C-type lectins constitute a key class of pathogen receptors on dendritic cells (DCs), one of the main antigen presenting cell types in the immune system. One such receptor, DC-SIGN, binds to a large variety of pathogens, including dengue virus, HIV and yeasts, by recognizing carbohydrates on the pathogen surfaces. DC-SIGN is found assembled in small domains on the plasma membranes of DCs, and this clustering is required for DC-SIGN to successfully bind to and internalize pathogens. These domains possess some striking properties. First, the DC-SIGN molecules appear to be immobilized within the domains. Second, the origin of this immobilization is initiated in the extracellular carbohydrate recognition moiety rather than the membrane apposed cytoskeleton. Third, super-resolution microscopy shows that the microdomains, as imaged by conventional fluorescence microscopy, are actually composed of arrays of ~75nm nanodomains containing only about 3 tetramers of DC-SIGN, which indicates the presence of other proteins and lipids within these "nanodomains". To elucidate how these nanodomains carry out their function in DCs, we will define their nanoscale properties after pathogens are bound by using super- resolution microscopies and, furthermore, investigate the mechanisms of transport from binding to internalization sites. We will also define the at present unknown nanodomain protein composition using modern proteomic technologies complemented by state of the art domain isolation methods. Information on this level of detail will not only provide an increased understanding of membrane domains in general, but will also provide a much more complete picture of the initial recognition and processing of infectious agents by the human immune system.

Public Health Relevance

The studies to be conducted will provide a much more detailed understanding of the manner in which previously identified molecular clusters found on the surfaces of certain human cells belonging to the immune system initially recognize and subsequently process (for further immune response) a very large variety of human pathogens. The range of infectious agents recognized and processed by these cell-surface molecular clusters is quite broad, including a number of bacteria (e.g., E. Coli, H. pylori), yeasts (e.g, Candida albicans), viruses (e.g., HIV, Ebola &Dengue), and parasites (e.g., Leishmania). Greatly increased understanding of the mechanism through which these molecular clusters function will thus provide a variety of new opportunities for therapeutic interventions aimed at suppressing and/or curing a large variety of human diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
3R01GM041402-23S1
Application #
8928279
Study Section
Biochemistry and Biophysics of Membranes Study Section (BBM)
Program Officer
Chin, Jean
Project Start
1988-12-01
Project End
2017-07-31
Budget Start
2014-08-01
Budget End
2015-07-31
Support Year
23
Fiscal Year
2014
Total Cost
$48,953
Indirect Cost
Name
University of North Carolina Chapel Hill
Department
Physiology
Type
Schools of Medicine
DUNS #
608195277
City
Chapel Hill
State
NC
Country
United States
Zip Code
27599
Itano, Michelle S; Graus, Matthew S; Pehlke, Carolyn et al. (2014) Super-resolution imaging of C-type lectin spatial rearrangement within the dendritic cell plasma membrane at fungal microbe contact sites. Front Phys 2:
Liu, Ping; Wang, Xiang; Itano, Michelle S et al. (2014) Low copy numbers of DC-SIGN in cell membrane microdomains: implications for structure and function. Traffic 15:179-96
Thompson, Nancy L; Navaratnarajah, Punya; Wang, Xiang (2011) Measuring surface binding thermodynamics and kinetics by using total internal reflection with fluorescence correlation spectroscopy: practical considerations. J Phys Chem B 115:120-31
Neumann, Aaron K; Itano, Michelle S; Jacobson, Ken (2010) Understanding lipid rafts and other related membrane domains. F1000 Biol Rep 2:31
Neumann, Aaron K; Jacobson, Ken (2010) A novel pseudopodial component of the dendritic cell anti-fungal response: the fungipod. PLoS Pathog 6:e1000760
Thompson, Nancy L; Wang, Xiang; Navaratnarajah, Punya (2009) Total internal reflection with fluorescence correlation spectroscopy: Applications to substrate-supported planar membranes. J Struct Biol 168:95-106
Neumann, Aaron K; Thompson, Nancy L; Jacobson, Ken (2008) Distribution and lateral mobility of DC-SIGN on immature dendritic cells--implications for pathogen uptake. J Cell Sci 121:634-43
Lagerholm, B Christoffer (2007) Peptide-mediated intracellular delivery of quantum dots. Methods Mol Biol 374:105-12