This Article Full Text Full Text (PDF) Purchase Article View Shopping Cart Alert me when this article is cited Alert me if a correction is posted Citation Map 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 Request Copyright Permission Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Haft, C. R. Articles by Taylor, S. I. Search for Related Content PubMed PubMed Citation Articles by Haft, C. R. Articles by Taylor, S. I.

Analysis of the Juxtamembrane Dileucine Motif in the Insulin Receptor¹

Diabetes Branch, National Institutes of Diabetes and Digestive and Kidney Diseases, National Institutes of Health (C.R.H., M.d.L.L.S., S.I.T.), Bethesda, Maryland 20892; and the Department of Morphology, University of Geneva (I.H., J.-L.C.), Geneva, Switzerland

Address all correspondence and requests for reprints to: Simeon I. Taylor, M.D., Ph.D., National Institutes of Health, Building 10, Room 9S-213, 10 Center Drive, MSC-1829, Bethesda, Maryland 20892-1829.

Dileucine-containing motifs are involved in trans-Golgi sorting, lysosomal targeting, and internalization. Previously, we have shown that the dileucine motif (EKITLL, residues 982–987) in the juxtamembrane region of the insulin receptor is involved in receptor internalization. Substitution of alanine residues for Leu⁹⁸⁶ and Leu⁹⁸⁷ led to a 3- to 5-fold decrease in the ability of the receptors to mediate insulin uptake. In the current study, we show that mutation of the same motif to Met⁹⁸⁶Ser⁹⁸⁷, the sequence found in the homologous position in the type I insulin-like growth factor receptor, did not affect insulin uptake. Therefore, we inquired whether the sequence EKITMS as an isolated motif could mediate the targeting of a reporter molecule to endosomes and then lysosomes, as was shown previously with the EKITLL motif of the normal receptor. Chimeric molecules containing Tac antigen fused to different hexapeptide sequences showed distinct patterns of subcellular localization by immunofluorescence microscopy. Tac-EKITLL and Tac-EKITAA were found predominantly in lysosomes and the plasma membrane, respectively. In contrast, Tac-EKITMS was found at the plasma membrane, in the trans-Golgi network, and in endosomes, but only small amounts were found in lysosomes. Thus, the dileucine motif (EKITLL) plays an important role in directing endocytosis of the intact insulin receptor and in mediating efficient endocytosis and lysosomal targeting as an isolated motif. Substitution of AA for LL inhibits endocytosis and lysosomal targeting in both systems. In contrast, substitution of MS for LL permits rapid endocytosis in the intact receptor, but mediates modest endocytosis and very little targeting to lysosomes as an isolated motif. Our observations support the idea that sorting signals are recognized at multiple steps in the cell, and that specific amino acid substitutions may differentially affect each of these sorting steps.

This article has been cited by other articles:

				N. Marr, D. G Bichet, M. Lonergan, M.-F. Arthus, N. Jeck, H. W. Seyberth, W. Rosenthal, C. H. van Os, A. Oksche, and P. M. T. Deen Heteroligomerization of an Aquaporin-2 mutant with wild-type Aquaporin-2 and their misrouting to late endosomes/lysosomes explains dominant nephrogenic diabetes insipidus Hum. Mol. Genet., April 1, 2002; 11(7): 779 - 789. [Abstract] [Full Text] [PDF]

				P. van Kerkhof, R. Govers, C. M. Alves dos Santos, and G. J. Strous Endocytosis and Degradation of the Growth Hormone Receptor Are Proteasome-dependent J. Biol. Chem., January 21, 2000; 275(3): 1575 - 1580. [Abstract] [Full Text] [PDF]

				G. Strous and R Govers The ubiquitin-proteasome system and endocytosis J. Cell Sci., January 5, 1999; 112(10): 1417 - 1423. [Abstract] [PDF]

				I. V. Sandoval, S. Martinez-Arca, J. Valdueza, S. Palacios, and G. D. Holman Distinct Reading of Different Structural Determinants Modulates the Dileucine-mediated Transport Steps of the Lysosomal Membrane Protein LIMPII and the Insulin-sensitive Glucose Transporter GLUT4 J. Biol. Chem., December 15, 2000; 275(51): 39874 - 39885. [Abstract] [Full Text] [PDF]