ENDODEP · Molecular mechanisms of endosome departure from the spindle in asymmetric cell division

Molecular mechanisms of endosome departure from the spindle in asymmetric cell division

Asymmetric cell division (ACD) allows developing organisms to balance self-renewal and diversification of cells during development. ACD generates sibling cells that differ in terms of cell content and therefore adopt different fates. One means of achieving such biased segregation is by enrolling the endosomal machinery to package and ship elements of cell content to one or the other pole of the dividing cell. The first step in this asymmetric distribution: directed trafficking, has been described in good detail. The present project aimed at gaining understanding of the last step of the process: How are endosomes actually released from microtubules, into the target cell, after being trafficked? We use the first division of the Drosophila sensory organ precursor (SOP) as a model of ACD. The SOP undergoes ACD and generates an anterior –pIIb- and a posterior –pIIa- cell. PIIb and PIIa have different Notch status and one way the asymmetry of Notch levels is achieved, is through the biased trafficking of Notch already present in the mother cell. A population of Notch-containing Rab5 endosomes marked with the protein Smad Anchor for Receptor Activation (SARA) is trafficked posteriorly, sliding along microtubules organized in two overlapping antiparallel bundles. Sara endosomes are preferentially released into PIIa upon cytokinesis. Collectively, Sara endosomes are first targeted to the overlapping, antiparallel area of the spindle, accumulate at this particular location for 500ms after the onset of anaphase B, when they start vacating this area, showing a preferential departure towards PIIa. Previously, levels of Sara at the endosome surface were found to affect targeting to the spindle and Klp98A has been identified as the motor in charge of Sara endosomes; thus both Sara and Klp98A are involved in the timely release of Sara endosomes in the posterior cell. Here, we aimed at gaining understanding of the mechanisms underlying departure by i) clarifying the interactions between Sara and the motor, and ii) measuring endosomal motility parameters: we hypothesized identifying changes in these parameters could hint at spatial or temporal changes in the microtubule-motor interactions. We found that the motor-endosome association persists after departure. We also found that although the endosome motility is impaired in Sara mutant and in Klp98A mutant backgrounds, their ‘partner’ (resp. Klp98A and Sara) localizes correctly at the endosome surface. We reconstructed 91 endosome tracks and are in the process of extracting and analyzing their motility parameters.

Data: CORDIS, © European Union