A key player in mast cell activation is the high affinity IgE receptor, Fc5RI. While much is known about the chain of events initiated by crosslinking of Fc?RI through multivalent antigen, there is a fundamental gap in our understanding of how protein dynamics facilitate signaling. Our long term goal is to understand how the dynamic and stochastic behavior of protein-protein interactions influences signal propagation. The objective of this proposal is to quantify dynamic protein interactions during early events in Fc?RI signaling. Our central hypothesis is that the duration of protein-protein interactions modulates the signaling outcome. The rationale for the proposed research is that understanding the role of protein-protein interaction dynamics in signaling is the next step in understanding how the cell shapes the strength and quality of an immune response. We will integrate multiple imaging modalities, both novel and established, to capture and quantify early events that govern Fc?RI membrane associated signaling in order to achieve three specific aims: 1) To determine the interaction lifetime and diffusional dynamics that facilitate Fc?RI activation;2) To quantify the kinetics of Syk adaptor protein binding to the signaling complex;and 3) To determine the cluster stability of LAT and the interplay between Fc?RI and LAT signaling patches. The approach is innovative because we will develop methodology for imaging biochemical events at the molecular level that will allow us to obtain dynamic information about signaling events that cannot be determined using traditional biochemistry techniques. The proposed research is significant because the quantitative information that we propose to obtain has not been directly measured before and will bring new perspectives to the cell biology community. Furthermore, the development of multi-color single molecule imaging techniques described here will be applicable to many other fundamental biological questions. The information we gain about Fc5RI signaling will help to fill the gaps in our knowledge of how Fc5RI initiates signaling. Ultimately, we expect that this information will open new avenues for drug design that target protein interactions and localization.

Public Health Relevance

This proposal is relevant to public health since the activation of mast cells is implicated in many physiological responses from allergy and asthma to rheumatoid arthritis and cancer. In order to further our fundamental understanding of mast cell signaling, we will quantify protein interactions and dynamics at the molecular level using innovative imaging techniques. Therefore, the proposal is relevant to the part of NIH's mission that is in pursuit of fostering fundamental creative discoveries, innovative research strategies, and their applications to biomedical problems.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM100114-02
Application #
8463220
Study Section
Enabling Bioanalytical and Imaging Technologies Study Section (EBIT)
Program Officer
Lewis, Catherine D
Project Start
2012-05-01
Project End
2017-02-28
Budget Start
2013-03-01
Budget End
2014-02-28
Support Year
2
Fiscal Year
2013
Total Cost
$275,280
Indirect Cost
$84,210
Name
University of New Mexico Health Sciences Center
Department
Pathology
Type
Schools of Medicine
DUNS #
829868723
City
Albuquerque
State
NM
Country
United States
Zip Code
87131
Schwartz, Samantha L; Yan, Qi; Telmer, Cheryl A et al. (2015) Fluorogen-activating proteins provide tunable labeling densities for tracking Fc?RI independent of IgE. ACS Chem Biol 10:539-46
Yan, Qi; Schwartz, Samantha L; Maji, Suvrajit et al. (2014) Localization microscopy using noncovalent fluorogen activation by genetically encoded fluorogen-activating proteins. Chemphyschem 15:687-95
Cutler, Patrick J; Malik, Michael D; Liu, Sheng et al. (2013) Multi-color quantum dot tracking using a high-speed hyperspectral line-scanning microscope. PLoS One 8:e64320