Microtubule bundles in the mitotic spindle: probing how mechanical and functional robustness emerge from molecular architecture

Project Summary/Abstract
The mitotic spindle is a microtubule-based machine that segregates chromosomes into two new
daughter cells when cells divide. Accurate spindle function is critical: mistakes lead to extra or missing
chromosomes, which are associated with cancer, birth defects, and miscarriage. Spindle function requires
robust coupling of biochemistry and mechanics. Yet, understanding how this self-organizing machine
generates the required forces in the right place at the right time remains a challenge. Our long term goal is to
determine how micron-scale mechanical properties of the spindle emerge from molecular-scale biochemistry.
We focus on microtubule bundles, which provide organization and underpin rigidity in the spindle and in other
microtubule-based structures. We do not understand what material properties bundling molecules impart to
spindle bundles, how their molecular properties allow them to do so, or how these emergent mechanical
properties are tuned for biological functions. To address these questions, we will measure quantitative
readouts of how bundles respond to perturbations that alter mechanics. We take a multi-system approach to
understanding bundle mechanics in mammalian kinetochore-fibers (k-fibers), which attach and segregate
chromosomes; in fission yeast S. pombe spindles, whose stereotyped organization facilitates probing how
specific crosslinker properties affect bundles overall; and in vitro, where we have more precise control.
Our approach is organized into two programs: (1) probing the molecular and mechanical organization of
spindle microtubule bundles, and (2) controlling spindle microtubule bundles to alter function through novel
mechanics. In Program 1, we will determine how k-fiber organization balances competing mechanical
constraints of robust force-transmission for chromosome segregation with flexibility to adapt to changing
spindle morphology. We will also develop new tools to measure force between microtubules within spindle
bundles, determining how these bundles effectively transmit force to achieve their mechanical functions. In
Program 2, we will determine how the geometric and mechanical properties of microtubule crosslinkers impart
bundle-scale properties that are adapted to particular functions. We will create engineered crosslinkers whose
mechanical and geometric properties we will control, and use them to build reconstituted microtubule bundles
in vitro, and to alter bundle properties in vivo. By measuring the response of these bundles to molecular-scale
changes, we will determine how micron-scale properties emerge.
In sum, the proposed work will map how molecular scale parts impart spindle bundles with properties
that balance competing mechanical constraints. In the long term, this approach may lead to new insight into
how altering the cell’s “building code” can be harnessed to target microtubule architectures with key roles in
disease, or to build novel architectures. This approach can extend to understand the emergent mechanics of
microtubule bundle architectures beyond the spindle, such as in cilia and axons.