Current Research Focus:     Establish the physical basis of the dramatic force generation and exquisite cytoskeletal rearrangements that control cell division

Every time a cell divides, a microtubule-based structure called the mitotic spindle assembles around duplicate copies of condensed DNA, and then pulls one copy to each side of the dividing cell. In animal cells, an actin-based contractile ring then assembles around the mid-plane of the cell, pinching off daughter cells in a process called cytokinesis. Comprehensive biochemical studies have identified both accessory proteins that alter network morphology and motor proteins that exert force and drive out-of-equilibrium rearrangements. However, we are just beginning to develop a detailed understanding of how the various filament systems and proteins cooperate to generate the strong forces and complex movements that drive cells to literally tear themselves in two. Many important questions remain:

---

How do members of the kinesin family of motor proteins organize microtubules and generate forces in the mitotic spindle?

Kinesin-related molecular motors are known to modulate microtubule polymerization and exert force by crosslinking, sliding, and pushing microtubules. Using high-resolution optical trapping and fluorescence methods, we probe the nanomechanical properties of these mitotic kinesins, and explore how ensembles of motors cooperate to generate large forces in vivo. By comparing the force-dependent enzymology of different classes of motor proteins, we hope to understand how cellular function influences structure, kinetics, and force response in the kinesin superfamily.

---

What determines the mechanical strength of bipolar spindles and how are forces balanced at each stage of cell division?

Cytoplasm extracted from mature Xenopus laevis (frog) eggs is an established model system for biological studies of cell division. Extracts can be biochemically synchronized and controllably cycled through each stage of cell division, can be depleted of or supplemented with proteins of interest to control molecular composition, are easily imaged, and most importantly, are amenable to the mechanical perturbation required for biophysical characterization. We use these extracts to explore the mechanisms that drive self-assembly and force generation during cell division. We then recapitulate cellular behavior using only essential proteins in order to test theories of how small ensembles of proteins interact. Ultimately, we aim to develop predictive models of the force-balance and tension-sensing mechanisms that cells use to replicate with high fidelity.

---

How is mechanical response at long length scales related to the properties and distribution of individual motor and crosslinking proteins and filaments in cytoplasm?

The structural changes that drive cell division undoubtedly arise from complex interactions of multiple cytoskeletal filament systems with numerous motor and binding proteins. To probe these complicated and likely synergistic interactions, we study in vitro networks of purified cytoskeletal filaments that are copolymerized in the presence of crosslinking and motor proteins. We observe network morphology as a function of molecular composition, develop micromechanical methods to measure viscoelasticity and interaction forces in situ, and construct predictive models that relate the single-molecule structural properties of crosslinkers to network rheology. Our aim is to establish the molecular basis of cytoskeletal mechanics while simultaneously exploring how specific protein characteristics impart strength and flexibility to biomaterials. This work may ultimately lead to the generation of unique classes of materials based on novel engineered proteins with tailored nanomechanical properties.