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1.
Hum Mutat ; 2019 Nov 07.
Artigo em Inglês | MEDLINE | ID: mdl-31696996

RESUMO

Hereditary spastic paraplegia (HSP) is a group of disorders with predominant symptoms of lower-extremity weakness and spasticity. Despite the delineation of numerous genetic causes of HSP, a significant portion of individuals with HSP remain molecularly undiagnosed. Through exome sequencing, we identified five unrelated families with childhood-onset nonsyndromic HSP, all presenting with progressive spastic gait, leg clonus, and toe walking starting from 7 to 8 years old. A recurrent two-base pair deletion (c.426_427delGA, p.K143Sfs*15) in the UBAP1 gene was found in four families, and a similar variant (c.475_476delTT, p.F159*) was detected in a fifth family. The variant was confirmed to be de novo in two families and inherited from an affected parent in two other families. RNA studies performed in lymphocytes from one patient with the de novo c.426_427delGA variant demonstrated escape of nonsense-mediated decay of the UBAP1 mutant transcript, suggesting the generation of a truncated protein. Both variants identified in this study are predicted to result in truncated proteins losing the capacity of binding to ubiquitinated proteins, hence appearing to exhibit a dominant-negative effect on the normal function of the endosome-specific endosomal sorting complexes required for the transport-I complex.

2.
Genet Med ; 21(11): 2663, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31267042

RESUMO

In the Acknowledgements section of the paper the authors neglected to mention that the study was supported by a grant from the National Human Genome Research Institute (NHGRI) UM1HG007301 (S.H., M.L.T.). In addition, the award of MD was associated with the authors Michelle L. Thompson and Susan Hiatt instead of PhD. The PDF and HTML versions of the Article have been modified accordingly.

3.
Curr Opin Neurol ; 32(5): 722-727, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31343429

RESUMO

PURPOSE OF REVIEW: The purpose of this review is to highlight updates in the standard of care recommendations for DMD, and to describe approaches to and recent advances in genetic therapies for DMD. RECENT FINDINGS: Treatment of DMD patients with the corticosteroids prednisone or deflazacort remains the standard of care, and recent data shows that early treatment (as young as 5 months) with a weekend dosing regimen results in measurable improvement in motor outcomes. A mutation-specific therapy directed at restoring an open reading frame by skipping exon 51 is FDA-approved, and therapies directed at other exons are in trials. Gene replacement therapy shows significant promise in animal models, and trials are underway. Genome editing has received significant attention because of results in animal models, but challenges to implementation in humans remain. SUMMARY: The mainstay of treatment remains meeting well defined standards of care that have been shown to influence morbidity and mortality. These include use of systemic steroids, early nocturnal ventilatory support, appropriate cardiac care and prophylaxis, and wherever appropriate, scoliosis surgery. Early and accurate molecular diagnosis, along with appropriate and multidisciplinary care, provides the best opportunity for maximum benefit of both current standard and upcoming novel therapies for boys with DMD. Among the most promising of these is AAV-based gene replacement therapy, which is currently in clinical trials.

4.
Genet Med ; 2019 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-31155615

RESUMO

PURPOSE: Mediator is a multiprotein complex that allows the transfer of genetic information from DNA binding proteins to the RNA polymerase II during transcription initiation. MED12L is a subunit of the kinase module, which is one of the four subcomplexes of the mediator complex. Other subunits of the kinase module have been already implicated in intellectual disability, namely MED12, MED13L, MED13, and CDK19. METHODS: We describe an international cohort of seven affected individuals harboring variants involving MED12L identified by array CGH, exome or genome sequencing. RESULTS: All affected individuals presented with intellectual disability and/or developmental delay, including speech impairment. Other features included autism spectrum disorder, aggressive behavior, corpus callosum abnormality, and mild facial morphological features. Three individuals had a MED12L deletion or duplication. The other four individuals harbored single-nucleotide variants (one nonsense, one frameshift, and two splicing variants). Functional analysis confirmed a moderate and significant alteration of RNA synthesis in two individuals. CONCLUSION: Overall data suggest that MED12L haploinsufficiency is responsible for intellectual disability and transcriptional defect. Our findings confirm that the integrity of this kinase module is a critical factor for neurological development.

