Forum 10/30/2003

“Microtubule dynamic instability in yeast mitosis”

Gaudenz Danuser – Department of Cell Biology, The Scripps Research Institute, La Jolla, CA

We study microtubule dynamic instability (MDI) as the driving mechanism of chromosome segregation in yeast mitosis. We test the hypothesis that during mitosis, MDI is not only powered by GTP hydrolysis, as suggested for microtubules in vitro, but by the motors and microtubule associated proteins (MAPs) of the kinetochore and the forces exerted on microtubule ends. To study chromosome segregation we are building a multi-scale model of the mitotic spindle. The model will span the range from a single tubulin dimer integrated in the microtubule lattice to the whole spindle apparatus represented by a complex mechanical system with hundreds of visco-elastic and contractile components. We rigorously calibrate the model parameters by solving the so-called inverse dynamics problem, i.e. given experimental trajectories of chromosome movement relative to the spindle poles and cell cortex we seek to estimate the various forces and kinetic parameters that dictate the kinematics of the spindle. Using genetic mutations, we can then identify the molecules which have a critical impact on force generation and MDI. To obtain large data pools supporting the robust solution of the inverse problem, we have implemented an automated readout of chromosome and spindle pole movement from 3D fluorescence light microscope movies. In yeast, the tracking of markers indicating the position of centromeric DNA and spindle pole becomes a challenge because of the short inter-marker distances and the small but significant displacements, both at or below the resolution limits of light microscopy. We will describe techniques that allow us to break the diffraction limit and extract spindle dynamics with super-resolution. We will report first implementations of numerical models of MDI and how we fit the governing parameters to data pools of chromosome trajectories.

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