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v-Src SH3-enhanced interaction with focal adhesion kinase at beta 1 integrin-containing invadopodia promotes cell
Christof R Hauck1, Datsun A Hsia, Dusko Ilic
1Department of Immunology, The Scripps Research Institute, 10550 N. Torrey Pines Road, La Jolla, CA 92037, USA.
Abstract:
In viral Src (v-Src)-transformed cells, focal adhesion kinase (FAK) associates with v-Src by combined v-Src SH2 and gain-of-function v-Src SH3 domain binding to FAK. Here we assess the significance of the Arg-95 to Trp gain-of-function mutation in the v-Src SH3 domain through comparisons of Src-/- fibroblasts transformed with either Prague C v-Src or a point mutant (v-Src-RT) containing a normal (Arg-95) SH3 domain. Both v-Src isoforms exhibited equivalent kinase activity, enhanced Src-/- cell motility, and stimulated cell growth in both low serum and soft agar. The stability of a v-Src-RT.FAK signaling complex and FAK phosphorylation at Tyr-861 and Tyr-925 were reduced in v-Src-RT- compared with v-Src-transformed cells. v-Src but not v-Src-RT promoted Src-/- cell invasion through a reconstituted Matrigel basement membrane barrier and v-Src co-localized with FAK and beta(1) integrin at invadopodia. In contrast, v-Src-RT exhibited a partial perinuclear and focal contact distribution in Src-/- cells. Adenovirus-mediated FAK overexpression promoted v-Src-RT recruitment to invadopodia, the formation of a v-Src-RT.FAK signaling complex, and reversed the v-Src-RT invasion deficit. Adenovirus-mediated inhibition of FAK blocked v-Src-stimulated cell invasion. These studies establish that gain-of-function v-Src SH3 targeting interactions with FAK at beta(1) integrin-containing invadopodia act to stabilize a v-Src.FAK signaling complex promoting cell invasion.
Insights
Viral Src (v-Src) SH3 domain interactions with focal adhesion kinase (FAK) are crucial for cell invasion. Specific mutations disrupt this complex, impairing invasion, but FAK re-expression restores it, highlighting FAK
Area of Science:
- Cell Biology
- Molecular Oncology
- Virology
Background:
- Viral Src (v-Src) protein kinase is a potent oncogene.
- Focal Adhesion Kinase (FAK) is a key regulator of cell motility and invasion.
- v-Src and FAK form signaling complexes that influence cell behavior.
Purpose of the Study:
- To investigate the role of the v-Src SH3 domain, specifically the Arg-95 to Trp mutation, in v-Src-mediated cell invasion.
- To determine the significance of v-Src and FAK complex stability in invadopodia for cell invasion.
- To elucidate the contribution of FAK to v-Src-induced cell invasion.
Main Methods:
- Comparison of wild-type v-Src and a v-Src SH3 domain mutant (v-Src-RT) in Src-/- fibroblasts.
- Assessment of cell motility, growth, invasion through Matrigel, and signaling complex formation.
- Utilizing adenovirus-mediated FAK overexpression and inhibition to study FAK's role.
Main Results:
- The v-Src SH3 domain mutation (v-Src-RT) reduced v-Src-FAK complex stability and FAK phosphorylation.
- v-Src, but not v-Src-RT, promoted cell invasion and co-localized with FAK and β1 integrin at invadopodia.
- FAK overexpression rescued the invasion defect of v-Src-RT, while FAK inhibition blocked v-Src-induced invasion.
Conclusions:
- Gain-of-function v-Src SH3 domain interactions with FAK at β1 integrin-containing invadopodia stabilize the v-Src-FAK complex.
- This stabilized complex is essential for promoting v-Src-mediated cell invasion.
- Targeting these specific interactions could offer therapeutic strategies against v-Src-driven cancers.