5.
Neuropediatrics ; 50(2): 96-102, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30665247

RESUMO

Next-generation sequencing is a powerful diagnostic tool, yet it has proven inadequate to establish a diagnosis in all cases of congenital hypotonia or childhood onset weakness. We sought to describe the impact of whole exome sequencing (WES), which has only recently become widely available clinically, on molecular diagnosis in the Nationwide Children's Hospital Neuromuscular clinics. We reviewed records of all patients in our clinic with pediatric onset of symptoms who had WES done since 2013. Patients were included if clinical suspicion was high for a neuromuscular disease. Clinical WES was performed in 30 families, representing 31 patients, all of whom were seen for hypotonia, weakness, or gait disturbance. Probands had between 2 and 12 genetic diagnostic tests prior to obtaining WES. A genetic diagnosis was established in 11 families (37%), and in 12 patients (39%), with mutations in 10 different genes. Five of these genes have only been associated with disease since 2013, and were not previously represented on clinically available disease gene panels. Our results confirm the utility of WES in the clinical setting, particularly for genetically heterogeneous syndromes. The availability of WES can provide an end to the diagnostic odyssey for parents and allow for expansion of phenotypes.

6.
Artigo em Inglês | MEDLINE | ID: mdl-30054298

RESUMO

We describe two unrelated patients, a 12-yr-old female and a 6-yr-old male, with congenital contractures and severe congenital muscular atrophy. Exome and genome sequencing of the probands and their unaffected parents revealed that they have the same de novo deletion in BICD2 (c.1636_1638delAAT). The variant, which has never been reported, results in an in-frame 3-bp deletion and is predicted to cause loss of an evolutionarily conserved asparagine residue at position 546 in the protein. Missense mutations in BICD2 cause autosomal dominant spinal muscular atrophy, lower-extremity predominant 2 (SMALED2), a disease characterized by muscle weakness and arthrogryposis of early onset and slow progression. The p.Asn546del clusters with four pathogenic missense variants in a region that likely binds molecular motor KIF5A. Protein modeling suggests that removing the highly conserved asparagine residue alters BICD2 protein structure. Our findings support a broader phenotypic spectrum of BICD2 mutations that may include severe manifestations such as cerebral atrophy, seizures, dysmorphic facial features, and profound muscular atrophy.

7.
Am J Med Genet A ; 176(5): 1207-1211, 2018 May.
Artigo em Inglês | MEDLINE | ID: mdl-29681093

RESUMO

Vici syndrome is a multisystem disorder characterized by agenesis of the corpus callosum, oculocutaneous hypopigmentation, cataracts, cardiomyopathy, combined immunodeficiency, failure to thrive, profound developmental delay, and acquired microcephaly. Most individuals are severely affected and have a markedly reduced life span. Here we describe an 8-year-old boy with a history of developmental delay, agenesis of the corpus callosum, failure to thrive, myopathy, and well-controlled epilepsy. He was initially diagnosed with a mitochondrial disorder, based in part upon nonspecific muscle biopsy findings, but mitochondrial DNA mutation analysis revealed no mutations. Whole exome sequencing revealed compound heterozygosity for two EPG5 variants, inherited in trans. One was a known pathogenic mutation in exon 13 (c.2461C > T, p.Arg821X). The second was reported as a variant of unknown significance found within intron 16, six nucleotides before the exon 17 splice acceptor site (c.3099-6C > G). Reverse transcription-polymerase chain reaction of the EPG5 mRNA showed skipping of exon 17-which maintains an open reading frame-in 77% of the transcript, along with 23% expression of wild-type mRNA suggesting that intronic mutations may affect splicing of the EPG5 gene and result in symptoms. However, the expression of 23% wild-type mRNA may result in a significantly attenuated Vici syndrome phenotype.

