Huntingtin-associated Protein 1 Interacts with Hepatocyte Growth Factor-regulated Tyrosine Kinase Substrate and Functions in Endosomal Trafficking

Yankun Li,Lih-Shen Chin,Allan I Levey,Lian Li

doi:10.1074/jbc.m111612200

Abstract

Huntingtin-associated protein 1 (HAP1) is a novel protein of unknown function with a higher binding affinity for the mutant form of Huntington's disease protein huntingtin. Here we report that HAP1 interacts with hepatocyte growth factor-regulated tyrosine kinase substrate (Hrs), a mammalian homologue of yeast vacuolar protein sorting protein Vps27p involved in the endosome-to-lysosome trafficking. This novel interaction was identified in a yeast two-hybrid screen using full-length Hrs as bait, and confirmed by in vitro binding assays and co-immunoprecipitation experiments. Deletion analysis reveals that the association of HAP1 with Hrs is mediated via a coiled-coil interaction between the central coiled-coil domains of both proteins. Immunofluorescence and subcellular fractionation studies show that HAP1 co-localizes with Hrs on early endosomes. Like Hrs, overexpression of HAP1 causes the formation of enlarged early endosomes, and inhibits the degradation of internalized epidermal growth factor receptors. Whereas overexpression of HAP1 does not affect either constitutive or ligand-induced receptor-mediated endocytosis, it potently blocks the trafficking of endocytosed epidermal growth factor receptors from early endosomes to late endosomes. These findings implicate, for the first time, the involvement of HAP1 in the regulation of vesicular trafficking from early endosomes to the late endocytic compartments.

Highlights

Huntington’s disease (HD)1 is a neurodegenerative disorder caused by the expansion of a polyglutamine stretch (Ն36 glutamines) at the N terminus of huntingtin, a ubiquitously expressed protein of unknown function [1, 2]
We report that Huntingtin-associated protein 1 (HAP1) interacts with hepatocyte growth factor-regulated tyrosine kinase substrate (Hrs), a mammalian homologue of yeast vacuolar protein sorting protein Vps27p involved in the endosome-to-lysosome trafficking
We identify a novel interaction between HAP1 and hepatocyte growth factor-regulated tyrosine kinase substrate (Hrs), an early endosome-associated phosphoprotein involved in the regulation of vesicular trafficking and signal transduction (10 –12)

Summary

Introduction

Huntington’s disease (HD)1 is a neurodegenerative disorder caused by the expansion of a polyglutamine stretch (Ն36 glutamines) at the N terminus of huntingtin, a ubiquitously expressed protein of unknown function [1, 2]. In support of this role, our previous work demonstrates that Hrs regulates the trafficking of EGF receptors from early endosomes to lysosomes via its interaction with sorting nexin 1, a mammalian homologue of yeast Vps5p that binds to the lysosomal targeting region of the EGF receptor [20].

Results

Conclusion