This proposal examines mechanisms of cell motility at a fundamental level. We focus on clarifying the relationship between lamellipodial dynamics and focal complex formation, at the leading edge of a migrating cell. We are in a unique position to conduct these studies at the molecular level, because our published and preliminary data have unveiled a core mechanism whereby cortactin couples lamellipodial persistence with focal complex assembly.
In Aim 1 we will test the hypothesis that this coupling is in effect akin to a """"""""rack and pinion"""""""" steering mechanism for moving cells, which allows for a dominant lamellipodia to form and guide the cell in its direction.
In Aim 2 we will define how lamellipodia persistence is generated and positively regulated. Our existing data already conclusively show that binding of cortactin to branched actin is the basis for persistence, and we will test whether cortactin stabilizes branches, super-activates Arp2/3 complex, or both.
In Aim 3 we will define how persistence is spatially constrained and negatively regulated at areas of lamellipodia activity, by investigating molecular mechanisms of cortactin inhibition by phospholipids and cofilin, at the front and rear of the lamellipodia, respectively. An essential aspect of our studies is that we combine biochemistry with quantitative microscopy in order to determine not only whether molecular mechanisms can happen (e.g., in a test tube with purified components), but also whether in fact they occur in a living cell. These approaches include: chemotaxis and other cell motility assays, quantitative analyses of lamellipodial and adhesion dynamics in living cells, and careful biochemical characterization of actin binding protein mutants followed by quantification of cell phenotypes they produce. With the proposed studies on lamellipodial persistence, we are positioning ourselves at the interface of the adhesion and actin biology fields, whose integration will hopefully generate a new exciting discipline. Significance: Our studies are significant both for fundamental cell biology and human health. The molecular mechanisms we are attempting to solve are not only critical for cell motility, but also for the many cellular functions that depend on branched actin assembly, including vesicular trafficking and tissue morphogenesis. With respect to human health, these studies are particularly relevant to cancer metastasis, since cell motility is an essential component of cancer cell invasion. More directly, cortactin is well- documented to be overexpressed in a number of cancers via gene amplification, including 15% of breast and 30% of head and neck squamous cell cancer (HNSCC). Intriguingly, cortactin overexpression correlates with poor prognosis and decreased survival. Thus, the studies in this proposal are important for understanding both the fundamental regulation of dynamic branched actin assemblies and the possible role of cortactin specifically in cancer cell motility.
| Cleghorn, Whitney M; Branch, Kevin M; Kook, Seunghyi et al. (2015) Arrestins regulate cell spreading and motility via focal adhesion dynamics. Mol Biol Cell 26:622-35 |
| Hong, Nan Hyung; Qi, Aidong; Weaver, Alissa M (2015) PI(3,5)P2 controls endosomal branched actin dynamics by regulating cortactin-actin interactions. J Cell Biol 210:753-69 |
| Benesh, Emily C; Miller, Paul M; Pfaltzgraff, Elise R et al. (2013) Bves and NDRG4 regulate directional epicardial cell migration through autocrine extracellular matrix deposition. Mol Biol Cell 24:3496-510 |
| Bryce, Nicole S; Reynolds, Albert B; Koleske, Anthony J et al. (2013) WAVE2 regulates epithelial morphology and cadherin isoform switching through regulation of Twist and Abl. PLoS One 8:e64533 |
| Hoshino, Daisuke; Branch, Kevin M; Weaver, Alissa M (2013) Signaling inputs to invadopodia and podosomes. J Cell Sci 126:2979-89 |
| Weaver, Alissa M; Page, Jonathan M; Guelcher, Scott A et al. (2013) Synthetic and tissue-derived models for studying rigidity effects on invadopodia activity. Methods Mol Biol 1046:171-89 |
| Kirkbride, Kellye C; Hong, Nan Hyung; French, Christi L et al. (2012) Regulation of late endosomal/lysosomal maturation and trafficking by cortactin affects Golgi morphology. Cytoskeleton (Hoboken) 69:625-43 |
| Hoshino, Daisuke; Jourquin, Jerome; Emmons, Shane Weller et al. (2012) Network analysis of the focal adhesion to invadopodia transition identifies a PI3K-PKC? invasive signaling axis. Sci Signal 5:ra66 |
| Sung, Bong Hwan; Weaver, Alissa (2011) Cell-cell fusion: a new function for invadosomes. Curr Biol 21:R121-3 |
| Parekh, Aron; Ruppender, Nazanin S; Branch, Kevin M et al. (2011) Sensing and modulation of invadopodia across a wide range of rigidities. Biophys J 100:573-582 |
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