8.
Neuromuscul Disord ; 28(2): 116-121, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-29305136

RESUMO

The reading frame rule suggests that Duchenne muscular dystrophy (DMD) results from DMD mutations causing an out-of-frame transcript, whereas the milder Becker muscular dystrophy results from mutations causing an in-frame transcript. However, predicted nonsense mutations may instead result in altered splicing and an in-frame transcript. Here we report a 10-year-old boy with a predicted nonsense mutation in exon 42 who had a 6-minute walk time of 157% of that of age matched DMD controls, characterized as intermediate muscular dystrophy. RNA sequencing analysis from a muscle biopsy revealed only 6.0-9.8% of DMD transcripts were in-frame, excluding exon 42, and immunoblot demonstrated only 3.2% dystrophin protein expression. Another potential genetic modifier noted was homozygosity for the protective IAAM LTBP4 haplotype. This case suggests that very low levels of DMD exon skipping and dystrophin protein expression may result in amelioration of skeletal muscle weakness, a finding relevant to current dystrophin-restoring therapies.

10.
Endocrinology ; 149(12): 6006-17, 2008 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-18755801

RESUMO

The composition of the beta-cell exocytic machinery is very similar to that of neuronal synapses, and the developmental pathway of beta-cells and neurons substantially overlap. beta-Cells secrete gamma-aminobutyric acid and express proteins that, in the brain, are specific markers of inhibitory synapses. Recently, neuronal coculture experiments have identified three families of synaptic cell-surface molecules (neurexins, neuroligins, and SynCAM) that drive synapse formation in vitro and that control the differentiation of nascent synapses into either excitatory or inhibitory fully mature nerve terminals. The inhibitory synapse-like character of the beta-cells led us to hypothesize that members of these families of synapse-inducing adhesion molecules would be expressed in beta-cells and that the pattern of expression would resemble that associated with neuronal inhibitory synaptogenesis. Here, we describe beta-cell expression of the neuroligins, neurexins, and SynCAM, and show that neuroligin expression affects insulin secretion in INS-1 beta-cells and rat islet cells. Our findings demonstrate that neuroligins and neurexins are expressed outside the central nervous system and help confer an inhibitory synaptic-like phenotype onto the beta-cell surface. Analogous to their role in synaptic neurotransmission, neurexin-neuroligin interactions may play a role in the formation of the submembrane insulin secretory apparatus.


Assuntos
Moléculas de Adesão Celular Neuronais/metabolismo , Células Secretoras de Insulina/metabolismo , Insulina/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Processamento Alternativo , Animais , Western Blotting , Encéfalo/metabolismo , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Moléculas de Adesão Celular Neuronais/genética , Linhagem Celular Tumoral , Glicoproteínas/genética , Glicoproteínas/metabolismo , Guanilato Quinases/genética , Guanilato Quinases/metabolismo , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Microscopia Confocal , Proteínas do Tecido Nervoso/genética , Neuropeptídeos/genética , Neuropeptídeos/metabolismo , Interferência de RNA , Ratos , Reação em Cadeia da Polimerase Via Transcriptase Reversa
11.
Endocrinology ; 148(10): 4572-8, 2007 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-17584960

