title: Insulin receptor substrate-4 binds to Slingshot-1 phosphatase and promotes cofilin dephosphorylation.
abstract: Cofilin plays an essential role in cell migration and morphogenesis by enhancing actin filament dynamics via its actin filament-severing activity. Slingshot-1 (SSH1) is a protein phosphatase that plays a crucial role in regulating actin dynamics by dephosphorylating and reactivating cofilin. In this study, we identified insulin receptor substrate (IRS)-4 as a novel SSH1-binding protein. Co-precipitation assays revealed the direct endogenous binding of IRS4 to SSH1. IRS4, but not IRS1 or IRS2, was bound to SSH1. IRS4 was bound to SSH1 mainly through the unique region (amino acids 335-400) adjacent to the C terminus of the phosphotyrosine-binding domain of IRS4. The N-terminal A, B, and phosphatase domains of SSH1 were bound to IRS4 independently. Whereas in vitro phosphatase assays revealed that IRS4 does not directly affect the cofilin phosphatase activity of SSH1, knockdown of IRS4 increased cofilin phosphorylation in cultured cells. Knockdown of IRS4 decreased phosphatidylinositol 3-kinase (PI3K) activity, and treatment with an inhibitor of PI3K increased cofilin phosphorylation. Akt preferentially phosphorylated SSH1 at Thr-826, but expression of a non-phosphorylatable T826A mutant of SSH1 did not affect insulin-induced cofilin dephosphorylation, and an inhibitor of Akt did not increase cofilin phosphorylation. These results suggest that IRS4 promotes cofilin dephosphorylation through sequential activation of PI3K and SSH1 but not through Akt. In addition, IRS4 co-localized with SSH1 in F-actin-rich membrane protrusions in insulin-stimulated cells, which suggests that the association of IRS4 with SSH1 contributes to localized activation of cofilin in membrane protrusions.
authors.affiliation: Department of Biomolecular Sciences, Graduate School of Life Sciences, Tohoku University, Sendai, Miyagi 980-8578, Japan.
authors.forename: Yuta
authors.initials: Y
authors.lastname: Homma
authors.affiliation: Division of Dynamic Proteome in Cancer and Aging, Institute of Development, Aging and Cancer, Tohoku University, Sendai, Miyagi 980-8575, Japan, and.
authors.forename: Shin-Ichiro
authors.initials: SI
authors.lastname: Kanno
authors.affiliation: Department of Biomolecular Sciences, Graduate School of Life Sciences, Tohoku University, Sendai, Miyagi 980-8578, Japan.
authors.forename: Kazutaka
authors.initials: K
authors.lastname: Sasaki
authors.affiliation: Department of Biomolecular Sciences, Graduate School of Life Sciences, Tohoku University, Sendai, Miyagi 980-8578, Japan.
authors.forename: Michiru
authors.initials: M
authors.lastname: Nishita
authors.affiliation: Division of Dynamic Proteome in Cancer and Aging, Institute of Development, Aging and Cancer, Tohoku University, Sendai, Miyagi 980-8575, Japan, and.
authors.forename: Akira
authors.initials: A
authors.lastname: Yasui
authors.affiliation: Department of Medical Science, Graduate School of Medicine, University of Hiroshima, Hiroshima, Hiroshima 734-8553, Japan.
authors.forename: Tomoichiro
authors.initials: T
authors.lastname: Asano
authors.affiliation: Department of Biomolecular Sciences, Graduate School of Life Sciences, Tohoku University, Sendai, Miyagi 980-8578, Japan.
authors.forename: Kazumasa
authors.initials: K
authors.lastname: Ohashi
authors.affiliation: Department of Biomolecular Sciences, Graduate School of Life Sciences, Tohoku University, Sendai, Miyagi 980-8578, Japan,. Electronic address: kmizuno@biology.tohoku.ac.jp.
authors.forename: Kensaku
authors.initials: K
authors.lastname: Mizuno
mesh_terms: D022081:Cell Surface Extensions / Q000378:metabolism; D051338:Cofilin 1 / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D057809:HEK293 Cells; D006801:Humans; D007328:Insulin / Q000502:physiology; D055504:Insulin Receptor Substrate Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D011499:Protein Processing, Post-Translational; D021381:Protein Transport; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D015398:Signal Transduction
chemical_list: C495810:CFL1 protein, human; D051338:Cofilin 1; C107686:IRS4 protein, human; D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt; D010749:Phosphoprotein Phosphatases; C484329:SSH1 protein, human
keywords: Actin; Akt PKB; Cofilin; IRS; Insulin Receptor Substrate-4; Phosphatase; Phosphatidylinositide 3-Kinase (PI 3-Kinase); Slingshot
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