Publicação
In vitro Modeling of Angelman Syndrome using the Neural Commitment of Patient-Specific iPSCs
| Resumo: | Angelman syndrome (AS) is a rare neurodevelopmental disorder with no cure, characterized by a severe developmental delay, speech impairment, motor dysfunction, and a characteristic happy behavior. AS is caused by the loss of functional UBE3A protein, an E3 ubiquitin ligase that targets proteins for degradation by the ubiquitin-proteasome system. Disruption of UBE3A activity in neurons impairs ubiquitination leading to accumulation of UBE3A-specific targets which results in neuronal dysfunction. The UBE3A gene undergoes tissue-specific genomic imprinting, an epigenetic phenomenon that leads to parent-of-origin-specific monoallelic expression. Thus, UBE3A is exclusively expressed from the maternally inherited allele in mature neurons, since the paternal allele is silenced upon neuronal differentiation due to transcriptional interference of an antisense RNA called UBE3A-ATS. Loss of maternal UBE3A allele results in complete absence of UBE3A protein ultimately leading to AS. Most of what we know about AS has been studied in animal models, however, this pre-clinical model has several limitations for direct translation to the human disease. Recently, somatic cell reprogramming technology and neuronal differentiation protocols have contributed to overcome such difficulty and establish in vitro models using patient-derived induced pluripotent stem cells (iPSCs). These models enable the generation of disease-relevant cells in limitless amounts while faithfully recapitulating neuronal developmental hallmarks, which allows the detailed elucidation of the disease mechanisms responsible for the clinical features observed in patients. In this project, we submitted healthy control and AS-derived iPSCs to a neuronal differentiation protocol for 80 days to obtain fully mature neurons. Our data suggest AS-derived neuronal cultures display increased neuronal apoptosis, enhanced gliogenesis and persistence or progenitor-like neural rosettes at the final stages of differentiation. We hypothesize that loss of UBE3A function impairs neuronal viability and consequently prompts neural progenitors to continue proliferating and differentiating, causing a precocious neuro-to-glia switch. Our results show the suitability of patient-derived iPSCs to be used as a disease modelling approach to study Angelman syndrome and future works will use this system to tackle the pathophysiological mechanisms at the molecular level that causes AS. |
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| Autores principais: | Vieira, Adriana Andrade |
| Assunto: | Síndrome de Angelman UBE3A imprinting genómico iPSCs diferenciação neuronal modelos de doença doença neurológica Teses de mestrado - 2020 |
| Ano: | 2020 |
| País: | Portugal |
| Tipo de documento: | dissertação de mestrado |
| Tipo de acesso: | acesso aberto |
| Instituição associada: | Universidade de Lisboa |
| Idioma: | inglês |
| Origem: | Repositório da Universidade de Lisboa |
| Resumo: | Angelman syndrome (AS) is a rare neurodevelopmental disorder with no cure, characterized by a severe developmental delay, speech impairment, motor dysfunction, and a characteristic happy behavior. AS is caused by the loss of functional UBE3A protein, an E3 ubiquitin ligase that targets proteins for degradation by the ubiquitin-proteasome system. Disruption of UBE3A activity in neurons impairs ubiquitination leading to accumulation of UBE3A-specific targets which results in neuronal dysfunction. The UBE3A gene undergoes tissue-specific genomic imprinting, an epigenetic phenomenon that leads to parent-of-origin-specific monoallelic expression. Thus, UBE3A is exclusively expressed from the maternally inherited allele in mature neurons, since the paternal allele is silenced upon neuronal differentiation due to transcriptional interference of an antisense RNA called UBE3A-ATS. Loss of maternal UBE3A allele results in complete absence of UBE3A protein ultimately leading to AS. Most of what we know about AS has been studied in animal models, however, this pre-clinical model has several limitations for direct translation to the human disease. Recently, somatic cell reprogramming technology and neuronal differentiation protocols have contributed to overcome such difficulty and establish in vitro models using patient-derived induced pluripotent stem cells (iPSCs). These models enable the generation of disease-relevant cells in limitless amounts while faithfully recapitulating neuronal developmental hallmarks, which allows the detailed elucidation of the disease mechanisms responsible for the clinical features observed in patients. In this project, we submitted healthy control and AS-derived iPSCs to a neuronal differentiation protocol for 80 days to obtain fully mature neurons. Our data suggest AS-derived neuronal cultures display increased neuronal apoptosis, enhanced gliogenesis and persistence or progenitor-like neural rosettes at the final stages of differentiation. We hypothesize that loss of UBE3A function impairs neuronal viability and consequently prompts neural progenitors to continue proliferating and differentiating, causing a precocious neuro-to-glia switch. Our results show the suitability of patient-derived iPSCs to be used as a disease modelling approach to study Angelman syndrome and future works will use this system to tackle the pathophysiological mechanisms at the molecular level that causes AS. |
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