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3.
Nat Genet ; 51(10): 1438-1441, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31570889

RESUMO

Hypopigmentation along Blaschko's lines is a hallmark of a poorly defined group of mosaic syndromes whose genetic causes are unknown. Here we show that postzygotic inactivating mutations of RHOA cause a neuroectodermal syndrome combining linear hypopigmentation, alopecia, apparently asymptomatic leukoencephalopathy, and facial, ocular, dental and acral anomalies. Our findings pave the way toward elucidating the etiology of pigmentary mosaicism and highlight the role of RHOA in human development and disease.

4.
Oncogene ; 38(48): 7357-7365, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31417180

RESUMO

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematological malignancy characterized by an accumulation of immature T cells. Although patient outcomes have improved, novel targeted therapies are needed to reduce the intensity of chemotherapy and improve the prognosis of high-risk patients. Interleukin-7 (IL-7) modulates the survival and proliferation of normal and malignant T cells. Targeting the IL-7 signaling pathway is thus a potentially effective therapeutic strategy. To achieve such aim, it is essential to first understand how the IL-7 signaling pathway is activated. Although IL-7 production has been observed from multiple stromal tissues, T-ALL autocrine IL-7 secretion has not yet been described. Interestingly, using T-ALL cell lines, primary and patient-derived xenotransplanted (PDX) T-ALL cells, we demonstrate that T-ALL cells produce IL-7 whereas normal T cells do not. Finally, using knock down of IL7 gene in T-ALL cells, we describe to what extent IL-7 autocrine secretion is involved in the T-ALL cells propagation in bone marrow and how it affects the number of leukemia-initiating cells in PDX mice. Together, these results demonstrate how the autocrine production of the IL-7 cytokine mediated by T-ALL cells can be involved in the oncogenic development of T-ALL and offer novel insights into T-ALL spreading.

5.
Cell Mol Life Sci ; 2019 Jun 19.
Artigo em Inglês | MEDLINE | ID: mdl-31218450

RESUMO

The sperm acrosome is a lysosome-related organelle that develops using membrane trafficking from the Golgi apparatus as well as the endolysosomal compartment. How vesicular trafficking is regulated in spermatids to form the acrosome remains to be elucidated. VPS13B, a RAB6-interactor, was recently shown involved in endomembrane trafficking. Here, we report the generation of the first Vps13b-knockout mouse model and show that male mutant mice are infertile due to oligoasthenoteratozoospermia. This phenotype was explained by a failure of Vps13b deficient spermatids to form an acrosome. In wild-type spermatids, immunostaining of Vps13b and Rab6 revealed that they transiently locate to the acrosomal inner membrane. Spermatids lacking Vps13b did not present with the Golgi structure that characterizes wild-type spermatids and showed abnormal targeting of PNA- and Rab6-positive Golgi-derived vesicles to Eea1- and Lamp2-positive structures. Altogether, our results uncover a function of Vps13b in the regulation of the vesicular transport between Golgi apparatus, acrosome, and endolysosome.

6.
J Mol Med (Berl) ; 97(5): 633-645, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30843084

RESUMO

Cohen syndrome (CS) is a rare genetic disorder due to mutations in VPS13B gene. Among various clinical and biological features, CS patients suffer from inconsistent neutropenia, which is associated with recurrent but minor infections. We demonstrate here that this neutropenia results from an exaggerate rate of neutrophil apoptosis. Besides this increased cell death, which occurs in the absence of any endoplasmic reticulum stress or defect in neutrophil elastase (ELANE) expression or localization, all neutrophil functions appeared to be normal. We showed a disorganization of the Golgi apparatus in CS neutrophils precursors, that correlates with an altered glycosylation of ICAM-1 in these cells, as evidenced by a migration shift of the protein. Furthermore, a striking decrease in the expression of SERPINB1 gene, which encodes a critical component of neutrophil survival, was detected in CS neutrophils. These abnormalities may account for the excessive apoptosis of neutrophils leading to neutropenia in CS. KEY MESSAGES: Cohen syndrome patients' neutrophils display normal morphology and functions. Cohen syndrome patients' neutrophils have an increased rate of spontaneous apoptosis compared to healthy donors' neutrophils. No ER stress or defective ELA2 expression or glycosylation was observed in Cohen syndrome patients' neutrophils. SerpinB1 expression is significantly decreased in Cohen syndrome neutrophils as well as in VPS13B-deficient cells.

