Coordinated regulation of AP2 uncoating from clathrin-coated vesicles by rab5 and hRME-6

J Cell Biol. 2008 Nov 3;183(3):499-511. doi: 10.1083/jcb.200806016.


Here we investigate the role of rab5 and its cognate exchange factors rabex-5 and hRME-6 in the regulation of AP2 uncoating from endocytic clathrin-coated vesicles (CCVs). In vitro, we show that the rate of AP2 uncoating from CCVs is dependent on the level of functional rab5. In vivo, overexpression of dominant-negative rab5(S34N), or small interfering RNA (siRNA)-mediated depletion of hRME-6, but not rabex-5, resulted in increased steady-state levels of AP2 associated with endocytic vesicles, which is consistent with reduced uncoating efficiency. hRME-6 guanine nucleotide exchange factor activity requires hRME-6 binding to alpha-adaptin ear, which displaces the ear-associated mu2 kinase AAK1. siRNA-mediated depletion of hRME-6 increases phospho-mu2 levels, and expression of a phosphomimetic mu2 mutant increases levels of endocytic vesicle-associated AP2. Depletion of hRME-6 or rab5(S35N) expression also increases the levels of phosphoinositide 4,5-bisphosphate (PtdIns(4,5)P(2)) associated with endocytic vesicles. These data are consistent with a model in which hRME-6 and rab5 regulate AP2 uncoating in vivo by coordinately regulating mu2 dephosphorylation and PtdIns(4,5)P(2) levels in CCVs.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adaptor Protein Complex 2 / physiology*
  • Animals
  • Cell Line
  • Cell Membrane / physiology
  • Clathrin-Coated Vesicles / physiology*
  • Cytosol / physiology
  • Endocytosis / physiology*
  • Humans
  • Kidney
  • Membrane Proteins / isolation & purification
  • Membrane Proteins / physiology
  • Mice
  • Mice, Knockout
  • Nerve Tissue Proteins / deficiency
  • Nerve Tissue Proteins / physiology
  • Neurons / physiology*
  • Phosphoric Monoester Hydrolases / deficiency
  • Phosphoric Monoester Hydrolases / physiology
  • rab5 GTP-Binding Proteins / physiology*


  • Adaptor Protein Complex 2
  • Membrane Proteins
  • Nerve Tissue Proteins
  • synaptojanin
  • Phosphoric Monoester Hydrolases
  • rab5 GTP-Binding Proteins