QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

First published online June 23, 2005
doi: 10.1242/10.1242/jcs.02434

This Article Figures Only Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Nair, P. Articles by Rohrer, J. Search for Related Content PubMed PubMed Citation Articles by Nair, P. Articles by Rohrer, J.

Characterization of the TGN exit signal of the human mannose 6-phosphate uncovering enzyme

Prashant Nair^1,*, Beat E. Schaub^1,*, Kai Huang², Xiang Chen², Robert F. Murphy^2,3, Janice M. Griffith⁴, Hans J. Geuze⁴ and Jack Rohrer^1,

¹ Institute of Physiology, University of Zurich, Zurich, 8057, Switzerland
² Department of Biological Sciences, Carnegie Mellon University, Pittsburgh, PA 15213, USA
³ Department of Biomedical Engineering, Carnegie Mellon University, Pittsburgh, PA 15213, USA
⁴ Department of Cell Biology and Institute of Biomembranes, Utrecht University School of Medicine, 3584 CX Utrecht, The Netherlands

Author for correspondence (e-mail: rohrer{at}access.unizh.ch)

Accepted 12 April 2005

The human mannose 6-phosphate uncovering enzyme participates in the uncovering of the mannose 6-phosphate recognition tag on lysosomal enzymes, a process that facilitates recognition of those enzymes by mannose 6-phosphate receptors to ensure delivery to lysosomes. Uncovering enzyme has been identified on the trans-Golgi network at steady state. It has been shown to traffic to the plasma membrane from where it is rapidly internalized via endosomal structures, the process being mediated by a tyrosine-based internalization motif, Y⁴⁸⁸HPL, in its cytoplasmic tail. Using immunogold electron microscopy a GFP-uncovering enzyme fusion construct was found to be colocalized with the cation-dependent mannose 6-phosophate receptor in regions of the trans-Golgi network, suggesting that uncovering enzyme might follow a similar pathway of exit from the trans-Golgi network as that of the cation-dependent mannose 6-phosohate receptor. In this study, we identified the signal sequence in the cytoplasmic tail of uncovering enzyme responsible for its exit from the trans-Golgi network. Using GFP fusion constructs of the transmembrane and cytoplasmic domains of uncovering enzyme, we could show, by automated analysis of confocal immunofluorescence images, that residues Q⁴⁹²EMN in the cytoplasmic tail of uncovering enzyme are involved in its exit from the trans-Golgi network. Detailed characterization of the exit signal revealed that residue Q⁴⁹² is the most important to the exit function while M⁴⁹⁴ and N⁴⁹⁵ also contribute. The cytoplasmic tail of the uncovering enzyme does not possess any of the known canonical signal sequences for interaction with Golgi-associated gamma ear-containing adaptor proteins. The identification of a trans-Golgi network exit signal in its cytoplasmic tail elucidates the trafficking pathway of uncovering enzyme, a crucial player in the process of lysosomal biogenesis.

Key words: Lysosomal biogenesis, Uncovering enzyme, trans-Golgi network, TGN exit, Subcellular location features, Protein distribution comparison