All ETDs from UAB

Advisory Committee Chair

Gail Johnson-Voll

Advisory Committee Members

Etty Benveniste

Stuart Frank

Mathieu Lesort

Joanne Murphy-Ullrich

Document Type

Dissertation

Date of Award

2010

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

Low density Lipoprotein receptor Related 6 (LRP6) functions as an essential co-receptor for Wnt/Beat;-catenin signaling as pathway activation, reflected by cytosolic Beta-catenin stabilization and TCF/LEF-1 transactivation, requires Glycogen Synthase Kinase 3Beta(GSK3Beta)-mediated phosphorylation of multiple PPP(S/T)P motifs within the membrane anchored LRP6 intracellular domain. Additionally, LRP6 undergoes a proteolytic cleavage event resulting in the formation of a soluble LRP6 intracellular domain (LRP6-ICD). LRP6-ICD can function within the Wnt/Beta-catenin pathway by interacting with GSK3Beta and attenuating the phosphorylation of GSK3Beta substrates such as cytosolic Beta-catenin as well as stimulate TCF/LEF-1 activity. However, LRP6-ICD's role(s) within the Wnt/Beta-catenin pathway and the mechanism(s) that govern LRP6-ICD activity are not clear. The cytosolic function of LRP6-ICD has been examined, but its possible function in the nucleus remains unexplored. A better understanding of LRP6-ICD biology will improve our understanding of the regulation of the Wnt/Beta-catenin pathway. The objective for the first part of the project was to determine if LRP6-ICD requires GSK3Beta-mediated phosphorylation of its PPP(S/T)P motifs to function within the Wnt/Beta-catenin pathway, similar to LRP6. Results show LRP6-ICD is functionally distinct from LRP6 as LRP6-ICD functions within the pathway in the absence of PPP(S/T)P phosphorylation and functions as a pathway modulatory protein not as a primary activating component. We also showed LRP6-ICD modulates the pathway by functioning as a true GSK3Beta effector protein capable of directly attenuating GSK3Beta kinase activity. In the second part, we analyzed a potential role for LRP6-ICD in the nucleus. Our findings showed LRP6-ICD is a nucleocytoplasmic protein that differentially regulates Wnt/Beta-catenin pathway activity depending upon its localization and protein-protein interaction. We validated the hypothesis that cytosolic LRP6-ICD positively modulates Wnt/Beta-catenin activity through GSK3Beta and enhanced cytosolic Beta-catenin stabilization. Surprisingly, nuclear LRP6-ICD negatively modulates the pathway by interacting with and attenuating Amino-terminal Enhancer of Split (AES), a positive Wnt/Beta-catenin modulatory protein. To summarize, our studies reveal that LRP6-ICD differentially modulates the Wnt/Beta-catenin pathway in a context and localization dependent manner. Our studies also provide further insight concerning the growing role modulatory proteins play in the Wnt/Beta-catenin pathway and mechanistic diversity of the pathway.

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