7.
Hum Reprod ; 34(4): 612-622, 2019 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-30865273

RESUMO

STUDY QUESTION: Do assisted reproductive technologies (ARTs) impact on the expression of transposable elements (TEs) in preimplantation embryos? SUMMARY ANSWER: The expression of all TE families is globally increased with mouse embryo culture with differences according to culture medium composition. WHAT IS KNOWN ALREADY: Mammalian genomes are subject to global epigenetic reprogramming during early embryogenesis. Whether ARTs could have consequences on this period of acute epigenetic sensitivity is the matter of intense research. So far, most studies have examined the impact of ARTs on the regulation of imprinted genes. However, very little attention has been given to the control of TEs, which exceed by far the number of genes and account for half of the mammalian genomic mass. This is of particular interest as TEs have the ability to modulate gene structure and expression, and show unique regulatory dynamics during the preimplantation period. STUDY DESIGN, SIZE, DURATION: Here, we evaluated for the first time the impact of ART procedures (superovulation, in-vitro fertilisation and embryo culture) on the control of different TE types throughout preimplantation development of mouse embryos. We also made use of a mouse model carrying a LINE-1 retrotransposition-reporter transgene to follow parental patterns of transmission and mobilisation. PARTICIPANTS/MATERIALS, SETTING, METHODS: Hybrid B6CBA/F1 mice were used for the expression analyses. Relative TE expression was evaluated by using the nCounter quantification methodology (Nanostring®). This quantitative method allowed us to simultaneously follow 15 TE targets. Another technique of quantification (RTqPCR) was also used.A mouse model carrying a LINE-1 retrotransposition-reporter transgene (LINE-1 GF21) was used to follow parental patterns of transmission and mobilisation. MAIN RESULTS AND THE ROLE OF CHANCE: We found that the superovulation step did not modify the dynamics nor the level of TE transcription across the preimplantation period. However, upon in-vitro culture, TE expression was globally increased at the blastocyst stage in comparison with in-vivo development. Finally, by monitoring the transmission and mobilisation of a transgenic LINE-1 transposon, we found that in-vitro fertilisation may alter the mendelian rate of paternal inheritance. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: Even though the Nanostring results concerning the dynamics of transcription throughout preimplantation development were based on pools of embryos originating from several females, only two pools were analysed per developmental stage. However, at the blastocyst stage, consistent expressional results were found between the Nanostring technology and the other technique of quantification used, RTqPCR. WIDER IMPLICATIONS OF THE FINDINGS: Our findings highlight the sensitivity of TEs to the ART environment and their great potential as biomarkers of culture medium-based effects. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by funding from the 'Agence de la Biomedecine', 'Conseil Régional de Bourgogne' and 'RCT grant from INSERM-DGOS'. The authors have no conflicts of interest to declare.

