טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentAbu Shah Enas
SubjectReconstitution of Actin-Based Motility in a Cell-Like
Compartment - Towards the Generation of an
Artificial Crawling Cell
DepartmentDepartment of Nanoscience and Nanotechnology
Supervisor Professor Kinneret Keren
Full Thesis textFull thesis text - English Version


Abstract

The actin cytoskeleton plays a central role in many cellular processes including polarisation, cell shape determination, intracellular transport, cell division and movement. The structure and function of the cytoskeleton arise from the self-organised dynamics of numerous molecular building blocks. One of the prevailing cytoskeletal structures in the cell is the actin cortex. This thin actin shell underneath the plasma membrane provides mechanical support to the cell and plays a central role in cell division, polarisation and motility. In all these contexts, the homogeneous actin cortex has to break symmetry to generate polar cytoskeletal dynamics. Despite extensive research, the mechanisms responsible for regulating cortical dynamics in vivo and inducing symmetry breaking are still unclear. More generally, the principles governing large-scale coordination of cytoskeletal components are still not well-understood.  Biomimetic systems for reconstituting actin dynamics in vitro, detached from the complexity of the whole cell, present a powerful tool for studying the organisation and function of the cytoskeleton. Using a bottom-up reconstitution approach, where individual components are used to give rise to more complex assemblies, we have developed a model system that self-organises into dynamic actin cortices at the inner interface of water-in-oil emulsions. This artificial system undergoes spontaneous symmetry breaking, driven by myosin-induced cortical actin flows, which appear remarkably similar to the initial polarisation of the embryo in many species. The contractile behaviour of the reconstituted cortices exhibits a sharp temperature-dependent transition, facilitating the use of temperature as an external parameter to control the onset of symmetry breaking. The reconstituted cortices further display occasional local detachment from the interface as observed in cellular blebs, and polar force generation which can lead to deformation of the interface. Our in vitro model system recapitulates the rich cortical dynamics observed in vivo, allowing us to reveal the basic biophysical and biochemical requirements for actin cortex formation and symmetry breaking. Moreover, using this synthetic system we were able to observe artificial motility of droplets. Hence, this set-up paves the way towards the realisation of a minimal model system for studying actin dynamics and cell motility.