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Author Notes:

Correspondence: Lih-Shen Chin, Department of Pharmacology, Emory University School of Medicine, 1510 Clifton Road, Atlanta, Georgia, USA, 30322. Fax: 1-404-727-0365; Phone: 1-404-727-0361; Email: chinl@pharm.emory.edu; Website: http://www.pharm.emory.edu/

J.W.P., E.A.W., and D.S. contributed equally to this work.

We thank Drs. Tom Südhof and Cam Patterson for providing the expression constructs for Munc18-1 and Hsc70, respectively.

Subjects:

Research Funding:

J.W.P was supported by National Institute of Neurological Disorders and Stroke Training Grant T32NS007480.

This work was supported by grants from National Institutes of Health (NS047575 and GM082828 to L.L. and NS050650 to L.-S.C.).

Keywords:

  • Ubiquitination
  • ubiquitin-interacting motif
  • Hrs
  • endocytic trafficking
  • in vitro expression cloning

Proteomic analysis reveals Hrs UIM-mediated ubiquitin signaling in multiple cellular processes

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Journal Title:

FEBS Journal

Volume:

Volume 276, Number 1

Publisher:

, Pages 118-131

Type of Work:

Article | Post-print: After Peer Review

Abstract:

Despite the critical importance of protein ubiquitination in regulation of diverse cellular processes, the molecular mechanisms by which cells recognize and transmit ubiquitin signals remain poorly understood. The endosomal sorting machinery component hepatocyte growth factor regulated tyrosine kinase substrate (Hrs) contains an ubiquitin-interacting motif (UIM), which is believed to bind ubiquitinated membrane cargo proteins and mediate their sorting to the lysosomal degradation pathway. To gain insight into the role of Hrs UIM-mediated ubiquitin signaling in cells, we performed a proteomic screen for Hrs UIM-interacting ubiquitinated proteins in human brain by using an in vitro expression cloning (IVEC) screening approach. We have identified 48 ubiquitinated proteins that are specifically recognized by the UIM domain of Hrs. Among them, 12 are membrane proteins which are likely to be Hrs cargo proteins, and 4 are membrane protein-associated adaptor proteins whose ubiquitination may act as a signal to target their associated membrane cargo for Hrs-mediated endosomal sorting. Other classes of the identified proteins include components of the vesicular trafficking machinery, cell signaling molecules, proteins associated with the cytoskeleton and cytoskeleton-dependent transport, and enzymes involved in ubiquitination and metabolism, suggesting the involvement of Hrs UIM-mediated ubiquitin signaling in regulation of multiple cellular processes. We have characterized the ubiquitination of two identified proteins, Munc18-1 and Hsc70, and their interaction with Hrs UIM and provided functional evidence supporting a role for Hsc70 in regulation of Hrs-mediated endosome-to-lysosome trafficking.

Copyright information:

© 2008 The Authors Journal compilation © 2008 FEBS

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