8.
Genet Med ; 21(9): 2025-2035, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-30723320

RESUMO

PURPOSE: Lanosterol synthase (LSS) gene was initially described in families with extensive congenital cataracts. Recently, a study has highlighted LSS associated with hypotrichosis simplex. We expanded the phenotypic spectrum of LSS to a recessive neuroectodermal syndrome formerly named alopecia with mental retardation (APMR) syndrome. It is a rare autosomal recessive condition characterized by hypotrichosis and intellectual disability (ID) or developmental delay (DD), frequently associated with early-onset epilepsy and other dermatological features. METHODS: Through a multicenter international collaborative study, we identified LSS pathogenic variants in APMR individuals either by exome sequencing or LSS Sanger sequencing. Splicing defects were assessed by transcript analysis and minigene assay. RESULTS: We reported ten APMR individuals from six unrelated families with biallelic variants in LSS. We additionally identified one affected individual with a single rare variant in LSS and an allelic imbalance suggesting a second event. Among the identified variants, two were truncating, seven were missense, and two were splicing variants. Quantification of cholesterol and its precursors did not reveal noticeable imbalance. CONCLUSION: In the cholesterol biosynthesis pathway, lanosterol synthase leads to the cyclization of (S)-2,3-oxidosqualene into lanosterol. Our data suggest LSS as a major gene causing a rare recessive neuroectodermal syndrome.

9.
Genet Med ; 20(6): 645-654, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29095811

RESUMO

PurposeCongenital anomalies and intellectual disability (CA/ID) are a major diagnostic challenge in medical genetics-50% of patients still have no molecular diagnosis after a long and stressful diagnostic "odyssey." Solo clinical whole-exome sequencing (WES) was applied in our genetics center to improve diagnosis in patients with CA/ID.MethodsThis retrospective study examined 416 consecutive tests performed over 3 years to demonstrate the effectiveness of periodically reanalyzing WES data. The raw data from each nonpositive test was reanalyzed at 12 months with the most recent pipeline and in the light of new data in the literature. The results of the reanalysis for patients enrolled in the third year are not yet available.ResultsOf the 416 patients included, data for 156 without a diagnosis were reanalyzed. We obtained 24 (15.4%) additional diagnoses: 12 through the usual diagnostic process (7 new publications, 4 initially misclassified, and 1 copy-number variant), and 12 through translational research by international data sharing. The final yield of positive results was 27.9% through a strict diagnostic approach, and 2.9% through an additional research strategy.ConclusionThis article highlights the effectiveness of periodically combining diagnostic reinterpretation of clinical WES data with translational research involving data sharing for candidate genes.


Assuntos
Anormalidades Congênitas/genética , Deficiência Intelectual/genética , Sequenciamento Completo do Exoma/métodos , Bases de Dados Genéticas , Exoma , Testes Genéticos/métodos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Doenças Raras/genética , Estudos Retrospectivos , Análise de Sequência de DNA/métodos
10.
Hum Mol Genet ; 26(23): 4680-4688, 2017 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-28973148

RESUMO

The main identified function of BCL2 protein is to prevent cell death by apoptosis. Mouse knock-out for Bcl2 demonstrates growth retardation, severe polycystic kidney disease (PKD), grey hair and lymphopenia, and die prematurely after birth. Here, we report a 40-year-old male referred to for abdominal and thoracic aortic dissection with associated aortic root aneurysm, PKD, lymphocytopenia with a history of T cell lymphoblastic lymphoma, white hair since the age of 20, and learning difficulties. PKD, which was also detected in the father and sister, was related to an inherited PKD1 mutation. The combination of PKD with grey hair and lymphocytopenia was also reminiscent of Bcl2-/- mouse phenotype. BCL2 gene transcript and protein level were observed to be dramatically decreased in patient peripheral blood T-cells and in his aorta vascular wall cells, which was not detected in parents and sister T-cells, suggesting an autosomal recessive inheritance. Accordingly, spontaneous apoptosis of patient T-cells was increased and could be rescued through stimulation with an anti-CD3 antibody. Direct sequencing of BCL2 gene exons, promoter and 3'UTR region as well as BCL2 mRNA sequencing failed in identifying any pathogenic variant. Array-CGH was also normal and whole exome sequencing of the patient, parents and sister DNA did not detect any significant variant in genes encoding BCL2-interacting proteins. miRNA array identified an up-regulation of miR-181a, which is a known regulator of BCL2 expression. Altogether, miR-181a-mediated decrease in BCL2 gene expression could be a modifying factor that aggravates the phenotype of a PKD1 constitutive variant.


