RESUMO
Myelin basic protein (MBP) is a major component of the myelin sheaths of oligodendrocytes in the central nervous system and Schwann cells of the peripheral nervous system. Here we generated heterozygous fluorescent reporter of MBP gene in human induced pluripotent stem cells (hiPSCs). CRISPR/Cas9 genome editing technology was employed to knock in fused tdTomato fluorescent protein and EF1 alpha promoter-driven Bleomycin (Zeocin) resistance gene to the translational MBP C-terminal region. The resulting line, MBP-TEZ, showed tdTomato fluorescence upon oligodendrocyte differentiation. This reporter hiPSC line provides a precedential opportunity for monitoring human myelin formation and degeneration and purifying MBP-expressing cell lineages.
Assuntos
Células-Tronco Pluripotentes Induzidas , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Células-Tronco Pluripotentes Induzidas/citologia , Proteína Básica da Mielina/metabolismo , Proteína Básica da Mielina/genética , Bainha de Mielina/metabolismo , Linhagem Celular , Diferenciação Celular , Sistemas CRISPR-Cas , Genes Reporter , Proteínas Luminescentes/metabolismo , Proteínas Luminescentes/genética , Edição de Genes , Proteína Vermelha FluorescenteRESUMO
Rett syndrome is characterized by severe global developmental impairments with autistic features and loss of purposeful hand skills. Here we show that human induced pluripotent stem cell (hiPSC) lines derived from four Japanese female patients with Rett syndrome are generated from peripheral blood mononuclear cells using Sendai virus vectors. The generated hiPSC lines showed self-renewal and pluripotency and carried heterozygous frameshift, missense, or nonsense mutations in the MECP2 gene. Since the molecular pathogenesis caused by MECP2 dysfunction remains unclear, these cell resources are useful tools to establish disease models and develop new therapies for Rett syndrome.