CAREER:Elucidating the Mechanism of Microtubule Dynamics through Cold-stable Tubulin Mutants

University Of Colorado At Denver, Aurora CO

Investigators

Abstract

The goal of this CAREER project is to understand the molecular mechanism by which microtubule polymers assemble and how changes in the protein conformation lead to microtubule disassembly. Microtubule polymers play vital cellular roles in transport and signaling processes in all higher organisms. These processes depend on the ability of microtubules to switch between states of assembly and disassembly. To gain new insight into the mechanism of microtubule dynamics (assembly and disassembly), the research program will generate tubulin mutants that mimic sequence differences from cold-adapted organisms, which exhibit highly stable microtubules, and determine how these mutations impact microtubule dynamics. The results of these experiments will address a critical gap in our understanding of how changes in tubulin structure determine its propensity to assemble or disassemble, thereby providing a major advance for the microtubule field. In addition, this project will impact our broader understanding of molecular mechanisms of cold adaptation. To further broaden the impact of the project, the Principal Investigator's laboratory will partner with students and teachers from Denver public high schools with underrepresented minority populations in a collaborative research-education program. This program will advance the research goals of the project while training high school teachers and students in the process of hypothesis-driven science, techniques in molecular biology, communication skills, and gaining exposure to the broader research community, thereby exciting and empowering future scientists. This project will uniquely contribute to ongoing structural/function studies of tubulin proteins and microtubules. Whereas crystallographic approaches provide detailed molecular snapshots of stable conformations of the tubulin proteins, the project's genetic approach is uniquely suited to define the roles of dynamic regions of tubulins that are not accessible to crystallography. The Principal Investigator will use a combination of sequence data from cold tolerant organisms and unbiased genetic screens to identify amino acid changes that enhance microtubule stability, and to determine the basis of enhanced stability using in silico protein modeling and quantitative assays of microtubule activity in cells and in a minimal system with purified proteins.