The microtubule network is important for many cell functions such as intracellular transport or cell division, and our lab is particularly interested in how the microtubule cytoskeleton contributes to complex cell behaviors such as directed migration and cell shape changes. Generally, we approach these cell biological problems by a combination of quantitative confocal live cell imaging and biochemical methods. In addition to actin-driven lamella/lamellipodia protrusion in the direction of migration, migrating cells are characterized by orientation of the microtubule-organizing center and a large population of microtubules toward the leading edge. This polarity of the microtubule network is thought to be important for directional migration. Microtubules are dynamic polymers that stochastically switch between phases of polymerization and depolymerization. Stabilization of lamella microtubules is mediated by a bias of microtubule polymerization dynamics toward net growth downstream of intracellular signaling pathways, which may result from spatiotemporally regulated interactions of these MTs with the cortical cytoskeleton in the cell’s lamella.

+TIPs are a heterogeneous class of proteins that specifically accumulate at or near the plus ends of growing MTs in cells. Different +TIP complexes have been suggested to participate in MT linkage to cortical sites. CLIP-170 may mediate lamella MT attachments through interactions with activated Rac1, IQGAP1, and components of the dynein/dynactin complex. APC may link lamella MTs to the actin cytoskeleton through EB1-mediated binding to the formin mDia1, and CLASPs may link lamella MTs to adhesion sites by interactions with the spectraplakin ACF7 and the phosphatidylinositol (3,4,5)-trisphosphate sensor LL5β.

We are particularly interested in the function and regulation of CLASP proteins because CLASPs display very unusual intracellular spatiotemporal dynamics. In directionally migrating cells, CLASPs display a spatiotemporal gradient of microtubule association that correlates with the cell’s direction of migration. In fibroblasts, CLASPs accumulate in cortical patches near the cell’s leading edge. In epithelial cells, CLASPs associate along lamella MTs independently of their plus-end-tracking activity in the cell body. In addition, CLASPs may anchor MTs at the Golgi apparatus.

Several projects in the lab center on the molecular mechanism by which a gradient of CLASP-microtubule association is established in migrating cells, which may involve regulation of CLASP function by GSK3β phosphorylation. We identified GSK3β phosphorylation sites in CLASP and started to analyze the function of these sites in both interphase and mitotic cells. We are also interested in the basic molecular mechanisms by which +TIPs recognize growing microtubule ends, and specifically how a protein like CLASP can switch between different modes of microtubule binding. In addition, we recently observed that clusters of CLASP-decorated microtubules surround focal adhesion sites shortly before adhesion turnover in migrating epithelial cells, and are now investigating whether CLASPs function in adhesion turnover or stabilize microtubules in the vicinity of adhesions.

Other projects in the lab investigate microtubule function and dynamics in more physiological three-dimensional tissue culture models such as apical-basal polarized epithelial cells embedded in extracellular matrix, or in in vitro angiogenesis assays.