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Regulation of insulin receptor substrate 1 pleckstrin homology domain by protein kinase C: role of serine 24 phosphorylation

Regulation of insulin receptor substrate 1 pleckstrin homology domain by protein kinase C: role of serine 24 phosphorylation
Regulation of insulin receptor substrate 1 pleckstrin homology domain by protein kinase C: role of serine 24 phosphorylation

Phosphorylation of insulin receptor substrate (IRS) proteins on serine residues is an important post-translational modification that is linked to insulin resistance. Several phosphoserine sites on IRS1 have been identified; the majority are located proximal to the phosphotryosine-binding domain or near key receptor tyrosine kinase substrate and/or Src-homology 2 domain-binding sites. Here we report on the characterization of a serine phosphorylation site in the N-terminal pleckstrin homology (PH) domain of IRS1. Bioinformatic tools identify serine 24 (Ser24) as a putative substrate site for the protein kinase C (PKC) family of serine kinases. We demonstrate that this site is indeed a bona fide substrate for conventional PKC. In vivo, IRS-1 is also phosphorylated on Ser24 after phorbol 12-myristate 13-acetate treatment of cells, and isoform-selective inhibitor studies suggest the involvement of PKCα. By comparing the pharmacological characteristics of phorbol 12-myristate 13-acetate-stimulated Ser24 phosphorylation with phosphorylation at two other sites previously linked to PKC activity (Ser307 and Ser612), we show that PKCα is likely to be directly involved in Ser24 phosphorylation, but indirectly involved in Ser307 and Ser612 phosphorylation. Using Ser24Asp IRS-1 mutants to mimic the phosphorylated residue, we demonstrate that the phosphorylation status of Ser24 does play an important role in regulating phosphoinositide binding to, and the intracellular localization of, the IRS1-PH domain, which can ultimately impinge on insulin-stimulated glucose uptake. Hence we provide evidence that IRS1-PH domain function is important for normal insulin signaling and is regulated by serine phosphorylation in a manner that could contribute to insulin resistance.

0888-8809
1838-1852
Nawaratne, Ranmali
856a362a-2829-4e57-b1c5-c1b81497eb27
Gray, Alexander
a908261e-9c70-4f81-aea0-1674f6e587a7
Jørgensen, Christina H.
7298d16c-9f9c-405e-8d90-c7cf8106576f
Downes, C. Peter
5ce6b445-b02e-446e-ae10-bb51f51b8fa7
Siddle, Kenneth
1af6301b-41be-4982-8857-9f4bfa4b1eaa
Sethi, Jaswinder K.
923f1a81-91e4-46cd-8853-bb4a979f5a85
Nawaratne, Ranmali
856a362a-2829-4e57-b1c5-c1b81497eb27
Gray, Alexander
a908261e-9c70-4f81-aea0-1674f6e587a7
Jørgensen, Christina H.
7298d16c-9f9c-405e-8d90-c7cf8106576f
Downes, C. Peter
5ce6b445-b02e-446e-ae10-bb51f51b8fa7
Siddle, Kenneth
1af6301b-41be-4982-8857-9f4bfa4b1eaa
Sethi, Jaswinder K.
923f1a81-91e4-46cd-8853-bb4a979f5a85

Nawaratne, Ranmali, Gray, Alexander, Jørgensen, Christina H., Downes, C. Peter, Siddle, Kenneth and Sethi, Jaswinder K. (2006) Regulation of insulin receptor substrate 1 pleckstrin homology domain by protein kinase C: role of serine 24 phosphorylation. Molecular endocrinology, 20 (8), 1838-1852. (doi:10.1210/me.2005-0536).

Record type: Article

Abstract

Phosphorylation of insulin receptor substrate (IRS) proteins on serine residues is an important post-translational modification that is linked to insulin resistance. Several phosphoserine sites on IRS1 have been identified; the majority are located proximal to the phosphotryosine-binding domain or near key receptor tyrosine kinase substrate and/or Src-homology 2 domain-binding sites. Here we report on the characterization of a serine phosphorylation site in the N-terminal pleckstrin homology (PH) domain of IRS1. Bioinformatic tools identify serine 24 (Ser24) as a putative substrate site for the protein kinase C (PKC) family of serine kinases. We demonstrate that this site is indeed a bona fide substrate for conventional PKC. In vivo, IRS-1 is also phosphorylated on Ser24 after phorbol 12-myristate 13-acetate treatment of cells, and isoform-selective inhibitor studies suggest the involvement of PKCα. By comparing the pharmacological characteristics of phorbol 12-myristate 13-acetate-stimulated Ser24 phosphorylation with phosphorylation at two other sites previously linked to PKC activity (Ser307 and Ser612), we show that PKCα is likely to be directly involved in Ser24 phosphorylation, but indirectly involved in Ser307 and Ser612 phosphorylation. Using Ser24Asp IRS-1 mutants to mimic the phosphorylated residue, we demonstrate that the phosphorylation status of Ser24 does play an important role in regulating phosphoinositide binding to, and the intracellular localization of, the IRS1-PH domain, which can ultimately impinge on insulin-stimulated glucose uptake. Hence we provide evidence that IRS1-PH domain function is important for normal insulin signaling and is regulated by serine phosphorylation in a manner that could contribute to insulin resistance.

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Published date: 1 August 2006

Identifiers

Local EPrints ID: 415422
URI: https://eprints.soton.ac.uk/id/eprint/415422
ISSN: 0888-8809
PURE UUID: b084e22c-209b-476b-b727-bc7197a407f0
ORCID for Jaswinder K. Sethi: ORCID iD orcid.org/0000-0003-4157-0475

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Date deposited: 09 Nov 2017 17:30
Last modified: 31 Jul 2019 00:25

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