The Wnt family of secretory glycoproteins plays important roles in a wide range of biological and pathophysiological processes, including embryonic development, organogenesis, tissue homeostasis, stem cell biology, lipid and glucose metabolism and tumor genesis. In this study we plan to work on one of the major gaps that remains in Wnt signaling mechanisms, which is how Wnt transduces the signals cross the plasma membrane. Wnt3a, a prototypical canonical Wnt, initiates its signaling by binding to its two cell surface receptor Frizzled (Fz) and LDL- related protein 5/6 (LRP), leading to the phosphorylation of LRP and subsequently stabilization of b-catenin. We previously showed that Wnt3a, via Fz and disheveled (Dvl), stimulates phosphatidylinositol (PI) kinases PI4K2 and PIP5K1 to produce phosphatidylinositol 4, 5- bisphosphate (PIP2). PIP2 facilitates the formation of LRP clusters or signalosomes on the cell surface, which is important for LRP phosphorylation and signaling downstream. We used super resolution fluorescence imaging techniques including 3D-STORM (STochastic Optical Reconstruction Microscopy) to directly visualized LRP6 and clathrin clusters and their co-localization at cell surfaces, and in this study we will extend our study to Fz and Dvl. In addition, we plan to characterize the molecular basis for the interactions of PI4K2 and PIP5K1 with Dvl and mechanisms for the activation of these PI kinases by Dvl.
The specific aims of this study are: 1) to further characterize the formation and organization of Wnt3a receptor signalosomes using both biochemical and advanced imaging approaches. 2) To elucidate the molecular mechanisms for Dvl interaction and activation of PI4K2 and PIP5K1. These studies will provide novel organizational and mechanistic insights into the formation of Wnt3a receptor signalosome and elucidate how Dvl, a protein previously only known as a scaffold, stimulates lipid kinase activity.

Public Health Relevance

The purpose is to study the mechanisms for Wnt signaling, a process by which cells communicate with each other. Defects in these types of processes contribute to human diseases including cancer and diabetes. Our study will thus help us to understand these basic cellular functions and related diseases and may uncover therapeutic solutions to the diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
9R01GM112182-06
Application #
8761217
Study Section
Intercellular Interactions (ICI)
Program Officer
Dunsmore, Sarah
Project Start
2009-04-01
Project End
2018-05-31
Budget Start
2014-09-05
Budget End
2015-05-31
Support Year
6
Fiscal Year
2014
Total Cost
$366,300
Indirect Cost
$146,300
Name
Yale University
Department
Pharmacology
Type
Schools of Medicine
DUNS #
043207562
City
New Haven
State
CT
Country
United States
Zip Code
06520