Characterization of Sorting Signals in the 
-Amyloid Precursor Protein Cytoplasmic Domain (*)
- From the (1)Department of Cancer Biology, The Salk Institute, San Diego, California 92186-5800, the
- (2)Biomedical Sciences Graduate Program, University of California at San Diego, La Jolla, California 92037, and the
- (3)Neuropathology Laboratory, Department of Pathology, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205
- § To whom correspondence should be sent. Tel.: 619-453-4100 (ext. 1241); Fax: 619-455-1349.
Abstract
The β-amyloid precursor protein (APP) is proteolytically processed to generate β-amyloid protein, the principal protein component
of neuropathological lesions characteristic of Alzheimer's disease. To investigate potential sorting signals in the cytoplasmic
tail of APP, we transplanted APP cytoplasmic tail sequences into the cytoplasmic tail of the human transferrin receptor (TR)
and showed that two sequence motifs from the APP cytoplasmic tail promote TR internalization. One sequence, GYENPTY, is related
to the low density lipoprotein receptor internalization signal, FDNPVY, but also involves a critical glycine residue; the
other, YTSI, conforms to the 4-residue tyrosine-based internalization signal consensus sequence. Furthermore, a chimeric molecule
(APP-TR) consisting of the cytoplasmic domain of APP and the transmembrane and external domains of TR was rapidly internalized
enabling the transport of iron into the cell at 
50% the rate of wild-type TR. Alanine scanning mutations indicated that the two sequences identified in transplantation experiments
were required for internalization of the chimera. Metabolic pulse-chase experiments showed that the APP-TR chimeras were degraded
in a post-Golgi membrane compartment within 2-4 h following normal glycosylation. Degradation was partially dependent upon
the two internalization signals and was inhibited by ammonium chloride. A fraction of APP-TR chimeras traffic to a degradative
endocytic compartment after appearing on the cell surface. Comparison of soluble APP released from cells expressing either
full-length human APP or mutant APP with the sequence YENPTY deleted indicated that this sequence is required for sorting
of full-length APP along similar trafficking pathways as the APP-TR chimera.
Footnotes
-
↵* This work was supported by National Institutes of Health Grant GM07198 and the Lucille P. Markey Charitable Trust (to A. L.), by National Institutes of Health Grants NSAG05146 and NS20471 and the American Health Assistance Foundation (to S. S. S.), and by National Institutes of Health Grant RO1-CA34787 (to I. S. T.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
-
↵1 The abbreviations used are:
- AD
-
Alzheimer's disease
- APP
-
human β-amyloid precursor protein
- Aβ
-
β-amyloid protein
- APPs
-
soluble APP fragment
- TR
-
human transferrin receptor
- Tf
-
human transferrin
- LDLR
-
low density lipoprotein receptor
- CEF
-
chick embryo fibroblasts
- CHO
-
Chinese hamster ovary
- DME
-
Dulbecco's modified Eagle's medium
- BSA
-
bovine serum albumin
- PBS
-
phosphate-buffered saline
- PAGE
-
polyacrylamide gel electrophoresis.
-
- Received September 27, 1994.
- Revision received November 14, 1994.
- © 1995 by The American Society for Biochemistry and Molecular Biology, Inc.
