Accumulating evidence highlights the importance of studying the migration of cells within the context of their natural environment as manipulating the substrate on which a cell is migrating can have a dramatic impact on the mode/mechanisms employed by cells during migration. Central to this phenomenon is the requirement of adhesion to the ECM in order to gain traction during migration. Integrins constitute the main family of cell receptors involved in mediating cell-ECM interactions during motility. Whilst traditionally two-dimensional cell culture studies have placed emphasis on the importance of these receptors for spreading and migration, it has become evident that within more confined environments these receptors, at least for some cell types, are less crucial. In this research we utilise Drosophila embryonic hemocytes as an in vivo model for cell migration. We show that whilst hemocytes migrate within confined environments in vivo, these cells depend on integrins for powering both developmental and inflammatory migrations. Given the close association between these receptors and the actin cytoskeleton we were surprised to discover that removal of the main β integrin subunit, Myospheroid, did not affect cell spreading in vivo and had only a small impact on lamellipodial structure and dynamics. Furthermore we discovered that, in contrast to other cell types previously analysed, removal of this integrin subunit in hemocytes was not accompanied by an increase in the rate of actin retrograde flow within the protrusions, which we believe could reflect abrogation of a positive feedback between Rho, ROCK and Myosin II contraction. Instead, we discover a key role for integrins in regulating the microtubule cytoskeleton, in the maintenance of a polarised microtubule bundle, termed a ‘microtubule-arm’. Although the molecular mechanisms by which this stabilisation is coordinated have yet to be identified, this provides important insight into the co-regulation of adhesion and microtubule cytoskeleton important for the migratory behaviour of these cells.Cell migration reflects the complex and integrated regulation of the actin cytoskeleton by diverse families of actin regulatory proteins. Using hemocytes as a model system, we also explore the regulatory interactions between two main actin regulatory proteins, Diaphanous and Enabled, in vivo. Whilst the function of these proteins in the formation of filopodial protrusions is overlapping, recent research has highlighted the ability of these proteins to regulate the activity of one another. We find that co-expression of Enabled in hemocytes is able to rescue the morphological and migratory defects resulting from overexpression of active Diaphanous. Thus, data here presents Enabled as a negative regulator of Diaphanous, which may play an important role in the migration of hemocytes in vivo.
|Date of Award||1 Jul 2014|
|Supervisor||William Wood (Supervisor) & Andrew Chalmers (Supervisor)|
- cell migration