RESUMO
Implementation of next-generation DNA sequencing (NGS) technology into routine diagnostic genome care requires strategic choices. Instead of theoretical discussions on the consequences of such choices, we compared NGS-based diagnostic practices in eight clinical genetic centers in the Netherlands, based on genetic testing of nine pre-selected patients with cardiomyopathy. We highlight critical implementation choices, including the specific contributions of laboratory and medical specialists, bioinformaticians and researchers to diagnostic genome care, and how these affect interpretation and reporting of variants. Reported pathogenic mutations were consistent for all but one patient. Of the two centers that were inconsistent in their diagnosis, one reported to have found 'no causal variant', thereby underdiagnosing this patient. The other provided an alternative diagnosis, identifying another variant as causal than the other centers. Ethical and legal analysis showed that informed consent procedures in all centers were generally adequate for diagnostic NGS applications that target a limited set of genes, but not for exome- and genome-based diagnosis. We propose changes to further improve and align these procedures, taking into account the blurring boundary between diagnostics and research, and specific counseling options for exome- and genome-based diagnostics. We conclude that alternative diagnoses may infer a certain level of 'greediness' to come to a positive diagnosis in interpreting sequencing results. Moreover, there is an increasing interdependence of clinic, diagnostics and research departments for comprehensive diagnostic genome care. Therefore, we invite clinical geneticists, physicians, researchers, bioinformatics experts and patients to reconsider their role and position in future diagnostic genome care.
Assuntos
Cardiomiopatias/diagnóstico , Testes Genéticos/normas , Genoma Humano , Sequenciamento de Nucleotídeos em Larga Escala/normas , Mutação , Proteínas de Ligação ao Cálcio/genética , Miosinas Cardíacas/genética , Cardiomiopatias/genética , Proteínas de Transporte/genética , Exoma , Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala/instrumentação , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Consentimento Livre e Esclarecido/legislação & jurisprudência , Ensaio de Proficiência Laboratorial/estatística & dados numéricos , MAP Quinase Quinase Quinases/genética , Cadeias Pesadas de Miosina/genética , Países Baixos , Proteínas Serina-Treonina QuinasesRESUMO
THOC5 is a nuclear/cytoplasmic protein member of the spliceosome complex which potentiates C/EBP expression in adipocyte differentiation. As C/EBP family members are important regulators of myelopoiesis and THOC5 is highly expressed in neutrophil/macrophage progenitor cells we assessed the role of THOC5 in cytokine-stimulated monocytic development. M-CSF stimulated maturation of the NFS60 cell line was associated with enhanced THOC5 expression and phosphorylation. THOC5 was also shown to form a complex with C/EBPbeta. Ectopic expression of THOC5 mimicked M-CSF mediated cell maturation and enhanced protein expression of the myeloid transcription factors C/EBPbeta, C/EBPalpha, Pu-1 and also GAB2 (a PI-3 Kinase and macrophage development regulator). Increased THOC5 expression also mimicked M-CSF stimulated increases in the lipid second messenger PtdInsP(3). Inhibition of THOC5-induced increases in PtdInsP(3) levels abrogated the elevated levels of C/EBPbeta. Thus THOC5 expression can potentiate receptor signalling to transcription factor expression and monocyte differentiation.