Transforming growth factor β1 treatment of AKR-2B cells is coupled through a pertussis-toxin-sensitive G-protein(s)

P. H. Howe, E. B. Leof

Research output: Contribution to journalArticle

44 Scopus citations

Abstract

Transforming growth factor β (TGF β1) is a potent regulator of DNA synthesis and cellular proliferation. In this study, we investigated whether the growth stimulatory signal of TGF β1 is transduced intracellularly by guanine nucleotide regulatory proteins (G-proteins). In plasma membranes from AKR-2B cells, TGF β1 increased binding of the radiolabelled, non-hydrolysable GTP analogue, guanosine 5'-[γ-[35S]thio]triphosphate (GTP[35S]), in a dose-dependent manner. Maximal effects occurred between 0.4 and 1.0 nM-TGFβ1. Specific binding of GTP[35S] occurred with a K(d) of 3.2 x 10-8 M which was not affected by addition of TGFβ1. Instead, TGFβ1 increased the number of available binding sites for GTP[35S] from 16.2 ± 1.2 to 21.6 ± 2.1 pmol/mg of protein. GTP[35S] binding was both nucleotide- and growth-factor-specific. Only guanine nucleotides were able to compete for binding, and of the growth factors tested (epidermal growth factor, platelet-derived growth factor, insulin, TGFβ1 and TGFβ2) only TGFβ1 affected GTP[35S] binding. TGFβ1 increased GTPase activity, as determined by the release of 32PO43- from GTPγ[32P], from 116 ± 5.5 to 175 ± 4.3 pmol/mg of protein following a 15 min incubation. Pretreatment of the membranes with pertussis toxin inhibited both TGFβ1-stimulated binding of GTP[35S] as well as TGFβ1-stimulated GTPase activity. These inhibitory actions of pertussis toxin were associated with toxin-induced ADP-ribosylation of a 41 kDa protein. Furthermore, the stimulatory effects of TGFβ1 on c-sis mRNA expression were shown to be pertussis-toxin sensitive and could be mimicked by direct activation of G-proteins with AlF4-. These results demonstrate that in AKR-2B cells a pertussis-toxin-sensitive guanine nucleotide regulatory protein(s) is coupled to TGFβ1 receptor binding.

Original languageEnglish (US)
Pages (from-to)879-886
Number of pages8
JournalBiochemical Journal
Volume261
Issue number3
DOIs
StatePublished - 1989

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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