Sabo SL, Ikin AF, Buxbaum JD, Greengard P. The Alzheimer amyloid precursor protein (APP) and FE65, an APP-binding protein, regulate cell movement. J Cell Biol. 2001 Jun 25;153(7):1403-14. PubMed Abstract

REAGENTS/MATERIAL:
H4 neuroglioma cells and MDCK cells maintained in DME supplemented medium. MDCK cells overexpressing the 695 amino acid isoform of APP (+APP/-FE65) were a gift of Dr. C. Haass (Ludwig Maximilians University, Munich, Germany). These cells were stably transfected with FE65 cDNA according to Sabo et al. Polyclonal FE65 antibodies 170/173, which recognize the WW domain of FE65, were affinity purified accorindg to Sabo et al. Polyclonal APP antibody 369, which recognizes the cytoplasmic domain of APP, was affinity purified with a peptide corresponding to the last 50 amino acids of APP. APP monoclonal antibodies 5A3/1G7, recognize epitopes in the extracellular domain of APP, gift from Dr. E.H. Koo, Univeristy of California at San Diego. Polyclonal antibodies against Mena, gift from Dr. F. Gertler (MIT, Cambridge, MA) Integrin b1 and phosphotyrosine monoclonal antibodies were purchased from Upstate Biotechnology Secondary antibodies 1:400 for Oregon green goat anti-mouse and Texas red goat anti-rabbit (Molecular Probes), 1:200 for Cy5 goat anti-mouse, and 1:10 for rhodamine red X goat anti-rabbit (Jackson ImmunoResearch Laboratories).

FUTURE DIRECTION:
Further studies needed in identifying the sginals that regulate the interaction between FE65 and Mena or FE65 and APP.