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Published on: March 8, 2017
Dynamic interaction between Src and C-terminal Src kinase in integrin alphaIIbbeta3-mediated signaling to the
Martin Vielreicher1, Gregory Harms2, Elke Butt3
1Institute of Clinical Biochemistry and Pathobiochemistry, University of Wurzburg, Wurzburg, D-97080, Germany; Molecular Microscopy Group, Rudolf-Virchow-Center, University of Wurzburg, D-97080 Wurzburg, Germany.
The Journal of Biological Chemistry
|September 15, 2007
Summary
C-terminal Src kinase (Csk) tightly controls Src activity, crucial for platelet adhesion. This study reveals dynamic Src-Csk interactions during cell adhesion, highlighting Csk
Area of Science:
- Cell Biology
- Biochemistry
- Molecular Biology
Background:
- Integrin alpha(IIb)beta(3) signaling is vital for platelet adhesion and thrombus formation, mediated by Src tyrosine kinase.
- Src activation at Tyr-418 occurs at lamellipodia leading edges, but its inactivation mechanism via C-terminal Src kinase (Csk) phosphorylation at Tyr-529 is less understood.
Purpose of the Study:
- To investigate the dynamic interactions between Src and Csk during alpha(IIb)beta(3)-mediated platelet adhesion.
- To elucidate the role of Csk-mediated Src phosphorylation at Tyr-529 in regulating Src activity and subsequent cell adhesion and spreading.
Main Methods:
- Live cell fluorescence resonance energy transfer (FRET) microscopy was used to visualize dynamic Src-Csk interactions in a platelet model cell line.
- Western blot analysis was employed to assess phosphorylation levels of Src at Tyr-418 and Tyr-529.
- Mutation analysis of Csk and Src was performed to determine their functional roles in cell adhesion.
Main Results:
- Highly dynamic Src-Csk interactions were observed at the leading edges of lamellipodia, with complexes reorganizing and forming focal adhesions during lamellipodia retraction.
- Increased Csk-mediated Src Tyr-529 phosphorylation correlated with decreased Src Tyr-418 phosphorylation, indicating Src inactivation.
- While Csk inactivation had no effect, constitutively active Src (SrcY529F) severely impaired cell adhesion and spreading, resulting in disorganized focal adhesions.
Conclusions:
- Tight regulation of Src by Csk through Tyr-529 phosphorylation is essential for proper cell adhesion and spreading.
- The study introduces a novel FRET approach for studying kinase inactivation, applicable to other integrin and Src family kinase signaling pathways.
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