Regulation of CB1 cannabinoid receptor internalization by a promiscuous phosphorylation-dependent mechanism

J Neurochem. 2008 Jul;106(1):70-82. doi: 10.1111/j.1471-4159.2008.05336.x. Epub 2008 Jul 1.


Agonists stimulate cannabinoid 1 receptor (CB(1)R) internalization. Previous work suggests that the extreme carboxy-terminus of the receptor regulates this internalization - likely through the phosphorylation of serines and threonines clustered within this region. While truncation of the carboxy-terminus (V460Z CB(1)) and consequent removal of these putative phosphorylation sites prevents endocytosis in AtT20 cells, the residues necessary for CB(1)R internalization remain elusive. To determine the structural requirements for internalization, we evaluated endocytosis of carboxy-terminal mutant CB(1)Rs stably expressed in HEK293 cells. In contrast to AtT20 cells, V460Z CB(1)R expressed in HEK293 cells internalized to the same extent and with similar kinetics as the wild-type receptor. However, mutation of serine and/or threonine residues within the extreme carboxy-terminal attenuated internalization when these receptors were expressed in HEK293 cells. These results establish that the extreme carboxy-terminal phosphorylation sites are not required for internalization of truncated receptors, but are required for internalization of full-length receptors in HEK293 cells. Analysis of beta-arrestin-2 recruitment to mutant CB(1)R suggests that putative carboxy-terminal phosphorylation sites mediate beta-arrestin-2 translocation. This study indicates that the local cellular environment affects the structural determinants of CB(1)R internalization. Additionally, phosphorylation likely regulates the internalization of (full-length) CB(1)Rs.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Amino Acid Sequence / physiology
  • Animals
  • Arrestins / metabolism
  • Cannabinoids / pharmacology
  • Cell Membrane / drug effects
  • Cell Membrane / metabolism*
  • Endocytosis / drug effects
  • Endocytosis / physiology*
  • Humans
  • Mice
  • Molecular Weight
  • Phosphorylation / drug effects
  • Protein Structure, Tertiary / physiology
  • Protein Transport / drug effects
  • Protein Transport / physiology
  • Receptor, Cannabinoid, CB1 / agonists
  • Receptor, Cannabinoid, CB1 / chemistry
  • Receptor, Cannabinoid, CB1 / metabolism*
  • beta-Arrestin 2
  • beta-Arrestins


  • ARRB2 protein, human
  • Arrb2 protein, mouse
  • Arrestins
  • Cannabinoids
  • Receptor, Cannabinoid, CB1
  • beta-Arrestin 2
  • beta-Arrestins