Assuntos
Rim Policístico Autossômico Dominante/genética , Proteínas Proto-Oncogênicas c-bcl-2/genética , Canais de Cátion TRPP/genética , Adulto , Animais , Apoptose/genética , Regulação para Baixo , Éxons , Predisposição Genética para Doença , Humanos , Masculino , Camundongos , Camundongos Knockout , MicroRNAs/metabolismo , Linhagem , Fenótipo , Rim Policístico Autossômico Dominante/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Canais de Cátion TRPP/metabolismo , Regulação para Cima
12.
Genet Med ; 19(9): 989-997, 2017 09.
Artigo em Inglês | MEDLINE | ID: mdl-28151489

RESUMO

PURPOSE: Postzygotic activating mutations of PIK3CA cause a wide range of mosaic disorders collectively referred to as PIK3CA-related overgrowth spectrum (PROS). We describe the diagnostic yield and characteristics of PIK3CA sequencing in PROS. METHODS: We performed ultradeep next-generation sequencing (NGS) of PIK3CA in various tissues from 162 patients referred to our clinical laboratory and assessed diagnostic yield by phenotype and tissue tested. RESULTS: We identified disease-causing mutations in 66.7% (108/162) of patients, with mutant allele levels as low as 1%. The diagnostic rate was higher (74%) in syndromic than in isolated cases (35.5%; P = 9.03 × 10-5). We identified 40 different mutations and found strong oncogenic mutations more frequently in patients without brain overgrowth (50.6%) than in those with brain overgrowth (15.2%; P = 0.00055). Mutant allele levels were higher in skin and overgrown tissues than in blood and buccal samples (P = 3.9 × 10-25), regardless of the phenotype. CONCLUSION: Our data demonstrate the value of ultradeep NGS for molecular diagnosis of PROS, highlight its substantial allelic heterogeneity, and confirm that optimal diagnosis requires fresh skin or surgical samples from affected regions. Our findings may be of value in guiding future recommendations for genetic testing in PROS and other mosaic conditions.Genet Med advance online publication 02 February 2017.


Assuntos
Classe I de Fosfatidilinositol 3-Quinases/genética , Estudos de Associação Genética , Testes Genéticos , Transtornos do Crescimento/diagnóstico , Transtornos do Crescimento/genética , Mutação , Adolescente , Adulto , Alelos , Substituição de Aminoácidos , Criança , Pré-Escolar , Classe I de Fosfatidilinositol 3-Quinases/metabolismo , Gerenciamento Clínico , Feminino , Predisposição Genética para Doença , Testes Genéticos/métodos , Genótipo , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Lactente , Recém-Nascido , Masculino , Mosaicismo , Fenótipo , Diagnóstico Pré-Natal , Análise de Sequência de DNA , Adulto Jovem
13.
Hum Mol Genet ; 24(23): 6603-13, 2015 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-26358774

RESUMO

Cohen Syndrome (CS) is a rare autosomal recessive disorder, with defective glycosylation secondary to mutations in the VPS13B gene, which encodes a protein of the Golgi apparatus. Besides congenital neutropenia, retinopathy and intellectual deficiency, CS patients are faced with truncal obesity. Metabolism investigations showed abnormal glucose tolerance tests and low HDL values in some patients, and these could be risk factors for the development of diabetes mellitus and/or cardiovascular complications. To understand the mechanisms involved in CS fat storage, we used two models of adipogenesis differentiation: (i) SGBS pre-adipocytes with VPS13B invalidation thanks to siRNA delivery and (ii) CS primary fibroblasts. In both models, VPS13B invalidation led to accelerated differentiation into fat cells, which was confirmed by the earlier and increased expression of specific adipogenic genes, consequent to the increased response of cells to insulin stimulation. At the end of the differentiation protocol, these fat cells exhibited decreased AKT2 phosphorylation after insulin stimulation, which suggests insulin resistance. This study, in association with the in-depth analysis of the metabolic status of the patients, thus allowed us to recommend appropriate nutritional education to prevent the occurrence of diabetes mellitus and to put forward recommendations for the follow-up of CS patients, in particular with regard to the development of metabolic syndrome. We also suggest replacing the term obesity by abnormal fat distribution in CS, which should reduce the number of inappropriate diagnoses in patients who are referred only on the basis of intellectual deficiency associated with obesity.