RESUMO

The enzyme glutamate decarboxylase-65 (GAD65) is a major autoantigen in autoimmune diabetes. The mechanism whereby autoreactivity to GAD65, an intracellular protein, is triggered is unknown, and it is possible that immunoreactive GAD65 is released by injured pancreatic islet beta-cells. There is a great need for methods by which to detect and monitor ongoing islet injury. If GAD65 were released and, furthermore, were able to reach the circulation, it could function as a marker of beta-cell injury. Here, a novel GAD65 plasma immunoassay is used to test the hypotheses that beta-cell injury induces GAD65 discharge in vivo and that discharged GAD65 reaches the bloodstream. Plasma GAD65 levels were determined in rats treated with alloxan, and with diabetogenic and low, subdiabetogenic doses of streptozotocin. beta-Cell injury resulted in GAD65 release into the circulation in a dose-dependent manner, and low-dose streptozotocin resulted in a more gradual increase in plasma GAD65 levels than did diabetogenic doses. Plasma GAD65 levels were reduced in rats that had undergone partial pancreatectomy and remained undetectable in mice. Together, these data demonstrate that GAD65 can be released into the circulation by injured beta-cells. Autoantigen shedding may contribute to the pathogenesis of islet autoimmunity in the multiple low-dose streptozocin model and perhaps, more generally, in other forms of autoimmune diabetes. These results demonstrate that, as is true with other tissues, islet injury, at least in some circumstances, can be monitored by use of discharged, circulating proteins. GAD65 is the first such confirmed protein marker of islet injury.


Assuntos
Aloxano/farmacologia , Glutamato Descarboxilase/sangue , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/enzimologia , Estreptozocina/farmacologia , Aloxano/administração & dosagem , Animais , Biomarcadores/sangue , Peptídeo C/sangue , Morte Celular , Relação Dose-Resposta a Droga , Meia-Vida , Injeções Intraperitoneais , Insulina/sangue , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pâncreas/efeitos dos fármacos , Pâncreas/patologia , Pancreatectomia , Ratos , Ratos Wistar , Estreptozocina/administração & dosagem
12.
Diabetes Technol Ther ; 8(2): 207-18, 2006 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-16734550

RESUMO

BACKGROUND: Glutamic acid decarboxylase-65 (GAD65) is a major autoantigen in autoimmune diabetes and is discharged from injured islet beta cells. GAD65 may also be released by transplanted islets undergoing immunological rejection. To test hypotheses regarding the utility of GAD65 as a biomarker for transplant rejection or diabetes-associated islet damage and also regarding the timing and instigators of GAD65 release in humans or animal models, a sensitive assay capable of measuring GAD65 in serum or plasma will be necessary. Ideally, this assay would also be resistant to interference by anti-GAD65 autoantibodies. METHODS: A novel, magnetic bead-based assay was developed based on GAD65 capture by a monoclonal antibody directed to the only region of the protein known not to be significantly targeted by autoantibodies. A subsequent denaturation step allows sensitive immunodetection to proceed using anti-GAD65 polyclonal antibodies that would otherwise potentially be blocked by bound autoantibodies. RESULTS: The GAD65 assay worked equally well with serum and plasma as with a solution of bovine serum albumin (BSA). The limit of blank was 31 pg/mL and did not differ significantly in the BSA solution (27 pg/mL). Mean recovery of GAD65 from the plasma of control subjects and GAD65 autoantibody-positive and -negative subjects with type 1 diabetes was 101 +/- 4.6%, 88 +/- 7.8%, and 99 +/- 7.0% (+/- SEM), respectively. The assay was used to quantify both recombinant GAD65 and the GAD65 content of human and rodent islets and other tissue extracts that were added to human plasma samples. CONCLUSIONS: A sensitive, autoantibody-resistant GAD65 assay has been developed that is compatible with detection in serum and plasma and therefore will likely also work with a variety of other biologic fluids. This assay may enable the use of circulating GAD65 as a biomarker of islet damage or transplant rejection and will facilitate in vivo studies of the pathogenesis of anti-GAD65 autoreactivity.


Assuntos
Glutamato Descarboxilase/sangue , Fragmentos de Peptídeos/sangue , Autoanticorpos , Autoantígenos/sangue , Diabetes Mellitus/sangue , Diabetes Mellitus/imunologia , Glutamato Descarboxilase/imunologia , Humanos , Técnicas de Imunoadsorção , Células Secretoras de Insulina/metabolismo , Fragmentos de Peptídeos/imunologia , Plasma/química , Proteínas Recombinantes/sangue , Soro/química , Extratos de Tecidos/química
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