Organelle movement through cytoplasm, which occurs in virtually all eukaryotic cells, plays an important role in transporting lipids and proteins between regions or compartments of the cell. Organelle transport was described ober a century ago, yet the mechanism responsible for generating this movement has remained a mystery, although it is now clear that microtubules and microtubule-based motility proteins are involved in this process. Using neurons as model systems for examining this important phenomenon, my research approach is to reconstitute microtubule- based transport in vitro using isolated components that can then be biochemically analyzed. The overall goals if this proposal are to identify the molecules that comprise the force generating machinery that powers organelle transport, to determine how these proteins generate movement and to understand how motility is regulated.
The specific aims are: 1) to study the biochemistry and in vitro motility of a newly described force generating protein termed kinesin, distinct from myosin and dynein, that appears to mediate anterograde movement of organelles in axons. Research will focus upon investigating the molecular interaction of kinesin with microtubules and nucleotides, the mechanism and regulation of force production, the association of kinesin with organelles, and the characterization of functional domains of kinesin. 2) to identify, purify and characterize a second motility protein in crude axoplasmic supernatants that generates movement along microtubules in the opposite direction to kinesin. This protein may be involved in the retrograde transport of organelles. 3) to ascertain how kinesin and the retrograde motor are involved in transporting organelles by reconstituting movement using purified proteins and organelles and to identify signals that control the direction of transport of organelles along microtubules. The in vitro motility assays, the ability to prepare sufficient quantities of kinesin for biochemical characterization and the availability of specific antibodies against kinesin have opened up new possibilities for understanding organelle transport and other forms of microtubule-based motility at a molecular level. The studies proposed here address fundamental problems in modern cell biology and contribute to an understanding of transport processes that are medically relevant such as hormone secretion and nerve regeneration.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM038499-05
Application #
3294937
Study Section
Cellular Biology and Physiology Subcommittee 1 (CBY)
Project Start
1988-02-01
Project End
1993-01-31
Budget Start
1992-02-01
Budget End
1993-01-31
Support Year
5
Fiscal Year
1992
Total Cost
Indirect Cost
Name
University of California San Francisco
Department
Type
Schools of Medicine
DUNS #
073133571
City
San Francisco
State
CA
Country
United States
Zip Code
94143
Jain, Ankur; Vale, Ronald D (2017) RNA phase transitions in repeat expansion disorders. Nature 546:243-247
Hui, Enfu; Cheung, Jeanne; Zhu, Jing et al. (2017) T cell costimulatory receptor CD28 is a primary target for PD-1-mediated inhibition. Science 355:1428-1433
Ori-McKenney, Kassandra M; McKenney, Richard J; Huang, Hector H et al. (2016) Phosphorylation of ?-Tubulin by the Down Syndrome Kinase, Minibrain/DYRK1a, Regulates Microtubule Dynamics and Dendrite Morphogenesis. Neuron 90:551-63
Yan, Xiaowei; Hoek, Tim A; Vale, Ronald D et al. (2016) Dynamics of Translation of Single mRNA Molecules In Vivo. Cell 165:976-89
Jonsson, Erik; Yamada, Moé; Vale, Ronald D et al. (2015) Clustering of a kinesin-14 motor enables processive retrograde microtubule-based transport in plants. Nat Plants 1:
Tanenbaum, Marvin E; Gilbert, Luke A; Qi, Lei S et al. (2014) A protein-tagging system for signal amplification in gene expression and fluorescence imaging. Cell 159:635-46
Smith, Benjamin A; Gelles, Jeff; Goode, Bruce L (2014) Single-molecule studies of actin assembly and disassembly factors. Methods Enzymol 540:95-117
Vale, Ronald D (2014) Preface: the role of reconstitution in cytoskeleton research. Methods Enzymol 540:xix-xxiii
Schroeder, Courtney M; Ostrem, Jonathan M L; Hertz, Nicholas T et al. (2014) A Ras-like domain in the light intermediate chain bridges the dynein motor to a cargo-binding region. Elife 3:e03351
Petry, Sabine; Groen, Aaron C; Ishihara, Keisuke et al. (2013) Branching microtubule nucleation in Xenopus egg extracts mediated by augmin and TPX2. Cell 152:768-77

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