Mechanics and Cellular Mechanisms Driving Zebrafish Epiboly = Mecánica y mecanismos celulares responsables de la epibolia del pez cebra

• Autores: Amayra Noemi Hernández Vega
• Directores de la Tesis: David Bueno i Torrens (dir. tes.), Enrique Martín Blanco (dir. tes.)
• Lectura: En la Universitat de Barcelona ( España ) en 2015
• Idioma: inglés
• Tribunal Calificador de la Tesis: Xavier Trepat Guixer (presid.), Cristina Pujades Corbi (secret.), Damian Brunner (voc.)
• Materias:
  • Física
    • Mecánica
• Enlaces
  • Tesis en acceso abierto en:  TDX  Dipòsit Digital de la UB 
• Resumen

  • Epithelial layers constitute the skeleton of early embryos and most tissues and organs and their remodelling during development is essential for reshaping the embryo and for tissue architecture. Epithelial expansion in particular, has fundamental roles during early embryogenesis in both vertebrates and invertebrates. Some well-established invertebrate models have contributed greatly to our knowledge of this process. However, we are still far from having a global understanding of the cellular, molecular and mechanical changes involved in this process, the diversity, and relationship between them. With this in mind, we decided to explore a less well known model for epithelial expansion, epiboly in the vertebrate zebrafish. Epiboly is the expansion of the blastoderm around a big yolk cell to finally engulf it. Three different layers are involved in this process, an epithelial layer, a mesenquimal layer and a big yolk cell. Although teleost epiboly has been studied for many years a clear understanding of the process was still missing. We analysed the cellular, molecular and mechanical elements involved in this process and found that epithelial expansion is in this process a passive event driven by the pulling of the adjacent layer, the yolk syncytium. The increase in area of this epithelia is achieved by cell shape changes (flattening) and the RhoGTPase Rac1 activity seems to be necessary for these passive changes in shape. The contraction in the yolk syncytium is accompanied by the formation of membrane folds and endocytic vesicles in this area and the different mechanical properties of the elements at both sides of these contractile domains are, together with endocytosis, essential to understand the expansion. In relation to this, we found that Rab5ab activity in the yolk is essential for the expansion and for doming of the internal part of the yolk. In addition, we showed that the main constituent of the embryo at this stage, the yolk granules, behave as an hydrodynamic fluid at low Reynolds number that passively flow during epiboly by the activity at the surface. We learned that the spherical geometry of the embryo together with volume conservation and the transmission of forces between the different elements involved in the process are essential to understand the changes observed in the blastoderm during this process and its global coordination. We generated a non-intrusivemethodology to extract the mechanical changes involved in a given morphogenetic event from microscopy data based on the relation between the active elements (elastic, visco-elastic) and the passive ones (fluids). We applied this method to the process of epiboly and validated the results obtained by atomic force microscopy (AFM) indentation and laser microsurgery experiments. Finally, we generated an enhancer trap screen using the Gal4/UAS binary system with the aim of being able to spatially restrict gene expression during epiboly. However, and although we found several interesting lines that drove specific gene expression, we could not find any with sufficient early expression to be useful for our epiboly studies. Overall, we learnt that an isotropic actomyosin contraction generates an anisotropic stress pattern and movement by the properties of the surrounding elements. To get a precise understanding of epiboly we had to consider the transmission of forces between the different layer and volume conservation. For that, it was important to take into account both the contribution from the active elements (cortex) and from the passive ones (fluid). The role that unselective membrane removal has in morphogenesis has been barely explored. We anticipate that membrane tension and removal and its relationship to actomyosin contraction and shape changes will become an emerging and exciting field and that zebrafish epiboly will become a great model to study these relationships