Assuntos
Adipogenia , Distribuição da Gordura Corporal , Diabetes Mellitus Tipo 2/fisiopatologia , Dedos/anormalidades , Insulina/fisiologia , Deficiência Intelectual/fisiopatologia , Microcefalia/fisiopatologia , Hipotonia Muscular/fisiopatologia , Miopia/fisiopatologia , Obesidade/fisiopatologia , Adolescente , Adulto , Criança , Pré-Escolar , Deficiências do Desenvolvimento/complicações , Deficiências do Desenvolvimento/fisiopatologia , Diabetes Mellitus Tipo 2/epidemiologia , Diabetes Mellitus Tipo 2/etiologia , Feminino , Dedos/fisiopatologia , Humanos , Deficiência Intelectual/complicações , Masculino , Microcefalia/complicações , Pessoa de Meia-Idade , Modelos Biológicos , Hipotonia Muscular/complicações , Mutação , Miopia/complicações , Obesidade/complicações , Degeneração Retiniana , Risco , Transdução de Sinais , Proteínas de Transporte Vesicular/genética , Adulto Jovem
14.
Clin Epigenetics ; 7: 87, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26300992

RESUMO

Today, there is growing interest in the potential epigenetic risk related to assisted reproductive technologies (ART). Much evidence in the literature supports the hypothesis that adverse pregnancy outcomes linked to ART are associated with abnormal trophoblastic invasion. The aim of this review is to investigate the relationship between epigenetic dysregulation caused by ART and subsequent placental response. The dialogue between the endometrium and the embryo is a crucial step to achieve successful trophoblastic invasion, thus ensuring a non-complicated pregnancy and healthy offspring. However, as described in this review, ART could impair both actors involved in this dialogue. First, ART may induce epigenetic defects in the conceptus by modifying the embryo environment. Second, as a result of hormone treatments, ART may impair endometrial receptivity. In some cases, it results in embryonic growth arrest but, when the development of the embryo continues, the placenta could bring adaptive responses throughout pregnancy. Amongst the different mechanisms, epigenetics, especially thanks to a finely tuned network of imprinted genes stimulated by foetal signals, may modify nutrient transfer, placental growth and vascularization. If these coping mechanisms are overwhelmed, improper maternal-foetal exchanges occur, potentially leading to adverse pregnancy outcomes such as abortion, preeclampsia or intra-uterine growth restriction. But in most cases, successful placental adaptation enables normal progress of the pregnancy. Nevertheless, the risks induced by these modifications during pregnancy are not fully understood. Metabolic diseases later in life could be exacerbated through the memory of epigenetic adaptation mechanisms established during pregnancy. Thus, more research is still needed to better understand abnormal interactions between the embryo and the milieu in artificial conditions. As trophectoderm cells are in direct contact with the environment, they deserve to be studied in more detail. The ultimate goal of these studies will be to render ART protocols safer. Optimization of the environment will be the key to improving the dialogue between the endometrium and embryo, so as to ensure that placentation after ART is similar to that following natural conception.

15.
Hum Mol Genet ; 24(12): 3314-21, 2015 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-25736213

RESUMO

Like genetic mutations, DNA methylation anomalies or epimutations can disrupt gene expression and lead to human diseases. However, unlike genetic mutations, epimutations can in theory be reverted through developmental epigenetic reprograming, which should limit their transmission across generations. Following the request for a parental project of a patient diagnosed with Silver-Russell syndrome (SRS), and the availability of both somatic and spermatozoa DNA from the proband and his father, we had the exceptional opportunity to evaluate the question of inheritance of an epimutation. We provide here for the first time evidence for efficient reversion of a constitutive epimutation in the spermatozoa of an SRS patient, which has important implication for genetic counseling.


