The Basic Biology of Aging at the University of Washington
Zheng Yang

Email:yangz@u.washington.edu

In the past year, one of Dr. Yang’s research interests was on searching ligand for APP, a type I transmembrane protein that generates beta-amyloid by proteolytic processing. Purified APP ectodomain was used to cross-link with cell membranes (either from cell culture or mouse tissue). But this method failed to produce a consistent result. Right now this project has been putting on hold until a more sensitive method is found.

A second project that Dr. Yang was working on was studying the functional difference between neuronal and nonneuronal forms of FE65. FE65 is an adaptor protein that interacts with APP C-terminal and is believe to play a role in regulating APP processing and function. Alternative splicing of the FE65 gene produces two isoforms of FE65 that are expressed in either neuronal or nonneuronal tissues separately. Neuronal form contains a potential tyrosine phosphorylation site. Immunoprecipitation and western blot was used to see whether endogenous FE65 got phosphorylated at tyrosine residue (using mouse brain tissue). The result showed that endogenous neuronal FE65 is not tyrosine phosphorylated. Dr. Yang also tried to see whether different isoforms localize in different cellular compartments by immunostaining of the MDCK cells. The result showed that they all located in nucleus of the cell. Currently Dr. Yang is focusing on testing whether two isoforms have different affinity for LRP and Tip60, which bind to FE65 in the alternatively spliced region.

The third project that Dr. Yang is still working on is to study the role of FE65 in the APP-FE65 –Tip60 transcription activation complex. Some evidence from our lab showed that APP might function to modulate the localization of FE65. By fusing FE65 to GAL4 DNA binding domain, we want to test the possibility of FE65 –gal4 can turn on transcription and APP plays regulatory role in the complex. Currently Dr. Yang is in the process of making constructs.