RESEARCH OVERVIEW

Our lab uses interdisciplinary methodologies to investigate the cell biology, biophysics, and biochemistry of mitosis. The lab is an ideal destination for students who want to obtain rigorous training encompassing molecular and cell biological methods, biochemistry, quantitative imaging, image analysis, and mathematical modeling.

• Uncovering mechanistic and regulatory integration of three kinetochore functions into a common protein framework

The human kinetochore is an incredibly complex protein machine. It consists of hundreds of copies and more than 10 different protein complexes. These proteins work together to implement three different functions: (1) a mechanical motor, (2) tension-sensitive regulatory mechanisms, and (3) signaling (see #2 below). How does the kinetochore integrate these functions into a conserved protein framework? Does the nanoscale organization of the kinetochore play a role in this integration? The Joglekar lab has been at the forefront of developing fluorescence microscopy assays to define the nanoscale architecture of the kinetochore. We have now initiated the next phase of research: use in vivo imaging to study the mechanics of kinetochore movement.

• Revealing the biochemical design and physiological tuning of the signaling cascade of the mitotic checkpoint

The mitotic checkpoint is a composite system consisting of biochemical sub-systems: (1) a mechanical switch that turns on and off: (2) a biochemical signaling cascade, which conveys the state of the mechanical switch to: (3) a biochemical switch that controls a cell-cycle transition. Investigation of this complex, composite system fall mainly into two categories: (A) identifying proteins, protein-protein interactions, and their regulation, and (B) computational analysis aimed at understanding the overall design of the signaling cascade. We have opened a new front: using extensive, quantitative analysis of the individual biochemical reactions in vivo to build a detailed mathematical model of the mitotic checkpoint. Our work relies heavily on CRISPR’d cell lines to label various signaling proteins involved in the mitotic checkpoint and a range of live-cell fluorescence microscopy methods.

• Reverse engineering the kinetochore using de novo designed proteins

This is an exciting new front. Decades worth of investigations of the kinetochore have built a reasonably complete picture of its structure, function, and regulation. What comes next? We are pioneering efforts to reverse engineer the kinetochore using de novo designed proteins. This project will rely heavily on a comobination of in vivo and in vitro investigations with applied protein design. Please contact the PI if you are interested in learning more about this effort.