Direct visualization of the gamma secretase-generated carboxyl-terminal domain of the amyloid precursor protein: association with Fe65 and translocation to the nucleus |
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Authors: | Kinoshita A Whelan C M Smith C J Berezovska O Hyman B T |
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Institution: | Alzheimer Disease Research Laboratory, Massachusetts General Hospital, Harvard Medical School, Charlestown, Massachusetts, USA. |
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Abstract: | Amyloid-beta, the peptide that deposits as senile plaques in Alzheimer's disease, is derived from the amyloid precursor protein (APP) by a gamma secretase-mediated intramembranous cleavage. In addition to amyloid-beta, this cleavage produces a carboxyl-terminal intracellular fragment which has an unknown function. The carboxyl-terminal domain of APP interacts in the cytoplasm with an adapter protein, Fe65. We demonstrate by laser scanning confocal microscopy that a gamma secretase generated APP carboxyl-terminal domain, tagged with green fluorescent protein (GFP), translocates to the nucleus in a manner dependent upon stabilization by the adapter protein Fe65; APP which has been mutated to block interactions with Fe65 cannot be detected in the nucleus. The APP-CT domain continues to interact with Fe65 in the nucleus, as determined by both colocalization and fluorescence resonance energy transfer (FRET). Visualization of the APP-CT-Fe65 complex in the nucleus may serve as a readout for processes that modify gamma secretase release of APP-CT. |
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Keywords: | adapter protein amyloid precursor protein Fe65 FRET gamma cleavage intracellular domain |
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