AT1 receptor mutant lacking heterotrimeric G protein coupling activates the Src-Ras-ERK pathway without nuclear translocation of ERKs

J Biol Chem. 2002 Mar 15;277(11):9268-77. doi: 10.1074/jbc.M109221200. Epub 2002 Jan 3.

Abstract

Angiotensin II (Ang II) type 1 receptors (AT1Rs) activate tyrosine kinases, including Src. Whether or not tyrosine kinase activation by AT1R occurs independently of heterotrimeric G protein coupling and, if so, the cellular function of such a mechanism are unknown. To address these questions, we used an AT1aR intracellular second loop mutant, which lacks heterotrimeric G protein coupling (AT1a-i2m). Surprisingly, Ang II-induced Src activation was preserved in AT1a-i2m, which was not attenuated by inhibiting protein kinase C and Ca(2+) or by inhibiting Galpha(i) or Galpha(q) in CHO-K1 cells. By contrast, Ang II-induced Src activation was abolished in a C-terminally truncated AT1a-(1--309), where Ang II-induced inositol phosphate response was preserved. Ang II activates ERKs via a Src-Ras-dependent mechanism in AT1a-i2m. ERKs activated by AT1a-i2m phosphorylate their cytoplasmic targets, including p90(RSK), but fail to translocate into the nucleus or to cause cell proliferation. Ang II-induced nuclear translocation of ERKs by wild type AT1aR was inhibited by overexpression of nuclear exportin Crm-1, while that by AT1a-i2m was restored by leptomycin B, an inhibitor of Crm-1. In summary, while Src and ERKs are activated by Ang II even without heterotrimeric G protein coupling, the carboxyl terminus of the AT1 receptor is required for activation of Src. Interestingly, ERKs activated by heterotrimeric G protein-independent mechanisms fail to phosphorylate nuclear targets due to lack of inhibition of Crm-1-induced nuclear export of ERKs. These results suggest that heterotrimeric G protein-dependent and -independent signaling mechanisms play distinct roles in Ang II-mediated cellular responses.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Active Transport, Cell Nucleus*
  • Angiotensin II / pharmacology
  • Animals
  • CHO Cells
  • Cell Division / drug effects
  • Cricetinae
  • DNA-Binding Proteins*
  • Enzyme Activation
  • Exportin 1 Protein
  • Heterotrimeric GTP-Binding Proteins / physiology*
  • Karyopherins / metabolism
  • Mitogen-Activated Protein Kinases / metabolism*
  • Proto-Oncogene Proteins / metabolism
  • Receptor, Angiotensin, Type 1
  • Receptors, Angiotensin / physiology*
  • Receptors, Cytoplasmic and Nuclear*
  • Transcription Factors*
  • Transfection
  • ets-Domain Protein Elk-1
  • ras Proteins / physiology*
  • src-Family Kinases / physiology*

Substances

  • DNA-Binding Proteins
  • Karyopherins
  • Proto-Oncogene Proteins
  • Receptor, Angiotensin, Type 1
  • Receptors, Angiotensin
  • Receptors, Cytoplasmic and Nuclear
  • Transcription Factors
  • ets-Domain Protein Elk-1
  • Angiotensin II
  • src-Family Kinases
  • Mitogen-Activated Protein Kinases
  • Heterotrimeric GTP-Binding Proteins
  • ras Proteins