This proposal is focused on developing our understanding of class 1 myosins: ubiquitously expressed monomeric, membrane binding, actin-based motors that participate in diverse cellular functions, including organelle trafficking, transcription, host defense, cell motility, and mechano-sensation. Our studies of myosin-1 are centered on the 'brush border', a tightly packed array of microvilli that extends from the apical surface of many transporting epithelial cells types. This organelle is home to a number of myosin superfamily members, with the most abundant being myosin-1a (Myo1a), one of eight vertebrate class 1 myosins. Our laboratory has leveraged a unique combination of cell biological and biophysical approaches to discover that: (i) Myo1a contributes to membrane-cytoskeleton adhesion, which is critical for maintaining normal brush border structure, and (ii) Myo1a powers the release of membrane vesicles enriched in host defense machinery from microvillar tips into the intestinal lumen. The physiological significance of Myo1a function is also underscored by recent studies showing that mutations in this motor are linked to colorectal tumor formation in humans. While we have made substantial progress toward elucidating the biological roles of Myo1a, the molecular properties, interactions, and events that govern the function of this and other myosins-1 remain poorly characterized. The goal of this proposal is to develop our understanding of the fundamental biochemical and biophysical properties that enable Myo1a to contribute to brush border function. To this end, Aim 1 will examine the unitary properties of single Myo1a molecules interacting with the plasma membrane of live cells and supported bilayers in vitro, Aim 2 will examine the force generating potential of Myo1a bound to supported bilayers, and Aim 3 will investigate the role of force sensing in the regulation of Myo1a dynamics and function. Because defects in brush border formation and maintenance are at the core of numerous diseases that pose significant threats to human health, developing insight on the molecular mechanisms that govern Myo1a behavior will provide information that may ultimately be used in the development of therapeutics aimed at repairing malformed or damaged brush borders.

Public Health Relevance

Brush border microvilli function as the major site of nutrient absorption and a barrier to bacteria that reside in the intestinal lumen. Functional microvilli are essential for maintaining normal human health;malformation of the brush border or microvillar destruction caused by disease or infection are associated with malabsorption, dehydration, and in severe cases, death. Understanding the mechanism of molecules such as Myo1a, which are important for microvillar maintenance and function, will be critical for understanding human illnesses that derive from brush border malformation, enteric infections, and intestinal cancers.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK075555-07
Application #
8596813
Study Section
Macromolecular Structure and Function C Study Section (MSFC)
Program Officer
Grey, Michael J
Project Start
2006-07-01
Project End
2016-12-31
Budget Start
2014-01-01
Budget End
2014-12-31
Support Year
7
Fiscal Year
2014
Total Cost
$294,497
Indirect Cost
$105,422
Name
Vanderbilt University Medical Center
Department
Anatomy/Cell Biology
Type
Schools of Medicine
DUNS #
004413456
City
Nashville
State
TN
Country
United States
Zip Code
37212
Shifrin Jr, David A; Crawley, Scott W; Grega-Larson, Nathan E et al. (2014) Dynamics of brush border remodeling induced by enteropathogenic E. coli. Gut Microbes 5:504-16
GĂ©rard, Audrey; Patino-Lopez, Genaro; Beemiller, Peter et al. (2014) Detection of rare antigen-presenting cells through T cell-intrinsic meandering motility, mediated by Myo1g. Cell 158:492-505
Crawley, Scott W; Shifrin Jr, David A; Grega-Larson, Nathan E et al. (2014) Intestinal brush border assembly driven by protocadherin-based intermicrovillar adhesion. Cell 157:433-46
Knowles, Byron C; Tyska, Matthew J; Goldenring, James R (2014) Apical vesicle trafficking takes center stage in neonatal enteropathies. Gastroenterology 147:15-7
Mazerik, Jessica N; Kraft, Lewis J; Kenworthy, Anne K et al. (2014) Motor and tail homology 1 (Th1) domains antagonistically control myosin-1 dynamics. Biophys J 106:649-58
Knowles, Byron C; Roland, Joseph T; Krishnan, Moorthy et al. (2014) Myosin Vb uncoupling from RAB8A and RAB11A elicits microvillus inclusion disease. J Clin Invest 124:2947-62
Shifrin Jr, David A; Demory Beckler, Michelle; Coffey, Robert J et al. (2013) Extracellular vesicles: communication, coercion, and conditioning. Mol Biol Cell 24:1253-9
Shifrin Jr, David A; McConnell, Russell E; Nambiar, Rajalakshmi et al. (2012) Enterocyte microvillus-derived vesicles detoxify bacterial products and regulate epithelial-microbial interactions. Curr Biol 22:627-31
Mazerik, Jessica N; Tyska, Matthew J (2012) Myosin-1A targets to microvilli using multiple membrane binding motifs in the tail homology 1 (TH1) domain. J Biol Chem 287:13104-15
Liu, Yin; Belkina, Natalya V; Park, Chung et al. (2012) Constitutively active ezrin increases membrane tension, slows migration, and impedes endothelial transmigration of lymphocytes in vivo in mice. Blood 119:445-53

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