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Regulation of the interaction between PIPKI gamma and talin by proline-directed protein kinases.

Lee SY, Voronov S, Letinic K, Nairn AC, Di Paolo G, De Camilli P - J. Cell Biol. (2005)

Bottom Line: Cell Biol. 163:1339-1349).We find that Y649 phosphorylation does not stimulate directly PIPKI gamma binding to talin, but may do so indirectly by inhibiting S650 phosphorylation.Conversely, S650 phosphorylation inhibits Y649 phosphorylation by Src.

View Article: PubMed Central - PubMed

Affiliation: Howard Hughes Medical Institute, Yale University School of Medicine, New Haven, CT 06510, USA.

ABSTRACT
The interaction of talin with phosphatidylinositol(4) phosphate 5 kinase type I gamma (PIPKI gamma) regulates PI(4,5)P2 synthesis at synapses and at focal adhesions. Here, we show that phosphorylation of serine 650 (S650) within the talin-binding sequence of human PIPKI gamma blocks this interaction. At synapses, S650 is phosphorylated by p35/Cdk5 and mitogen-activated protein kinase at rest, and dephosphorylated by calcineurin upon stimulation. S650 is also a substrate for cyclin B1/Cdk1 and its phosphorylation in mitosis correlates with focal adhesion disassembly. Phosphorylation by Src of the tyrosine adjacent to S650 (Y649 in human PIPKI gamma) was shown to enhance PIPKI gamma targeting to focal adhesions (Ling, K., R.L. Doughman, V.V. Iyer, A.J. Firestone, S.F. Bairstow, D.F. Mosher, M.D. Schaller, and R.A. Anderson. 2003. J. Cell Biol. 163:1339-1349). We find that Y649 phosphorylation does not stimulate directly PIPKI gamma binding to talin, but may do so indirectly by inhibiting S650 phosphorylation. Conversely, S650 phosphorylation inhibits Y649 phosphorylation by Src. The opposite effects of the phosphorylation of Y649 and S650 likely play a critical role in regulating synaptic function as well as the balance between cell adhesion and cell motility.

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Inhibitory effects of p35/Cdk5 overexpression on focal adhesion. NIH3T3 cells were cotransfected with both p35 and Cdk5 for 24 h, and the overexpressed proteins were detected by immunofluorescence microscopy with the antibodies indicated. Focal adhesions were visualized by antivinculin immunostaining, and actin was visualized by fluorescent phalloidin. Bar, 10 μm.
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fig5: Inhibitory effects of p35/Cdk5 overexpression on focal adhesion. NIH3T3 cells were cotransfected with both p35 and Cdk5 for 24 h, and the overexpressed proteins were detected by immunofluorescence microscopy with the antibodies indicated. Focal adhesions were visualized by antivinculin immunostaining, and actin was visualized by fluorescent phalloidin. Bar, 10 μm.

Mentions: If the binding of PIPKIγ90 to talin is important for focal adhesion dynamics, overexpression of p35/Cdk5 would be expected to affect focal adhesion dynamics by enhancing the phosphorylation state of S650. Consistent with this prediction, overexpression of p35/Cdk5 in NIH3T3 cells disrupted focal adhesions and stress fibers, as detected by antivinculin immunostaining and phalloidin staining, respectively (Fig. 5).


Regulation of the interaction between PIPKI gamma and talin by proline-directed protein kinases.

Lee SY, Voronov S, Letinic K, Nairn AC, Di Paolo G, De Camilli P - J. Cell Biol. (2005)

Inhibitory effects of p35/Cdk5 overexpression on focal adhesion. NIH3T3 cells were cotransfected with both p35 and Cdk5 for 24 h, and the overexpressed proteins were detected by immunofluorescence microscopy with the antibodies indicated. Focal adhesions were visualized by antivinculin immunostaining, and actin was visualized by fluorescent phalloidin. Bar, 10 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171813&req=5

fig5: Inhibitory effects of p35/Cdk5 overexpression on focal adhesion. NIH3T3 cells were cotransfected with both p35 and Cdk5 for 24 h, and the overexpressed proteins were detected by immunofluorescence microscopy with the antibodies indicated. Focal adhesions were visualized by antivinculin immunostaining, and actin was visualized by fluorescent phalloidin. Bar, 10 μm.
Mentions: If the binding of PIPKIγ90 to talin is important for focal adhesion dynamics, overexpression of p35/Cdk5 would be expected to affect focal adhesion dynamics by enhancing the phosphorylation state of S650. Consistent with this prediction, overexpression of p35/Cdk5 in NIH3T3 cells disrupted focal adhesions and stress fibers, as detected by antivinculin immunostaining and phalloidin staining, respectively (Fig. 5).

Bottom Line: Cell Biol. 163:1339-1349).We find that Y649 phosphorylation does not stimulate directly PIPKI gamma binding to talin, but may do so indirectly by inhibiting S650 phosphorylation.Conversely, S650 phosphorylation inhibits Y649 phosphorylation by Src.

View Article: PubMed Central - PubMed

Affiliation: Howard Hughes Medical Institute, Yale University School of Medicine, New Haven, CT 06510, USA.

ABSTRACT
The interaction of talin with phosphatidylinositol(4) phosphate 5 kinase type I gamma (PIPKI gamma) regulates PI(4,5)P2 synthesis at synapses and at focal adhesions. Here, we show that phosphorylation of serine 650 (S650) within the talin-binding sequence of human PIPKI gamma blocks this interaction. At synapses, S650 is phosphorylated by p35/Cdk5 and mitogen-activated protein kinase at rest, and dephosphorylated by calcineurin upon stimulation. S650 is also a substrate for cyclin B1/Cdk1 and its phosphorylation in mitosis correlates with focal adhesion disassembly. Phosphorylation by Src of the tyrosine adjacent to S650 (Y649 in human PIPKI gamma) was shown to enhance PIPKI gamma targeting to focal adhesions (Ling, K., R.L. Doughman, V.V. Iyer, A.J. Firestone, S.F. Bairstow, D.F. Mosher, M.D. Schaller, and R.A. Anderson. 2003. J. Cell Biol. 163:1339-1349). We find that Y649 phosphorylation does not stimulate directly PIPKI gamma binding to talin, but may do so indirectly by inhibiting S650 phosphorylation. Conversely, S650 phosphorylation inhibits Y649 phosphorylation by Src. The opposite effects of the phosphorylation of Y649 and S650 likely play a critical role in regulating synaptic function as well as the balance between cell adhesion and cell motility.

Show MeSH
Related in: MedlinePlus