Reciprocal interaction with G-actin and tropomyosin is essential for aquaporin-2 trafficking

J Cell Biol. 2008 Aug 11;182(3):587-601. doi: 10.1083/jcb.200709177. Epub 2008 Aug 4.


Trafficking of water channel aquaporin-2 (AQP2) to the apical membrane and its vasopressin and protein kinase A (PKA)-dependent regulation in renal collecting ducts is critical for body water homeostasis. We previously identified an AQP2 binding protein complex including actin and tropomyosin-5b (TM5b). We show that dynamic interactions between AQP2 and the actin cytoskeleton are critical for initiating AQP2 apical targeting. Specific binding of AQP2 to G-actin in reconstituted liposomes is negatively regulated by PKA phosphorylation. Dual color fluorescence cross-correlation spectroscopy reveals local AQP2 interaction with G-actin in live epithelial cells at single-molecule resolution. Cyclic adenosine monophosphate signaling and AQP2 phosphorylation release AQP2 from G-actin. In turn, AQP2 phosphorylation increases its affinity to TM5b, resulting in reduction of TM5b bound to F-actin, subsequently inducing F-actin destabilization. RNA interference-mediated knockdown and overexpression of TM5b confirm its inhibitory role in apical trafficking of AQP2. These findings indicate a novel mechanism of channel protein trafficking, in which the channel protein itself critically regulates local actin reorganization to initiate its movement.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Actins / metabolism*
  • Animals
  • Aquaporin 2 / metabolism*
  • Cell Line
  • Cell Polarity
  • Cell Survival
  • Cyclic AMP-Dependent Protein Kinases / metabolism
  • Dogs
  • Green Fluorescent Proteins / metabolism
  • Humans
  • Liposomes / metabolism
  • Models, Biological
  • Phosphorylation
  • Protein Binding
  • Protein Transport
  • RNA Interference
  • Rats
  • Recombinant Proteins / metabolism
  • Spectrometry, Fluorescence
  • Tropomyosin / metabolism*


  • Actins
  • Aquaporin 2
  • Liposomes
  • Recombinant Proteins
  • Tropomyosin
  • Green Fluorescent Proteins
  • Cyclic AMP-Dependent Protein Kinases