Regulation of smoothened by Drosophila G-protein-coupled receptor kinases

Dev Biol. 2010 Jan 1;337(1):99-109. doi: 10.1016/j.ydbio.2009.10.014. Epub 2009 Oct 20.

Abstract

The Hedgehog (Hh) signaling pathway plays a conserved and essential role in regulating development and homeostasis of numerous tissues. Cytoplasmic signaling is initiated by Smoothened (Smo), a G-protein-coupled receptor (GPCR) family member, whose levels and activity are regulated by the Hh receptor Patched (Ptc). In response to Hh binding to Ptc, Ptc-mediated repression of Smo is relieved, leading to Smo activation, surface accumulation, and downstream signaling. We find that downregulation of Drosophila Smo protein in Hh-responding imaginal disc cells is dependent on the activity of G-protein-coupled receptor kinase 2 (Gprk2). By analyzing gain- and null loss-of-function phenotypes, we provide evidence that Gprk2 promotes Smo internalization subsequent to its activation, most likely by direct phosphorylation. Ptc-dependent regulation of Smo accumulation is normal in gprk2 mutants, indicating that Gprk2 and Ptc downregulate Smo by different mechanisms. Finally, we show that both Drosophila G-protein-coupled receptor kinase orthologues, Gprk1 and Gprk2, act in a partially redundant manner to promote Hh signaling. Our results suggest that Smo is regulated by distinct Ptc-dependent and Gprk2-dependent trafficking mechanisms in vivo, analogous to constitutive and activity-dependent regulation of GPCRs. G-protein-coupled receptor kinase activity is also important for efficient downstream signaling.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Arrestins / physiology
  • Cells, Cultured
  • Drosophila Proteins / genetics*
  • Drosophila Proteins / physiology*
  • Drosophila melanogaster
  • G-Protein-Coupled Receptor Kinase 1 / physiology
  • G-Protein-Coupled Receptor Kinase 2 / physiology*
  • Gene Expression Regulation*
  • Hedgehog Proteins / physiology
  • Phosphorylation
  • Receptors, Cell Surface / physiology
  • Receptors, G-Protein-Coupled / genetics*
  • Signal Transduction
  • Smoothened Receptor
  • Temperature
  • beta-Arrestins

Substances

  • Arrestins
  • Drosophila Proteins
  • Hedgehog Proteins
  • Receptors, Cell Surface
  • Receptors, G-Protein-Coupled
  • Smoothened Receptor
  • beta-Arrestins
  • ptc protein, Drosophila
  • smo protein, Drosophila
  • hh protein, Drosophila
  • G-Protein-Coupled Receptor Kinase 1
  • G-Protein-Coupled Receptor Kinase 2
  • Gprk2 protein, Drosophila