The microtubule and actin cytoskeletons are essential for the movement, polarization, sorting, and morphogenesis of intracellular membrane compartments. Molecular motors, scaffolding proteins, and associated filaments are recruited to membranes to power diverse trafficking events that have different force, power, kinetic, and transport requirements. Defining the mechanisms of trafficking requires understanding how different motor isoforms and families work alone and in teams, and how motors and filaments work with the scaffolding proteins and adapters that link them to membranes. Determining these mechanisms requires a detailed understanding of: the cellular organization and dynamics of the cytoskeleton and membranes;the spatial, kinetic, and mechanical relationship of different motors and filaments;the structural and biophysical properties of cytoskeletal-membrane assemblies;and the biophysical parameters that define the capabilities and mechanisms of motors and scaffolds when operating under working conditions. To this end, we assembled an extraordinarily strong scientific team with expertise in cell biology, biochemistry, structural biology, structural dynamics, and technology-development to define the role of the cytoskeleton and molecular motors in trafficking. Our team includes pioneers in the use of state-of-the-art imaging, single-molecule, and structural techniques to discover how cytoskeletal proteins function in complex cellular events. High-resolution live-cell microscopy, reconstituted cytoskeletal geometries using microfabrication and dielectrophoresis, X-ray crystallography, nanometer-resolved fluorescence tracking, single-molecule fluorescence polarization, optical trapping, and advanced biochemical techniques will be applied in highly collaborative studies to understand how motors, scaffolds, and filaments work together to power membrane dynamics. The projects and investigators are interdependent and are closely linked through research goals and common technologies, and the Aims were formulated to capitalize on the unique strengths of the team members while taking advantage of extensive synergies between the groups. We will focus on the following four Aims: (1) Investigate the Dynamics of Molecular Motors in Organelle Transport and Membrane Remodeling;(2) Investigate the Structural, Biochemical, and Cellular Properties of Cytoskeleton-Membrane Scaffolds in Organelle Morphogenesis and Motility;(3) Discover the Mechanical and Biochemical Adaptations of Membrane- Associated Motors and Scaffolds;(4) Investigate the Structural Dynamics of Myosin, Dynein and Motor Collections.

Public Health Relevance

Cytoskeletal motors and filaments are crucial for several normal and pathological processes, including: cell and tissue development, endocytosis, wound healing, hearing, and cell movement. However, the molecular details of motors in many of these functions are unknown. Therefore, we will define the cellular, biochemical, and biophysical properties of these motors to better understand the molecular basis of cell physiology and pathology of health-care problems such as cancer, neuronal defects, and cardiovascular disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Program Projects (P01)
Project #
2P01GM087253-11
Application #
8742368
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Gindhart, Joseph G
Project Start
2004-04-26
Project End
2019-07-31
Budget Start
2014-09-01
Budget End
2015-07-31
Support Year
11
Fiscal Year
2014
Total Cost
Indirect Cost
Name
University of Pennsylvania
Department
Physiology
Type
Schools of Medicine
DUNS #
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
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Moore, Andrew S; Holzbaur, Erika L F (2018) Mitochondrial-cytoskeletal interactions: dynamic associations that facilitate network function and remodeling. Curr Opin Physiol 3:94-100
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Lee, In-Gyun; Olenick, Mara A; Boczkowska, Malgorzata et al. (2018) A conserved interaction of the dynein light intermediate chain with dynein-dynactin effectors necessary for processivity. Nat Commun 9:986
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Lippert, Lisa G; Dadosh, Tali; Hadden, Jodi A et al. (2017) Angular measurements of the dynein ring reveal a stepping mechanism dependent on a flexible stalk. Proc Natl Acad Sci U S A 114:E4564-E4573
Pyrpassopoulos, Serapion; Shuman, Henry; Ostap, E Michael (2017) Adhesion force and attachment lifetime of the KIF16B-PX domain interaction with lipid membranes. Mol Biol Cell 28:3315-3322
Lewis, John H; Jamiolkowski, Ryan M; Woody, Michael S et al. (2017) Deconvolution of Camera Instrument Response Functions. Biophys J 112:1214-1220
Greenberg, Michael J; Shuman, Henry; Ostap, E Michael (2017) Measuring the Kinetic and Mechanical Properties of Non-processive Myosins Using Optical Tweezers. Methods Mol Biol 1486:483-509

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