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

Koichi Seta, Masakatsu Nanamori, J. Gregory Modrall, Richard R. Neubig, Junichi Sadoshima

Research output: Contribution to journalArticle

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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 ATlaR 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 Ca2+ or by inhibiting Gαi or Gαq 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 p90RSK, 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.

Original languageEnglish (US)
Pages (from-to)9268-9277
Number of pages10
JournalJournal of Biological Chemistry
Volume277
Issue number11
DOIs
StatePublished - Mar 15 2002

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Heterotrimeric GTP-Binding Proteins
MAP Kinase Signaling System
Angiotensin II
Chemical activation
Protein-Tyrosine Kinases
Karyopherins
Angiotensin Type 1 Receptor
Inositol Phosphates
Cell Nucleus Active Transport
src-Family Kinases
CHO Cells
Cell proliferation
Protein Kinase C
Cell Proliferation

ASJC Scopus subject areas

  • Biochemistry

Cite this

AT1 receptor mutant lacking heterotrimeric G protein coupling activates the Src-Ras-ERK pathway without nuclear translocation of ERKs. / Seta, Koichi; Nanamori, Masakatsu; Gregory Modrall, J.; Neubig, Richard R.; Sadoshima, Junichi.

In: Journal of Biological Chemistry, Vol. 277, No. 11, 15.03.2002, p. 9268-9277.

Research output: Contribution to journalArticle

Seta, Koichi ; Nanamori, Masakatsu ; Gregory Modrall, J. ; Neubig, Richard R. ; Sadoshima, Junichi. / AT1 receptor mutant lacking heterotrimeric G protein coupling activates the Src-Ras-ERK pathway without nuclear translocation of ERKs. In: Journal of Biological Chemistry. 2002 ; Vol. 277, No. 11. pp. 9268-9277.
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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 ATlaR 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 Ca2+ or by inhibiting Gαi or Gαq 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 p90RSK, 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.",
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T1 - AT1 receptor mutant lacking heterotrimeric G protein coupling activates the Src-Ras-ERK pathway without nuclear translocation of ERKs

AU - Seta, Koichi

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AU - Neubig, Richard R.

AU - Sadoshima, Junichi

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