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 ≈50%. The inhibition of IRS-1-associated PI3K activity by carbachol (i) was rapid (<1 min), persistent (≥60 min) and potent (half-maximal concentration ≈1 µM); (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.
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Research Article|
May 01 2004
Muscarinic-receptor-mediated inhibition of insulin-like growth factor-1 receptor-stimulated phosphoinositide 3-kinase signalling in 1321N1 astrocytoma cells
Ian H. BATTY;
Ian H. BATTY
1
The Division of Cell Signalling, School of Life Sciences, MSI/WTB Complex, University of Dundee, Dow St, Dundee DD1 5EH, Scotland, U.K.
1To whom correspondence should be addressed (e-mail i.h.batty@dundee.ac.uk).
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Ian N. FLEMING;
Ian N. FLEMING
2
The Division of Cell Signalling, School of Life Sciences, MSI/WTB Complex, University of Dundee, Dow St, Dundee DD1 5EH, Scotland, U.K.
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C. Peter DOWNES
C. Peter DOWNES
The Division of Cell Signalling, School of Life Sciences, MSI/WTB Complex, University of Dundee, Dow St, Dundee DD1 5EH, Scotland, U.K.
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Publisher: Portland Press Ltd
Received:
November 06 2003
Revision Received:
January 22 2004
Accepted:
February 09 2004
Accepted Manuscript online:
February 09 2004
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London ©2004
2004
Biochem J (2004) 379 (3): 641–651.
Article history
Received:
November 06 2003
Revision Received:
January 22 2004
Accepted:
February 09 2004
Accepted Manuscript online:
February 09 2004
Citation
Ian H. BATTY, Ian N. FLEMING, C. Peter DOWNES; Muscarinic-receptor-mediated inhibition of insulin-like growth factor-1 receptor-stimulated phosphoinositide 3-kinase signalling in 1321N1 astrocytoma cells. Biochem J 1 May 2004; 379 (3): 641–651. doi: https://doi.org/10.1042/bj20031700
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