Abstract.
Aims/hypothesis:
In primary adipocytes, although IRS-1 and IRS-3 are expressed in comparable amounts, these proteins manifest distinct distribution and significance in insulin signalling. We investigated the molecular basis of the difference between these two proteins.
Methods:
In Cos-1 cells transiently expressing rat IRS-1, IRS-3, or chimeric proteins of these two proteins we examined the tyrosine phosphorylation via the wild-type or mutant insulin receptors and evaluated their targeting to the plasma membrane by immunostaining the membrane ghost.
Results:
In contrast to IRS-1, IRS-3 was tyrosine-phosphorylated by the insulin receptor altering Tyr960 to Phe (Y960F), which disrupts the binding site of the PTB domain of IRSs, to an extent comparable to the wild-type receptor. The tyrosine phosphorylation of IRS-3 with the PH domain replacement via the Y960F insulin receptor markedly decreased, whereas that of IRS-3 with the PTB domain alteration was mildly impaired. Insulin-stimulated translocation of IRS-1 to the plasma membrane, as well as that of IRS-3 with the PH domain replacement, was wortmannin-sensitive, although that of IRS-3 was insulin-independent and wortmannin-resistant.
Conclusions/interpretation:
The affinity of the PH domain for the phospholipids in the plasma membrane seems to influence the receptor-substrate interaction required for IRS tyrosine phosphorylation, indicating that the PH domain and the PTB domain of IRSs cooperatively function in insulin-stimulated tyrosine phosphorylation of these proteins. [Diabetologia (2001) 44: 992–1004]
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Received: 17 October 2000 and in revised form: 27 March 2001
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Kaburagi, Y., Satoh, S., Yamamoto-Honda, R. et al. Insulin-independent and wortmannin-resistant targeting of IRS-3 to the plasma membrane via its pleckstrin homology domain mediates a different interaction with the insulin receptor from that of IRS-1. Diabetologia 44, 992–1004 (2001). https://doi.org/10.1007/s001250100587
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DOI: https://doi.org/10.1007/s001250100587