RESUMEN
Megalencephalic leukoencephalopathy with subcortical cysts (MLC) is a type of leukodystrophy characterized by white matter edema, and it is caused mainly by recessive mutations in MLC1 and GLIALCAM genes. These variants are called MLC1 and MLC2A with both types of patients sharing the same clinical phenotype. In addition, dominant mutations in GLIALCAM have also been identified in a subtype of MLC patients with a remitting phenotype. This variant has been named MLC2B. GLIALCAM encodes for an adhesion protein containing two immunoglobulin (Ig) domains and it is needed for MLC1 targeting to astrocyte-astrocyte junctions. Most mutations identified in GLIALCAM abolish GlialCAM targeting to junctions. However, it is unclear why some mutations behave as recessive or dominant. Here, we used a combination of biochemistry methods with a new developed anti-GlialCAM nanobody, double-mutants and cysteine cross-links experiments, together with computer docking, to create a structural model of GlialCAM homo-interactions. Using this model, we suggest that dominant mutations affect different GlialCAM-GlialCAM interacting surfaces in the first Ig domain, which can occur between GlialCAM molecules present in the same cell (cis) or present in neighbouring cells (trans). Our results provide a framework that can be used to understand the molecular basis of pathogenesis of all identified GLIALCAM mutations.
Asunto(s)
Encéfalo/metabolismo , Proteínas de Ciclo Celular/genética , Quistes/genética , Enfermedades Desmielinizantes del Sistema Nervioso Central Hereditarias/genética , Proteínas de la Membrana/genética , Conformación Proteica , Astrocitos , Encéfalo/patología , Encéfalo/ultraestructura , Proteínas de Ciclo Celular/ultraestructura , Cisteína/genética , Quistes/química , Quistes/patología , Edema/genética , Edema/patología , Células HeLa , Enfermedades Desmielinizantes del Sistema Nervioso Central Hereditarias/patología , Humanos , Proteínas de la Membrana/ultraestructura , Simulación del Acoplamiento Molecular , Mutación , Fenotipo , Multimerización de Proteína , Sustancia Blanca/metabolismo , Sustancia Blanca/patología , Sustancia Blanca/ultraestructuraRESUMEN
Cells perceive their microenvironment through chemical and physical cues. However, how the mechanical signals are interpreted during embryonic tissue deformation to result in specific cell behaviors is largely unknown. The Yap/Taz family of transcriptional co-activators has emerged as an important regulator of tissue growth and regeneration, responding to physical cues from the extracellular matrix, and to cell shape and actomyosin cytoskeletal changes. In this study, we demonstrate the role of Yap/Taz-TEAD activity as a sensor of mechanical signals in the regulation of the progenitor behavior of boundary cells during zebrafish hindbrain compartmentalization. Monitoring of in vivo Yap/Taz activity during hindbrain segmentation indicated that boundary cells responded to mechanical cues in a cell-autonomous manner through Yap/Taz-TEAD activity. Cell-lineage analysis revealed that Yap/Taz-TEAD boundary cells decreased their proliferative activity when Yap/Taz-TEAD activity ceased, which preceded changes in their cell fate from proliferating progenitors to differentiated neurons. Functional experiments demonstrated the pivotal role of Yap/Taz-TEAD signaling in maintaining progenitor features in the hindbrain boundary cell population.
Asunto(s)
División Celular/genética , Proteínas de Unión al ADN/fisiología , Péptidos y Proteínas de Señalización Intracelular/fisiología , Proteínas Nucleares/fisiología , Rombencéfalo/citología , Rombencéfalo/embriología , Células Madre/fisiología , Transactivadores/fisiología , Factores de Transcripción/fisiología , Proteínas de Pez Cebra/fisiología , Animales , Animales Modificados Genéticamente , Tipificación del Cuerpo/genética , Diferenciación Celular/genética , Movimiento Celular/genética , Proteínas de Unión al ADN/genética , Embrión no Mamífero , Péptidos y Proteínas de Señalización Intracelular/genética , Fenómenos Mecánicos , Mecanotransducción Celular/genética , Mecanotransducción Celular/fisiología , Neurogénesis/genética , Proteínas Nucleares/genética , Organogénesis/genética , Rombencéfalo/metabolismo , Transducción de Señal/genética , Células Madre/citología , Factores de Transcripción de Dominio TEA , Transactivadores/genética , Factores de Transcripción/genética , Proteínas Coactivadoras Transcripcionales con Motivo de Unión a PDZ , Proteínas Señalizadoras YAP , Pez Cebra/embriología , Pez Cebra/genética , Proteínas de Pez Cebra/genéticaRESUMEN
Elucidating the cellular and molecular mechanisms that regulate the balance between progenitor cell proliferation and neuronal differentiation in the construction of the embryonic brain demands the combination of cell lineage and functional approaches. Here, we generate the comprehensive lineage of hindbrain boundary cells by using a CRISPR-based knockin zebrafish transgenic line that specifically labels the boundaries. We unveil that boundary cells asynchronously engage in neurogenesis undergoing a functional transition from neuroepithelial progenitors to radial glia cells, coinciding with the onset of Notch3 signaling that triggers their asymmetrical cell division. Upon notch3 loss of function, boundary cells lose radial glia properties and symmetrically divide undergoing neuronal differentiation. Finally, we show that the fate of boundary cells is to become neurons, the subtype of which relies on their axial position, suggesting that boundary cells contribute to refine the number and proportion of the distinct neuronal populations.
Asunto(s)
División Celular Asimétrica , Pez Cebra , Animales , Diferenciación Celular , Neurogénesis , Rombencéfalo/metabolismo , Pez Cebra/metabolismo , Proteínas de Pez Cebra/genética , Proteínas de Pez Cebra/metabolismoRESUMEN
Cells in growing tissues receive both biochemical and physical cues from their microenvironment. Growing evidence has shown that mechanical signals are fundamental regulators of cell behavior. However, how physical properties of the microenvironment are transduced into critical cell behaviors, such as proliferation, progenitor maintenance, or differentiation during development, is still poorly understood. The transcriptional co-activators YAP/TAZ shuttle between the cytoplasm and the nucleus in response to multiple inputs and have emerged as important regulators of tissue growth and regeneration. YAP/TAZ sense and transduce physical cues, such as those from the extracellular matrix or the actomyosin cytoskeleton, to regulate gene expression, thus allowing them to function as gatekeepers of progenitor behavior in several developmental contexts. The Notch pathway is a key signaling pathway that controls binary cell fate decisions through cell-cell communication in a context-dependent manner. Recent reports now suggest that the crosstalk between these two pathways is critical for maintaining the balance between progenitor maintenance and cell differentiation in different tissues. How this crosstalk integrates with morphogenesis and changes in tissue architecture during development is still an open question. Here, we discuss how progenitor cell proliferation, specification, and differentiation are coordinated with morphogenesis to construct a functional organ. We will pay special attention to the interplay between YAP/TAZ and Notch signaling pathways in determining cell fate decisions and discuss whether this represents a general mechanism of regulating cell fate during development. We will focus on research carried out in vertebrate embryos that demonstrate the important roles of mechanical cues in stem cell biology and discuss future challenges.