression levels of all missense mutations of in vitro MCM protein were decreased (P<0.05) except those for I597R,and the MCM activity of the mutations was reduced in a permissive assay.Conclusion:The missense mutations L140P,A141T,G161V,W309G,I505T,Q514K,I597R and G723D affected the stability and enzymatic activity of MCM,indicating that they had a disease-causing capacity.
基金This study was supported by the grants from the National Natural Science Foundation of China （No. 81125009）, and the research foundation for distinguished scholar of Zhejiang University （No. 188020-193810101/089）.
文摘Background：Charcot-Marie-Tooth disease （CMT） is the most common inherited peripheral neuropathy.A great number of causative genes have been described in CMT,and among them,the heterozygous duplication of peripheral myelin protein-22 （PMP22） is the major cause.Although the missense mutation in PMP22 is rarely reported,it has been demonstrated to be associated with CMT.This study described a novel missense mutation of PMP22 in a Chinese family with CMT phenotype.Methods：Targeted next-generation sequencing （NGS） was used to screen the causative genes in a family featured with an autosomal dominant demyelinating form of CMT.The potential variants identified by targeted NGS were verified by Sanger sequencing and classified according to the American College of Medical Genetics and Genomics standards and guidelines.Further cell transfection studies were performed to characterize the function of the novel variant.Results：Using targeted NGS,a novel heterozygous missense variant in PMP22 （c.320G〉A,p.G107D） was identified.In vitro cell functional studies revealed that mutant PMP22 protein carrying p.G 107D mutation lost the ability to reach the plasma membrane,was mainly retained in the endoplasmic reticulum,and induced cell apoptosis.Conclusions：This study supported the notion that missense mutations in PMP22 give rise to a CMT phenotype,possibly through a toxic gain-of-function mechanism.
文摘To the Editor:Adams-Oliver syndrome (AOS,including 6 types) was initially reported in a three-generation family by Adams and Oliver in 1945,[1] with an estimated incidence of 1 in 225,000 live births.[2] Approximately 84% of AOS patients have terminal transverse limb defects,including amputations,syndactyly,brachydactyly,or oligodactyly.
基金equally supported by the National Natural Science Foundation of China of China(31970561 and 31722029 to X.J.L.)the National Key Research and Development Program of China(Stem Cell and Translational Research)(2016YFA0100900)+1 种基金the Innovative Research Team of Science and Technology department of Yunnan Province(2019HC004)the Key Research Project of Yunnan Province(2017FA008 to X.-J.L.)。
文摘The Psychiatric Genomics Consortium(PGC)has recently identified 10 potential functional coding variants for schizophrenia.However,how these coding variants confer schizophrenia risk remains largely unknown.Here,we investigate the associations between eight potential functional coding variants identified by PGC and schizophrenia in a large Han Chinese sample(n=4022 cases and 9270 controls).Among the eight tested single nucelotide polymorphisms(SNPs),rs3617(a missense variant,p.K315 Q in the ITIH3 gene)showed genome-wide significant association with schizophrenia in the Han Chinese population(P=8.36×10-16),with the same risk allele as in PGC.Interestingly,rs3617 is located in a genomic region that is highly evolutionarily conserved,and its schizophrenia risk allele(C allele)was associated with lower ITIH3 mRNA and protein expression.Intriguingly,mouse neural stem cells stably overexpressing ITIH3 with different alleles of rs3617 exhibited significant differences in proliferation,migration,and differentiation,suggesting the impact of rs3617 on neurodevelopment.Subsequent transcriptome analysis found that the differentially expressed genes in neural stem cells stably overexpressing different alleles of rs3617 were significantly enriched in schizophrenia-related pathways,including cell adhesion,synapse assembly,MAPK and PI3 K-AKT pathways.Our study provides convergent lines of evidence suggesting that rs3617 in ITIH3 likely affects protein function and neurodevelopment and thereby confers risk of schizophrenia.
