The goal of this research is to understand and control the mechanical properties of the cytoskeletal filament called microtubules. Microtubules are nano-scale filaments, and the mechanical properties have ramifications for the shape of cells, cell division, and cell motility. Understanding how the mechanics of single microtubules is controlled is essential for creating an accurate quantitative and predictive model to relate the strength of single microtubules to the strength of the entire cytoskeletal network. This work will systematically measure and model how lattice defects, associated proteins, and post-translational modifications can affect microtubule rigidity. This work represents the first systematic study of how the microtubule lattice can directly affect the mechanics of single microtubules. The proposed approach will reveal new information about how the structure of microtubules can affect its mechanics. The results will have an impact on basic cell biology, since microtubules are essential for many cellular processes. Moreover, microtubules are an entropically-driven, self-assembled system made from identical protein subunits. Elucidating how mechanical properties of this system can be altered by lattice defects, external binding partners, and modifications to the subunits will have ramifications for other self-assembled condensed matter systems made of colloids, polymers, and proteins. Microtubules and their related proteins can be harnessed as a biological-scaffold for nano-scale assemblies. This work will enable bio-engineering of novel bio-memetic materials from the microtubule-cytoskeletal system. Determining the mechanical properties of the individual microtubule structure is essential for future engineering processes that may involve them. The research proposed here is truly interdisiciplinary, combining the fields of condensed matter physics, bio-engineering, materials science, and cell biology.
During the past four years of study, we have made significant progress to understand how the mechanical properties of microtubules are regulated in cells using direct experimentation and simulations. We had 5 published articles in peer-reviewed journals. Using molecular dynamics simulation, we showed how microtubule stiffness is regulated at the molecular level. Using these results, we modeled and simulated a whole microtubule of 1 micrometer in length. Experimentally, we determined a new analysis methods for determining the persistence length of microtubules and we were able to determine the uncertainty of the measurement for the first time. We deduced that the spread in the measured data is due to real differences in the cross-sectional radius of the microtubules, but that microtubule-associated proteins have a large impact on the measured persistence length, as well. The cross-sectional radius can vary from filament to filament and can even change within a single microtubule. We used this new method to make measurements on the effects of two different microtubule stabilizers including the chemotherapeutic drug Taxol with stabilizing binding proteins or different nucleotides. We were the first to measure the stiffness of microtubules with the non-hydrolyzable analog of GTP, GTP-gamma-S, and found that this nucleotide makes microtubules more flexible, even though another non-hydrolyzable nucleotide, GMPCPP, makes them stiffer. Our results have impacts on cell division and cell differentiation, which are important to understanding cell and tissue development and have implications for biomedical problems like cancer and birth defects. During this grant, we trained numerous students and researchers at all levels including 3 high school students, 10 undergraduates, 3 graduates students, 2 postdocs, and 2 high school teachers. Of the undergraduate college students, several have continued in science, including 3 pursuing graduate degrees, 1 working as a technician at the National Institutes of Health, and several as industrial technicians. One of the postdocs is now a faculty member at a teaching-emphasis college. Our research was features in artistic form on the cover of the Biophysical Journal, pictured below.