Assuntos
Metilação de DNA , Epigênese Genética , Células Germinativas/metabolismo , Síndrome de Silver-Russell/genética , Adulto , Ilhas de CpG , Exoma , Feminino , Regulação da Expressão Gênica , Ordem dos Genes , Loci Gênicos , Impressão Genômica , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Fenótipo , Regiões Promotoras Genéticas , RNA Longo não Codificante/genética , Síndrome de Silver-Russell/diagnóstico
16.
Proc Natl Acad Sci U S A ; 111(29): 10592-7, 2014 Jul 22.
Artigo em Inglês | MEDLINE | ID: mdl-25002492

RESUMO

The hematopoietic system declines with age. Myeloid-biased differentiation and increased incidence of myeloid malignancies feature aging of hematopoietic stem cells (HSCs), but the mechanisms involved remain uncertain. Here, we report that 4-mo-old mice deleted for transcription intermediary factor 1γ (Tif1γ) in HSCs developed an accelerated aging phenotype. To reinforce this result, we also show that Tif1γ is down-regulated in HSCs during aging in 20-mo-old wild-type mice. We established that Tif1γ controls TGF-ß1 receptor (Tgfbr1) turnover. Compared with young HSCs, Tif1γ(-/-) and old HSCs are more sensitive to TGF-ß signaling. Importantly, we identified two populations of HSCs specifically discriminated by Tgfbr1 expression level and provided evidence of the capture of myeloid-biased (Tgfbr1(hi)) and myeloid-lymphoid-balanced (Tgfbr1(lo)) HSCs. In conclusion, our data provide a new paradigm for Tif1γ in regulating the balance between lymphoid- and myeloid-derived HSCs through TGF-ß signaling, leading to HSC aging.


Assuntos
Senescência Celular , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Fatores de Transcrição/metabolismo , Envelhecimento/metabolismo , Animais , Antígenos CD/metabolismo , Separação Celular , Senescência Celular/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Hematopoese/efeitos dos fármacos , Células-Tronco Hematopoéticas/efeitos dos fármacos , Camundongos , Células Mieloides/metabolismo , Fenótipo , Poliubiquitina/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptor do Fator de Crescimento Transformador beta Tipo I , Receptores de Superfície Celular/metabolismo , Transdução de Sinais/efeitos dos fármacos , Membro 1 da Família de Moléculas de Sinalização da Ativação Linfocitária , Fatores de Transcrição/deficiência , Fatores de Transcrição/genética , Fator de Crescimento Transformador beta1/farmacologia , Ubiquitinação/efeitos dos fármacos
17.
Am J Pathol ; 184(5): 1518-28, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24631023

RESUMO

Congenital muscular dystrophy, caused by mutations in LAMA2 (the gene encoding laminin α2 chain), is a severe and incapacitating disease for which no therapy is yet available. We have recently demonstrated that proteasome activity is increased in laminin α2 chain-deficient muscle and that treatment with the nonpharmaceutical proteasome inhibitor MG-132 reduces muscle pathology in laminin α2 chain-deficient dy(3K)/dy(3K) mice. Here, we explore the use of the selective and therapeutic proteasome inhibitor bortezomib (currently used for treatment of relapsed multiple myeloma and mantle cell lymphoma) in dy(3K)/dy(3K) mice and in congenital muscular dystrophy type 1A muscle cells. Outcome measures included quantitative muscle morphology, gene and miRNA expression analyses, proteasome activity, motor activity, and survival. Bortezomib improved several histological hallmarks of disease, partially normalized miRNA expression (miR-1 and miR-133a), and enhanced body weight, locomotion, and survival of dy(3K)/dy(3K) mice. In addition, bortezomib reduced proteasome activity in congenital muscular dystrophy type 1A myoblasts and myotubes. These findings provide evidence that the proteasome inhibitor bortezomib partially reduces laminin α2 chain-deficient muscular dystrophy. Investigation of the clinical efficacy of bortezomib administration in congenital muscular dystrophy type 1A clinical trials may be warranted.


