Behaviour, formation and function of oriented actin filament bundles in the establishment of fibroblast polarity.
Doctoral thesis, UCL (University College London).
To establish morphological polarity, a cell must form a rear that is distinct from its front. For subsequent directional migration it must continuously retract its rear and protrude its front, polarised in the direction of migration. Here, two methods for the analysis of the transition from a pre-polar to a polarised state are used; spontaneous and cue dependant polarisation. Time-lapse microscopy of primary chick embryo heart fibroblasts expressing GFP-actin showed that, prior to cell polarization, actin filament bundles in the cell body reorganise to form oriented filament bundles spanning the entire cell body. These actin bundles are already known to be necessary for cell migration. It was established that the first visual step in polarity acquisition was the organisation of actin from a disordered, non-oriented arrangement to an oriented array involving a range of acto-myosin II filament dynamics. This organisation was required to specify the position of the cell rear in spontaneous and in cue dependant polarity initiation. On average it took fibroblasts 5 minutes to spontaneously polarise once acto-myosin filaments were oriented. For polarization a localised net protrusion occurred at one end of the actin filament bundles, lagging retraction of the rear. Consequently, fully polarised cells started to migrate in the direction of the long axis of the acto-myosin II filament bundles. Jasplakinolide treatment and the introduction of either constitutively active LIM kinase or dominant negative ADF/Cofilin blocked formation of oriented actin filament bundles, with cells subsequently failing to polarise. It is therefore concluded that formation of oriented actin filament bundles in the cell body of fibroblasts requires ADF/Cofilin family of proteins, and is an early event needed to co-ordinate the spatial location of the cell rear and front during fibroblast polarization.
|Title:||Behaviour, formation and function of oriented actin filament bundles in the establishment of fibroblast polarity|
|Additional information:||Permission for digitisation not received|
|UCL classification:||UCL > School of Life and Medical Sciences > Faculty of Life Sciences > MRC/UCL Lab for Molecular Cell Biology|
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