Engineering a synthetic fission yeast chromosome

The spindle checkpoint is medically important, biologically fascinating, but mechanistically complex. All the major components have been known for decades, yet we still do not know how they work. Simple gene deletion/mutation approaches have reached their limitation as spindle checkpoint signals are transmitted by modulating protein-protein interactions and/or conformational changes, sometimes in response to post-translational modifications. We need a new approach to overcome these limitations.

I will use synthetic biology to artificially generate and transmit checkpoint signals from ectopic, non-centromere, locations. Signalling modules will be assembled de novo and assayed for activity. In the longer term we envisage analyses of multiple pathways to study cross-talk and inhibition. Our goals are: to definitively test models of spindle checkpoint signalling and find out how signalling scaffold is assembled and activated to generate a diffusible checkpoint signal. We will also construct synthetic arrays to provide a platform for functional dissection of this pathway in yeast. We will combine the array with chemical inducers of dimerisation (CIDs) and the SunTag. This will enable us to assemble and regulate a mitotic checkpoint complex (MCC) generator.

This research will help us to uncover the mechanisms that underlie the spindle checkpoint.