Assuntos
Ácidos Borônicos/farmacologia , Ácidos Borônicos/uso terapêutico , Laminina/deficiência , Distrofia Muscular Animal/tratamento farmacológico , Distrofia Muscular Animal/patologia , Pirazinas/farmacologia , Pirazinas/uso terapêutico , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Peso Corporal/efeitos dos fármacos , Bortezomib , Células Cultivadas , Modelos Animais de Doenças , Fibrose , Regulação da Expressão Gênica/efeitos dos fármacos , Laminina/metabolismo , Locomoção/efeitos dos fármacos , Camundongos Endogâmicos C57BL , MicroRNAs/genética , MicroRNAs/metabolismo , Células Musculares/efeitos dos fármacos , Células Musculares/metabolismo , Células Musculares/patologia , Músculos/efeitos dos fármacos , Músculos/metabolismo , Músculos/patologia , Distrofia Muscular Animal/genética , Especificidade de Órgãos/efeitos dos fármacos , Especificidade de Órgãos/genética , Complexo de Endopeptidases do Proteassoma/metabolismo , Análise de Sobrevida
18.
Am J Med Genet A ; 164A(2): 522-7, 2014 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-24311531

RESUMO

Over one hundred VPS13B mutations are reported in Cohen syndrome (CS). Most cases exhibit a homogeneous phenotype that includes intellectual deficiency (ID), microcephaly, facial dysmorphism, slender extremities, truncal obesity, progressive chorioretinal dystrophy, and neutropenia. We report on a patient carrying two VPS13B splicing mutations with an atypical phenotype that included microcephaly, retinopathy, and congenital neutropenia, but neither obesity nor ID. RNA analysis of the IVS34+2T_+3AinsT mutation did not reveal any abnormal splice fragments but mRNA quantification showed a significant decrease in VPS13B expression. RNA sequencing analysis up- and downstream from the IVS57+2T>C mutation showed abnormal splice isoforms. In contrast to patients with typical CS, who express only abnormal VPS13B mRNA and truncated protein, a dose effect of residual normal VPS13B protein possibly explains the incomplete phenotype in the patient. This observation emphasizes that VPS13B analysis should be performed in cases of congenital neutropenia associated with retinopathy, even in the absence of ID, therefore extending the VPS13B phenotype spectrum.


Assuntos
Deficiência Intelectual/genética , Mutação , Neutropenia/congênito , Obesidade/genética , Fenótipo , Doenças Retinianas/genética , Proteínas de Transporte Vesicular/genética , Adulto , Análise Mutacional de DNA , Facies , Feminino , Ordem dos Genes , Humanos , Deficiência Intelectual/diagnóstico , Neutropenia/diagnóstico , Neutropenia/genética , Obesidade/diagnóstico , Linhagem , Doenças Retinianas/diagnóstico , Síndrome
19.
Hum Mol Genet ; 23(9): 2391-9, 2014 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-24334764

