RESUMO
We identified a human embryonic stem cell subline that fails to respond to the differentiation cues needed to obtain endoderm derivatives, differentiating instead into extra-embryonic mesoderm. RNA-sequencing analysis showed that the subline has hyperactivation of the WNT and BMP4 signalling. Modulation of these pathways with small molecules confirmed them as the cause of the differentiation impairment. While activation of WNT and BMP4 in control cells resulted in a loss of endoderm differentiation and induction of extra-embryonic mesoderm markers, inhibition of these pathways in the subline restored its ability to differentiate. Karyotyping and exome sequencing analysis did not identify any changes in the genome that could account for the pathway deregulation. These findings add to the increasing evidence that different responses of stem cell lines to differentiation protocols are based on genetic and epigenetic factors, inherent to the line or acquired during cell culture.
Assuntos
Proteína Morfogenética Óssea 4/genética , Diferenciação Celular/genética , Células-Tronco Embrionárias Humanas/fisiologia , Proteínas Wnt/genética , Proteína Morfogenética Óssea 4/metabolismo , Células Cultivadas , Endoderma/citologia , Endoderma/fisiologia , Membranas Extraembrionárias/citologia , Membranas Extraembrionárias/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Células-Tronco Embrionárias Humanas/metabolismo , Humanos , Mesoderma/citologia , Mesoderma/fisiologia , Transdução de Sinais/genética , Transcriptoma , Proteínas Wnt/metabolismoRESUMO
ZNF335 plays an essential role in neurogenesis and biallelic variants in ZNF335 have been identified as the cause of severe primary autosomal recessive microcephaly in 2 unrelated families. We describe, herein, 2 additional affected individuals with biallelic ZNF335 variants, 1 individual with a homozygous c.1399 T > C, p.(Cys467Arg) variant, and a second individual with compound heterozygous c.2171_2173delTCT, p.(Phe724del) and c.3998A > G, p.(Glu1333Gly) variants with the latter variant predicted to affect splicing. Whereas the first case presented with early death and a severe phenotype characterized by anterior agyria with prominent extra-axial spaces, absent basal ganglia, and hypoplasia of the brainstem and cerebellum, the second case had a milder clinical presentation with hypomyelination and otherwise preserved brain structures on MRI. Our findings expand the clinical spectrum of ZNF335-associated microcephaly.
Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/genética , Microcefalia/genética , Degeneração Neural/genética , Neurogênese/genética , Proteínas Nucleares/genética , Alelos , Processamento Alternativo/genética , Gânglios da Base/patologia , Encéfalo/metabolismo , Encéfalo/fisiopatologia , Proteínas de Ligação a DNA , Feminino , Homozigoto , Humanos , Lactente , Recém-Nascido , Masculino , Microcefalia/epidemiologia , Microcefalia/fisiopatologia , Mutação , Degeneração Neural/epidemiologia , Degeneração Neural/fisiopatologia , Linhagem , Polimorfismo de Nucleotídeo Único/genética , Fatores de TranscriçãoRESUMO
Deletions on the long arm of the Y chromosome are a well-known cause of male infertility and it is generally accepted that deletions involving the AZFb region are not compatible with sperm production. Here, we report on two patients for whom basic diagnostic tests showed a deletion of the AZFb region. Unexpectedly, both patients had some residual sperm production. Subsequently, extension and additional analyses of the AZFb region disclosed an aberrant deletion pattern. Therefore, these results emphasize the need for a detailed and powerful analysis of cases where first-line Yq deletion tests reveal an AZFb deletion. Moreover, our study clearly demonstrated that only a very careful selection of test markers will avoid the pitfall of a 'no further treatment possible' wrongful conclusion.
Assuntos
Cromossomos Humanos Y , Fertilidade/genética , Infertilidade Masculina/genética , Oligospermia/genética , Transtornos do Cromossomo Sexual no Desenvolvimento Sexual/genética , Espermatogênese/genética , Deleção Cromossômica , Cromossomos Humanos Y/genética , Predisposição Genética para Doença , Humanos , Masculino , Reação em Cadeia da Polimerase Multiplex , Oligospermia/diagnóstico , Oligospermia/fisiopatologia , Oligospermia/terapia , Fenótipo , Valor Preditivo dos Testes , Reprodutibilidade dos Testes , Fatores de Risco , Índice de Gravidade de Doença , Aberrações dos Cromossomos Sexuais , Contagem de Espermatozoides , Motilidade dos EspermatozoidesRESUMO
Deregulation of signaling pathways that control differentiation, expansion and migration of neural crest-derived melanoblasts during normal development contributes also to melanoma progression and metastasis. Although several epithelial-to-mesenchymal (EMT) transcription factors, such as zinc finger E-box binding protein 1 (ZEB1) and ZEB2, have been implicated in neural crest cell biology, little is known about their role in melanocyte homeostasis and melanoma. Here we show that mice lacking Zeb2 in the melanocyte lineage exhibit a melanoblast migration defect and, unexpectedly, a severe melanocyte differentiation defect. Loss of Zeb2 in the melanocyte lineage results in a downregulation of the Microphthalmia-associated transcription factor (Mitf) and melanocyte differentiation markers concomitant with an upregulation of Zeb1. We identify a transcriptional signaling network in which the EMT transcription factor ZEB2 regulates MITF levels to control melanocyte differentiation. Moreover, our data are also relevant for human melanomagenesis as loss of ZEB2 expression is associated with reduced patient survival.