Microtubule organizing centers (MTOCs) generate specific arrangements of microtubules that are essential for many cellular functions, including cell division, polarization, and migration. Different subcellular sites can function as the MTOC in order to accommodate these different processes, and here we seek to understand how different MTOC locations are established during development. To divide, animal cells use centrosomes as MTOCs, localizing specific proteins to grow and anchor microtubules in order to build the mitotic spindle; the microtubules then promote the correct formation of two daughter cells. In differentiating cells, like some epithelial cells, the location of the MTOC changes: microtubule-organizing proteins leave the centrosome and move to a membrane surface, an ?MTOC switch? establishing the membrane as the MTOC. Epithelial cells have distinctive shapes and stick together tightly to form cellular sheets, and the microtubule configuration generated by the membrane MTOC is important for the structure and function of these cells. However, this arrangement of microtubules does not permit division, raising an important question: when an epithelial cell divides, how does it change the location of its MTOC to reorganize its microtubules into the mitotic spindle? Microtubule reorganization is critical for regulating cell division and shape both in normal development and in cancer. Centrosomal defects are classic hallmarks of many cancers [1], and recent studies have shown that increasing the microtubule organizing activity of the centrosome causes epithelial cells to divide more readily and display invasive behavior [11,12]. Despite its importance, little is known about how cells control their microtubule organization and the consequent effect on cell division and shape. The goal of this proposal is to understand how epithelial cells establish specific MTOC locations for different functions and how the change in MTOC location is coordinated with cell division. The cell cycle is a tightly controlled process that uses a series of checkpoints to ensure that a cell is prepared for division. Here we propose a series of genetic and biochemical experiments in intestinal epithelial cells in the model organism C. elegans to determine how the cell cycle and MTOC location are coordinated. Our lab has recently optimized a new strategy for targeted protein degradation. This method, coupled with recent methods for genome editing, will allow us to uncover the molecular connection between cell cycle regulators and microtubule organization.
Aim 1 will use both new and classic genetic approaches to determine which cell cycle genes regulate the MTOC location.
Aim 2 will use complementary genetic and biochemical methods to identify which of these proteins tether microtubules to specific locations.
In Aim 3, a genetic screen will uncover new regulators of the coordination of MTOC location and cell cycle. As the majority of cancers are epithelial in origin, we expect that understanding how epithelial cells coordinate their MTOC with cell division will be relevant to cancer biology and may lead to the discovery of useful cancer biomarkers or therapeutic targets.
Cells organize long structural polymers called microtubules in different ways depending on whether they will divide or specialize, and defects in microtubule organization are frequently observed in many cancers [1]. Despite its widespread importance, how cells choose one microtubule configuration over another is poorly understood at the molecular level. This proposal seeks to understand the molecular mechanisms used by specialized cells to control the organization of their microtubules in coordination with cell division.