RESUMO

Cohen syndrome (CS) is a rare autosomal recessive disorder with multisytemic clinical features due to mutations in the VPS13B gene, which has recently been described encoding a mandatory membrane protein involved in Golgi integrity. As the Golgi complex is the place where glycosylation of newly synthesized proteins occurs, we hypothesized that VPS13B deficiency, responsible of Golgi apparatus disturbance, could lead to glycosylation defects and/or mysfunction of this organelle, and thus be a cause of the main clinical manifestations of CS. The glycosylation status of CS serum proteins showed a very unusual pattern of glycosylation characterized by a significant accumulation of agalactosylated fucosylated structures as well as asialylated fucosylated structures demonstrating a major defect of glycan maturation in CS. However, CS transferrin and α1-AT profiles, two liver-derived proteins, were normal. We also showed that intercellular cell adhesion molecule 1 and LAMP-2, two highly glycosylated cellular proteins, presented an altered migration profile on SDS-PAGE in peripheral blood mononuclear cells from CS patients. RNA interference against VPS13B confirmed these glycosylation defects. Experiments with Brefeldin A demonstrated that intracellular retrograde cell trafficking was normal in CS fibroblasts. Furthermore, early endosomes were almost absent in these cells and lysosomes were abnormally enlarged, suggesting a crucial role of VPS13B in endosomal-lysosomal trafficking. Our work provides evidence that CS is associated to a tissue-specific major defect of glycosylation and endosomal-lysosomal trafficking defect, suggesting that this could be a new key element to decipher the mechanisms of CS physiopathology.


Assuntos
Dedos/anormalidades , Deficiência Intelectual/metabolismo , Microcefalia/metabolismo , Hipotonia Muscular/metabolismo , Miopia/metabolismo , Obesidade/metabolismo , Antígenos CD/metabolismo , Moléculas de Adesão Celular/metabolismo , Deficiências do Desenvolvimento/metabolismo , Eletroforese em Gel de Poliacrilamida , Fibroblastos/metabolismo , Glicosilação , Complexo de Golgi/metabolismo , Humanos , Molécula 1 de Adesão Intercelular/metabolismo , Interferência de RNA , Degeneração Retiniana , Transferrina/metabolismo , Proteínas de Transporte Vesicular/metabolismo
20.
Am J Hum Genet ; 93(1): 141-9, 2013 Jul 11.
Artigo em Inglês | MEDLINE | ID: mdl-23810378

RESUMO

Short stature, hyperextensibility of joints and/or inguinal hernia, ocular depression, Rieger anomaly, and teething delay (SHORT) syndrome is a developmental disorder with an unknown genetic cause and hallmarks that include insulin resistance and lack of subcutaneous fat. We ascertained two unrelated individuals with SHORT syndrome, hypothesized that the observed phenotype was most likely due to de novo mutations in the same gene, and performed whole-exome sequencing in the two probands and their unaffected parents. We then confirmed our initial observations in four other subjects with SHORT syndrome from three families, as well as 14 unrelated subjects presenting with syndromic insulin resistance and/or generalized lipoatrophy associated with dysmorphic features and growth retardation. Overall, we identified in nine affected individuals from eight families de novo or inherited PIK3R1 mutations, including a mutational hotspot (c.1945C>T [p.Arg649Trp]) present in four families. PIK3R1 encodes the p85α, p55α, and p50α regulatory subunits of class IA phosphatidylinositol 3 kinases (PI3Ks), which are known to play a key role in insulin signaling. Functional data from fibroblasts derived from individuals with PIK3R1 mutations showed severe insulin resistance for both proximal and distal PI3K-dependent signaling. Our findings extend the genetic causes of severe insulin-resistance syndromes and provide important information with respect to the function of PIK3R1 in normal development and its role in human diseases, including growth delay, Rieger anomaly and other ocular affections, insulin resistance, diabetes, paucity of fat, and ovarian cysts.


Assuntos
Transtornos do Crescimento/genética , Hipercalcemia/genética , Resistência à Insulina/genética , Doenças Metabólicas/genética , Nefrocalcinose/genética , Fosfatidilinositol 3-Quinases/metabolismo , Análise Mutacional de DNA , Exoma , Feminino , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibroblastos/patologia , Predisposição Genética para Doença , Idade Gestacional , Glucose/metabolismo , Glucose/farmacologia , Humanos , Insulina/metabolismo , Insulina/farmacologia , Masculino , Mutação , Linhagem , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais
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