Dynein light chain LC8 is a highly conserved, essential component of the microtubule-based molecular motor dynein. Its interactions with a number of non-dynein proteins led to the widely held view that it functions as a cargo adaptor. However, structural and thermodynamics work has recently shown that LC8 cannot serve as a cargo adaptor and has led to the paradigm-shifting proposition that LC8 is not primarily a dynein subunit, but is an essential component of diverse protein complexes that play roles in a variety of cellular systems. The far-reaching and unifying hypothesis of this proposal is that LC8 is a hub protein and functions as a dimerization facilitator in dynein and in all other complexes in which it participates, promoting the dimerization and ordering of the proteins with which it interacts. Work in this proposal will elucidate binding thermodynamics and structural characterization of the two best-known proteins that interact with LC8.
Aim 1 focuses on the interactions of LC8 with the intermediate chain (IC) in dynein assembly and aim 2 focuses on the interactions of LC8 with Swallow, a protein involved in localization of bicoid mRNA during Drosophila development, and the best-studied example of an LC8-interacting protein not associated with dynein. For both aims the mechanism and biological significance of LC8-promoted partner dimerization will be examined by both in vitro biophysical studies of representative protein segments and, when possible, full-length constructs, and in vivo assessment of phenotypes of cells and organisms expressing designed variants of IC and Swallow. These studies will provide a solid foundation for understanding the roles that LC8 plays in the variety of cellular systems in which it participates, and will also promote progress in the LC8 field in general as other LC8 binding partners are physiologically important. These studies also have broader applicability, because LC8 can serve as a model for other hub proteins that interact with a large number of disordered partners. In addition these studies will provide novel insights into the structural biology of dynein motor function, complementing the cell biological approaches that predominate the field of intracellular transport. Dr. Barbar's track record of pioneering work on structure-function relations of LC8, her role as developer of the novel hypothesis that LC8 is a dimerization hub for natively disordered proteins, her success in both producing useful protein constructs and handling these complex and partially disordered proteins, and her expertise in the battery of biophysical and biochemical solution techniques required to probe these questions, make her uniquely suited to lead this effort.

Public Health Relevance

In protein-protein interaction networks, a small proportion of proteins termed """"""""hubs"""""""" interact with many diverse partners. These studies will elucidate the mechanism of interaction of dynein light chain LC8, which we propose is a hub protein, and its role as a dimerization engine in the assembly and regulation of the microtubule-based motor complex dynein, and in assembly of non-dynein complexes that are involved in various physiological processes.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
1R01GM084276-01A2
Application #
7737697
Study Section
Macromolecular Structure and Function B Study Section (MSFB)
Program Officer
Wehrle, Janna P
Project Start
2009-09-30
Project End
2014-08-31
Budget Start
2009-09-30
Budget End
2010-08-31
Support Year
1
Fiscal Year
2009
Total Cost
$292,400
Indirect Cost
Name
Oregon State University
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
053599908
City
Corvallis
State
OR
Country
United States
Zip Code
97339
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Barbar, Elisar; Nyarko, Afua (2014) NMR Characterization of Self-Association Domains Promoted by Interactions with LC8 Hub Protein. Comput Struct Biotechnol J 9:e201402003
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Kidane, Ariam I; Song, Yujuan; Nyarko, Afua et al. (2013) Structural features of LC8-induced self-association of swallow. Biochemistry 52:6011-20

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