A specific dileucine motif is required for the GGA-dependent entry of newly synthesized insulin-responsive aminopeptidase into the insulin-responsive compartment

June Chunqiu Hou, Naoko Suzuki, Jeffrey E. Pessin, Robert T. Watson

Research output: Contribution to journalArticle

28 Scopus citations

Abstract

In muscle and adipose cells, the insulin-responsive aminopeptidase (IRAP) is localized to intracellular storage sites and undergoes insulin-dependent redistribution to the cell surface. Following expression, the newly synthesized IRAP protein traffics to the perinuclear insulin-sensitive compartment and acquires insulin sensitivity 6-9 h following biosynthesis. Knockdown of GGA1 by RNA interference prevented IRAP from entering, but not exiting, the insulin-responsive compartment. Mutation of the dileucine motif at positions 76 and 77 (EGFP-IRAP/AA76,77), but not the dileucine motif at positions 53 and 54, resulted in the rapid default of the reporter to the cell surface beginning at 3 h following biosynthesis. Alanine substitution of 9 residues amino- or carboxyl-terminal to LL76,77 did not perturb basal intracellular sequestration or abrogate insulin-stimulated IRAP translocation. Moreover, a dominant interfering GGA mutant (VHS-GAT) potently inhibited insulin-stimulated translocation of EGFP-IRAP/WT but did not block the constitutive exocytotic trafficking of EGFP-IRAP/AA76,77. In addition, the EGFP-IRAP/WT and EGFP-IRAP/AA76,77 constructs occupied morphologically distinct tubulovesicular compartments in the perinuclear region. Taken together, these data indicate that LL76,77 functions during the GGA-dependent sorting of newly made IRAP into the insulin-responsive storage compartment.

Original languageEnglish (US)
Pages (from-to)33457-33466
Number of pages10
JournalJournal of Biological Chemistry
Volume281
Issue number44
DOIs
StatePublished - Nov 3 2006
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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