Appointment period: 3/1/2012 to 2/28/2014

Project

Ultrastructural analysis of anaphase A/B spindle switch by correlative light and electron microscopy (CLEM)

The primary purpose of mitosis is the accurate segregation of genetic material. This is accomplished by a macromolecular machine, the mitotic spindle, which uses dynamic microtubules (MTs) and mitotic motors to separate chromosomes by orchestrating chromosome-to-pole motility (anaphase A) and spindle elongation (anaphase B). Our lab uses Drosophila syncytial embryos and a combination of in vivo, in vitro and in silico approaches to learn how the anaphase spindle functions as a macromolecular machine to elongate itself and pull apart sister chromosomes, and thus to provide insights into how defects in its function can give rise to aneuploidy, genomic instability, birth defects and cancer. Before anaphase B, the spindle achieve its steady-state length by balancing two opposing forces acting in concert: (1) an interpolar MTs (ipMTs) sliding mechanism pushing the spindle poles apart, generated by the homotetrameric kinesin-5 motor KLP61F, and (2) depolymerization of ipMTs at the spindle poles, generating poleward ipMT flux. Our working model suggests that, at anaphase A onset, ipMT depolymerization at the poles ceases, turning MTs flux “off” and tipping the balance of forces to allow outward ipMT sliding to push apart the spindle poles. My work focuses on investigating, at ultrastructural resolution, the reorganization of MTs and mitotic spindle components during anaphase B spindle elongation. Specifically, I am studying if KLP61f’s role in mitosis depends on it forming MT-MT crosslinks, associating with dynamic MT ends, binding to non-MT structures in spindles or assembling into multimeric arrays. I am also interested in determining to what extent the rate of anaphase B spindle elongation depends on KLP61F-mediated ipMT sliding.