title: BACE1 protein endocytosis and trafficking are differentially regulated by ubiquitination at lysine 501 and the Di-leucine motif in the carboxyl terminus.
abstract: β-Site amyloid precursor protein-cleaving enzyme (BACE1) is a membrane-tethered member of the aspartyl proteases that has been identified as β-secretase. BACE1 is targeted through the secretory pathway to the plasma membrane and then is internalized to endosomes. Sorting of membrane proteins to the endosomes and lysosomes is regulated by the interaction of signals present in their carboxyl-terminal fragment with specific trafficking molecules. The BACE1 carboxyl-terminal fragment contains a di-leucine sorting signal ((495)DDISLL(500)) and a ubiquitination site at Lys-501. Here, we report that lack of ubiquitination at Lys-501 (BACE1K501R) does not affect the rate of endocytosis but produces BACE1 stabilization and accumulation of BACE1 in early and late endosomes/lysosomes as well as at the cell membrane. In contrast, the disruption of the di-leucine motif (BACE1LLAA) greatly impairs BACE1 endocytosis and produces a delayed retrograde transport of BACE1 to the trans-Golgi network (TGN) and a delayed delivery of BACE1 to the lysosomes, thus decreasing its degradation. Moreover, the combination of the lack of ubiquitination at Lys-501 and the disruption of the di-leucine motif (BACE1LLAA/KR) produces additive effects on BACE1 stabilization and defective internalization. Finally, BACE1LLAA/KR accumulates in the TGN, while its levels are decreased in EEA1-positive compartments indicating that both ubiquitination at Lys-501 and the di-leucine motif are necessary for the trafficking of BACE1 from the TGN to early endosomes. Our studies have elucidated a differential role for the di-leucine motif and ubiquitination at Lys-501 in BACE1 endocytosis, trafficking, and degradation and suggest the involvement of multiple adaptor molecules.
authors.affiliation: Alzheimer's Disease Research Laboratory, Department of Neuroscience, Tufts University School of Medicine, Boston, Massachusetts 02111, USA.
authors.forename: Eugene L
authors.initials: EL
authors.lastname: Kang
authors.forename: Barbara
authors.initials: B
authors.lastname: Biscaro
authors.forename: Fabrizio
authors.initials: F
authors.lastname: Piazza
authors.forename: Giuseppina
authors.initials: G
authors.lastname: Tesco
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D053829:Amyloid Precursor Protein Secretases / Q000737:chemistry* / Q000378:metabolism*; D016564:Amyloid beta-Protein Precursor / Q000378:metabolism; D016282:Aspartic Acid Endopeptidases / Q000737:chemistry* / Q000378:metabolism*; D004705:Endocytosis; D011992:Endosomes / Q000201:enzymology; D004795:Enzyme Stability; D006801:Humans; D007930:Leucine / Q000378:metabolism*; D008239:Lysine / Q000378:metabolism*; D008247:Lysosomes / Q000201:enzymology; D008954:Models, Biological; D008969:Molecular Sequence Data; D011499:Protein Processing, Post-Translational; D021381:Protein Transport; D013347:Subcellular Fractions / Q000201:enzymology; D054875:Ubiquitination; D021601:trans-Golgi Network / Q000201:enzymology
chemical_list: D016564:Amyloid beta-Protein Precursor; D053829:Amyloid Precursor Protein Secretases; D016282:Aspartic Acid Endopeptidases; C509280:BACE1 protein, human; D007930:Leucine; D008239:Lysine