Transport of newly synthesized lysosomal membrane proteins from the TGN (trans-Golgi network) to the lysosomes is due to the presence of specific signals in their cytoplasmic domains that are recognized by cytosolic adaptors. p40, a hypothetical transporter of 372 amino acids localized in the lysosomal membrane, contains four putative lysosomal sorting motifs in its sequence: three of the YXXϕ-type (Y6QLF, Y106VAL, Y333NGL) and one of the [D/E]XXXL[L/I]-type (EQERL360L361). To test the role of these motifs in the biosynthetic transport of p40, we replaced the most critical residues of these consensus sequences, the tyrosine residue or the leucine–leucine pair, by alanine or alanine–valine respectively. We analysed the subcellular localization of the mutated p40 proteins in transfected HeLa cells by confocal microscopy and by biochemical approaches (subcellular fractionation on self-forming Percoll density gradients and cell surface biotinylation). The results of the present study show that p40 is mistargeted to the plasma membrane when its dileucine motif is disrupted. No role of the tyrosine motifs could be put forward. Taken together, our results provide evidence that the sorting of p40 from the TGN to the lysosomes is directed by the dileucine EQERL360L361 motif situated in its C-terminal tail.
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Research Article|
August 27 2008
A dileucine signal situated in the C-terminal tail of the lysosomal membrane protein p40 is responsible for its targeting to lysosomes
Marielle Boonen;
Marielle Boonen
1URΦM, Laboratoire de Chimie Physiologique, FUNDP, B-5000 Namur, Belgium
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Roberta Rezende de Castro;
Roberta Rezende de Castro
1URΦM, Laboratoire de Chimie Physiologique, FUNDP, B-5000 Namur, Belgium
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Gaëlle Cuvelier;
Gaëlle Cuvelier
1URΦM, Laboratoire de Chimie Physiologique, FUNDP, B-5000 Namur, Belgium
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Isabelle Hamer;
Isabelle Hamer
1URΦM, Laboratoire de Chimie Physiologique, FUNDP, B-5000 Namur, Belgium
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Michel Jadot
Michel Jadot
1
1URΦM, Laboratoire de Chimie Physiologique, FUNDP, B-5000 Namur, Belgium
1To whom correspondence should be addressed (email michel.jadot@fundp.ac.be).
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Publisher: Portland Press Ltd
Received:
November 30 2007
Revision Received:
May 12 2008
Accepted:
May 14 2008
Accepted Manuscript online:
May 14 2008
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2008 Biochemical Society
2008
Biochem J (2008) 414 (3): 431–440.
Article history
Received:
November 30 2007
Revision Received:
May 12 2008
Accepted:
May 14 2008
Accepted Manuscript online:
May 14 2008
Citation
Marielle Boonen, Roberta Rezende de Castro, Gaëlle Cuvelier, Isabelle Hamer, Michel Jadot; A dileucine signal situated in the C-terminal tail of the lysosomal membrane protein p40 is responsible for its targeting to lysosomes. Biochem J 15 September 2008; 414 (3): 431–440. doi: https://doi.org/10.1042/BJ20071626
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