Characterization of sorting signals in the beta-amyloid precursor protein cytoplasmic domain.
The beta-amyloid precursor protein (APP) is proteolytically processed to generate beta-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 approximately 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.
Lai
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Department of Cancer Biology, Salk Institute, San Diego, California 92186-5800.
Sisodia
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Trowbridge
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D000595:Amino Acid Sequence; D016564:Amyloid beta-Protein Precursor / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002451:Cell Compartmentation; D006224:Cricetinae; D003593:Cytoplasm / Q000378:metabolism; D017931:DNA Primers; D004705:Endocytosis; D006056:Golgi Apparatus / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D021382:Protein Sorting Signals / Q000378:metabolism*; D011990:Receptors, Transferrin / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D016564:Amyloid beta-Protein Precursor; D017931:DNA Primers; D021382:Protein Sorting Signals; D011990:Receptors, Transferrin; D011993:Recombinant Fusion Proteins