A Cytoplasmic Sequence in Human Tyrosinase Defines a Second Class of Di-leucine-based Sorting Signals for Late Endosomal and Lysosomal Delivery*
- From the Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104-6082
Abstract
Distinct cytoplasmic sorting signals target integral membrane proteins to late endosomal compartments, but it is not known whether different signals direct targeting by different pathways. The availability of multiple pathways may permit some cell types to divert proteins to specialized compartments, such as the melanosome of pigmented cells. To address this issue, we characterized sorting determinants of tyrosinase, a tissue-specific resident protein of the melanosome. The cytoplasmic domain of tyrosinase was both necessary and sufficient for internalization and steady state localization to late endosomes and lysosomes in HeLa cells. Mutagenesis of two leucine residues within a conventional di-leucine motif ablated late endosomal localization. However, the properties of this di-leucine-based signal were distinguished from that of CD3γ by overexpression studies; overexpression of the tyrosinase signal, but not the well characterized CD3γ signal, induced a 4-fold enlargement of late endosomes and lysosomes and interfered with endosomal sorting mediated by both tyrosine- and other di-leucine-based signals. These properties suggest that the tyrosinase and CD3γ di-leucine signals are distinctly recognized and sorted by distinct pathways to late endosomes in non-pigmented cells. We speculate that melanocytic cells utilize the second pathway to divert proteins to the melanosome.
Footnotes
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↵* This work was supported in part by Grant RPG 97-003-01-BE from the American Cancer Society and Grants R21 AI 42055 and R21 AI 42617 from the National Institutes of Health.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵‡ Supported in part by a National Research Service Award 1F32-AI09946 and by Training Grant T32-CA09671 from the National Institutes of Health.
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↵§ Supported in part by Training Grant T32-EY07131-06 from the National Institutes of Health.
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↵¶ To whom correspondence should be addressed: Dept. of Pathology and Laboratory Medicine, University of Pennsylvania School of Medicine, 277 John Morgan Bldg./6082, 36th and Hamilton Walk, Philadelphia, PA 19104-6082. Tel.: 215-898-3204; Fax: 215-573-4345; E-mail:marksm{at}mail.med.upenn.edu.
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↵2 J. F. Berson, D. W. Frank, P. A. Calvo, B. M. Bieler, and M. S. Marks, manuscript in preparation.
- Abbreviations:
- OCA
-
oculocutaneous albinism
- Ø
-
amino acid with bulky hydrophobic side chain
- LL
-
di-leucine
- YXXØ
-
tyrosine-based motif
- AP
-
adaptor-like complex
- TRP
-
tyrosinase-related protein
- mAb
-
monoclonal antibody
- IFM
-
immunofluorescence microscopy
- CHX
-
cycloheximide
- FITC
-
fluorescein isothiocyanate
- LRSC
-
lissamine rhodamine-sulfonyl chloride
- PE
-
phycoerythrin
- PBS
-
phosphate-buffered saline
- PCR
-
polymerase chain reaction
- ER
-
endoplasmic reticulum
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- Received October 27, 1998.
- Revision received January 5, 1999.
- The American Society for Biochemistry and Molecular Biology, Inc.
