TY - JOUR
T1 - YXXM Motifs in the PDGF-β Receptor Serve Dual Roles as Phosphoinositide 3-Kinase Binding Motifs and Tyrosine-based Endocytic Sorting Signals
AU - Wu, Haiyan
AU - Windmiller, David A.
AU - Wang, Ling
AU - Backer, Jonathan M.
PY - 2003/10/17
Y1 - 2003/10/17
N2 - Phosphoinositide 3-kinases (PI 3-kinases) are important regulators of endocytic trafficking. Previous studies have shown that mutant human platelet-derived growth factor-β receptors (PDGFR), which contain Phe in place of Tyr at the two p85/p110 PI 3-kinase binding sites (PDGFR-F/F), are defective for both p85 binding and ligand-stimulated degradation. This suggested that p85/p110 regulates PDGFR trafficking. However, more recent work has identified hVPS34, and not p85/p110, as the major PI 3-kinase regulating the movement of receptors through the endosomal system. To reconcile this discrepancy, we hypothesized that YXXM motifs in the PDGFR might play a second role as Tyr-based lysosomal sorting motifs (YXXφ). To test this, we replaced both YXXM motifs with a motif from LAMP-1, YQTI. This mutant PDGFR (PDGFR-YQTI) still underwent PDGF-stimulated autophosphorylation but did not bind p85. In CHO cells, both wild-type and YQTI receptors showed PDGF-stimulated turnover, whereas F/F receptors did not. In addition, uptake and degradation of cell surface-labeled YXXM and YQTI receptors was fast relative to F/F receptors. We also constructed chimeras containing extracellular and membrane-spanning domains from CD25 (Tac) and cytoplasmic tails containing the YQTI motif, two YXXM motifs, or two mutant FXXM motifs. The YXXM and YQTI chimeras mediated lysosomal delivery of fluorescein isothiocyanate-labeled anti-CD25 antibodies, whereas the F/F chimera was defective. Thus, YQTI motifs can target PDGFR for degradation in the absence of p85/p110 binding, and the p85/p110 binding motifs from PDGFR are sufficient to target Tac chimeras to the lysosome. These data suggest that the YXXM motifs in the PDGFR serve two distinct functions: nase recruitment and lysosomal targeting.
AB - Phosphoinositide 3-kinases (PI 3-kinases) are important regulators of endocytic trafficking. Previous studies have shown that mutant human platelet-derived growth factor-β receptors (PDGFR), which contain Phe in place of Tyr at the two p85/p110 PI 3-kinase binding sites (PDGFR-F/F), are defective for both p85 binding and ligand-stimulated degradation. This suggested that p85/p110 regulates PDGFR trafficking. However, more recent work has identified hVPS34, and not p85/p110, as the major PI 3-kinase regulating the movement of receptors through the endosomal system. To reconcile this discrepancy, we hypothesized that YXXM motifs in the PDGFR might play a second role as Tyr-based lysosomal sorting motifs (YXXφ). To test this, we replaced both YXXM motifs with a motif from LAMP-1, YQTI. This mutant PDGFR (PDGFR-YQTI) still underwent PDGF-stimulated autophosphorylation but did not bind p85. In CHO cells, both wild-type and YQTI receptors showed PDGF-stimulated turnover, whereas F/F receptors did not. In addition, uptake and degradation of cell surface-labeled YXXM and YQTI receptors was fast relative to F/F receptors. We also constructed chimeras containing extracellular and membrane-spanning domains from CD25 (Tac) and cytoplasmic tails containing the YQTI motif, two YXXM motifs, or two mutant FXXM motifs. The YXXM and YQTI chimeras mediated lysosomal delivery of fluorescein isothiocyanate-labeled anti-CD25 antibodies, whereas the F/F chimera was defective. Thus, YQTI motifs can target PDGFR for degradation in the absence of p85/p110 binding, and the p85/p110 binding motifs from PDGFR are sufficient to target Tac chimeras to the lysosome. These data suggest that the YXXM motifs in the PDGFR serve two distinct functions: nase recruitment and lysosomal targeting.
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U2 - 10.1074/jbc.C300225200
DO - 10.1074/jbc.C300225200
M3 - Article
C2 - 12941951
AN - SCOPUS:0142040128
SN - 0021-9258
VL - 278
SP - 40425
EP - 40428
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 42
ER -