Muscarinic-receptor-mediated inhibition of insulin-like growth factor-1 receptor-stimulated phosphoinositide 3-kinase signalling in 1321N1 astrocytoma cells

Ian H Batty, Ian Neil Fleming, C Peter Downes

Research output: Contribution to journalArticle

17 Citations (Scopus)

Abstract

In 1321N1 astrocytoma cells, stimulation of the IGF-1 (insulin-like growth factor-1) receptor increased the association of PI3K [phosphoinositide (PI) 3-kinase] activity with IRS-1 (insulin re-ceptor substrate 1), and increased the cellular concentration of PtdIns(3,4,5)P3. Carbachol, acting on M3 muscarinic receptors, inhibited insulin-, but not PDGF (platelet-derived growth factor)-, stimulated responses by approximately 50%. The inhibition of IRS-1-associated PI3K activity by carbachol (i) was rapid ( or =60 min) and potent (half-maximal concentration approximately 1 microM); (ii) was reproduced by stimuli for several phospholipase-C-coupled receptors; (iii) was prevented by the inhibition of protein kinase C, but not by chelation of intracellular Ca2+; and (iv) was not blocked or reproduced by inhibitors or stimuli respectively of mitogen-activated protein kinase, PI3K, protein kinase B or the mammalian target of rapamycin. However, the effects of carbachol were prevented by sodium vanadate, a protein tyrosine phosphatase inhibitor, and were accompanied by reduced insulin-stimulated IRS-1 tyrosine phosphorylation and recruitment of the 85 kDa regulatory subunit of PI3K to IRS-1, but not by reduced IGF-1 receptor kinase activity. The inhibitory effect of carbachol was reproduced by okadaic acid, a protein serine/threonine phosphatase inhibitor, but not by PDGF, yet all three agents stimulated the serine phosphorylation of IRS-1 at residues Ser312, Ser616 and Ser636/639, albeit to different extents. Thus muscarinic receptors may inhibit insulin signalling by promoting IRS-1 tyrosine dephosphorylation and/or by uncoupling IRS-1 from the stimulated IGF-1 receptor by stimulating IRS-1 serine phosphorylation. However, the proportion of IRS-1 molecules phosphorylated at a particular site or the phosphorylation of additional IRS-1 serine residues other than those noted above must be important.
Original languageEnglish
Pages (from-to)641-651
Number of pages11
JournalBiochemical Journal
Volume379
Issue number3
DOIs
Publication statusPublished - 1 May 2004

Keywords

  • astrocytoma
  • calcium
  • carbachol
  • tumor cell line
  • enzyme activation
  • humans
  • inositol 1,4,5-trisphosphate
  • insulin
  • insulin receptor substrate proteins
  • Ionomycin
  • mitogen-activated protein kinases
  • okadaic acid
  • phosphatidylinositol 3-kinases
  • phosphoproteins
  • phosphorylation
  • phosphoserine
  • phosphotyrosine
  • platelet-derived growth factor
  • protein kinase C
  • protein-serine-threonine kinases
  • proto-oncogene proteins
  • proto-oncogene proteins c-akt
  • IGF type 1 receptor
  • muscarinic receptors
  • ribosomal protein S6 kinases, 70-kDa
  • signal transduction
  • tetradecanoylphorbol acetate
  • type C phospholipases

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