Molecular mechanisms of myosin-X and filopodial function

About This Grant

Myosin-X (Myo10) is a molecular motor crucial for the formation and function of filopodia, finger-like protrusions cells use interact with their surroundings in processes such as brain development, blood vessel formation, and the spread of cancer cells. We have shown that Myo10 localizes to the tips of filopodia, increases the number and length of filopodia, and moves within filopodia in a process known as intrafilopodial motility. Our generation of Myo10 knock-out mice showed that loss of Myo10 causes developmental defects in brain, eye, and blood vessels, but is not essential for survival of adult mice. Myo10 promotes tumor growth and invasion in many cancers, including breast, lung, and melanoma. Myo10 also has important functions in cell division, where it is required for spindle orientation and for clustering the excess centrosomes that are a hallmark of cancer cells. Myo10’s important roles in biology, plus the need to understand the fundamental cell biology of filopodia, make it essential to investigate the molecular mechanisms of Myo10 and filopodial function at the cellular and organismal levels. To fill these knowledge gaps, we will address the following: -How does Myo10 promote filopodia and what are its molecular cargos? -What are the functions of headless Myo10, a form of Myo10 that lacks the motor domain and is expressed in brain and stem cells? -Purify filopodia and use modern proteomics approaches to identify and quantify the full set of their molecular components, including the filopodial cytoskeleton, tip, and plasma membrane. -What are Myo10’s organismal functions in epithelial tissues such as kidney where it localizes basolaterally and in eye where KO results in major developmental defects. Because filopodia are a major cellular organelle whose purification has not been reported, we will combine our recent progress purifying filopodia with quantitative proteomics to identify the molecular components of filopodia. We will also take advantage of the extensive set of tools and techniques we have established to investigate Myo10 and filopodia, including Myo10 floxed and knock-out mice, KO and stable cell lines, and deletion and point mutant constructs. Although the other members of the MyTH4-FERM family of myosins in vertebrates have important roles in human physiology and disease at the tips of other protrusions based on actin bundles like epithelial microvilli and inner ear stereocilia, Myo10 is the MyTH4-FERM myosin present in filopodia and most mammalian cells and tissues. This research will answer fundamental questions about Myo10 and filopodia as well as investigating Myo10 functions at the organismal level in health and disease.