基金supported by the National Natural Science Foundation of China (31671114) to H.G.the National Natural Science Foundation of China (81330027, 81525007, 81730036) to K.X.+5 种基金the National Natural Science Foundation of China (31500832) to J.Q.the National Natural Science Foundation of China (81671122) to Z.H.the National Natural Science Foundation of China (81501182) to Y.P.. H.G.the Natural Science Foundation of Hunan Province (2016RS2001, 2016JC2055) to K.X.supported by the Young Talent Lifts Project of the Chinese Association for Science and Technology (CAST)the Innovation-Driven Project of Central South University (2016CX038)
文摘Excess de novo likely gene-disruptive and missense variants within dozens of genes have been identified in autism spectrum disorder(ASD)and other neurodevelopmental disorders.However,many rare inherited missense variants of these high-risk genes have not been thoroughly evaluated.In this study,we analyzed the rare missense variant burden of POGZ in a large cohort of ASD patients from the Autism Clinical and Genetic Resources in China(ACGC)and further dissected the functional effect of diseaseassociated missense variants on neuronal development.Our results showed a significant burden of rare missense variants in ASD patients compared to the control population(P=4.6×10-5,OR=3.96),and missense variants in ASD patients showed more severe predicted functional outcomes than those in controls.Furthermore,by leveraging published large-scale sequencing data of neurodevelopmental disorders(NDDs)and sporadic case reports,we identified 8 de novo missense variants of POGZ in NDD patients.Functional analysis revealed that two inherited,but not de novo,missense variants influenced the cellular localization of POGZ and failed to rescue the defects in neurite and dendritic spine development caused by Pogz knockdown in cultured mouse primary cortical neurons.Significantly,L1CAM,an autism candidate risk gene,is differentially expressed in POGZ deficient cell lines.Reduced expression of L1cam was able to partially rescue the neurite length defects caused by Pogz knockdown.Our study showed the important roles of rare inherited missense variants of POGZ in ASD risk and neuronal development and identified the potential downstream targets of POGZ,which are important for further molecular mechanism studies.
文摘In this work, the most detrimental missense mutations of aspartoacylase that cause Canavan's disease were identified computationally and the substrate binding efficiencies of those missense mutations were analyzed. Out of 30 missense mutations, I-Mutant 2.0, SIFT and PolyPhen programs identified 22 variants that were less stable, deleterious and damaging respectively. Subsequently, modeling of these 22 variants was performed to understand the change in their conformations with respect to the native aspartoacylase by computing their root mean squared deviation (RMSD). Furthermore, the native protein and the 22 mutants were docked with the substrate NAA (N-Acetyl-Aspartic acid) to explain the substrate binding efficiencies of those detrimental missense mutations. Among the 22 mutants, the docking studies identified that 15 mutants caused lower binding affinity for NAA than the native protein. Finally, normal mode analysis determined that the loss of binding affinity of these 15 mutants was caused by altered flexibility in the amino acids that bind to NAA compared with the native protein. Thus, the pre- sent study showed that the majority of the substrate-binding amino acids in those 15 mutants displayed loss of flexibility, which could be the theoretical explanation of decreased binding affinity between the mutant aspartoacylases and NAA.
基金This study was supported by grants from Natural Science Foundation of China （No. 81371269） and Shanghai Research Program （No. 14140902600, No. 2013ZYJB0015, and No. 14DJ 1400103）.
文摘Background： Gaucher＇s disease （GD） is an autosomal recessive disorder caused by a deficiency of acid β-glucosidase （glucocerebrosidase [GBA]） that results in the accumulation of glucocerebroside within macrophages. Many mutations have been reported to be associated with this disorder. This study aimed to discover more mutations and provide data for the genetic pattern of the gene, which will help the development of quick and accurate genetic diagnostic tools for this disease. Methods： Genomic DNA was obtained from peripheral blood leukocytes of the patient and Sanger sequencing is used to sequence GBA gene. Sequence alignments of mammalian β-GBA （GCase） and three-dimensional protein structure prediction of the mutation were made. A construct of this mutant and its compound heterozygous counterpart were used to measure GCase in vitro. Results： GCase is relatively conserved at p.T219A. This novel mutation differs from its wild-type in structure. Moreover, it also causes a reduction in GCase enzyme activity. Conclusion： This novel mutation （c.655A〉G, p.T219A） is a pathogenic missense mutation, which contributes to GD.
文摘In this work, the most detrimental missense mutations of Madl protein that cause various types of cancer were identified computationally and the substrate binding efficiencies of those missense mutations were analyzed. Out of 13 missense mutations, I Mutant 2.0, SIFT and PolyPhen programs identified 3 variants that were less stable, deleterious and damaging respectively. Subsequently, modeling of these 3 variants was performed to understand the change in their conformations with respect to the native Madl by computing their root mean squared deviation （RMSD）. Furthermore, the native protein and the 3 mutants were docked with the binding partner Mad2 to explain the substrate binding efficiencies of those detrimental missense mutations. The docking studies identified that all the 3 mutants caused lower binding affinity for Mad2 than the native protein. Finally, normal mode analysis determined that the loss of binding affinity of these 3 mutants was caused by altered flexibility in the amino acids that bind to Mad2 compared with the native protein. Thus, the present study showed that majority of the substrate binding amino acids in those 3 mutants displayed loss of flexibility, which could be the theoretical explanation of decreased binding affinity between the mutant Madl and Mad2.
