Regulation of the Src kinase-associated phosphoprotein 55 homologue by the protein tyrosine phosphatase PTP-PEST in the control of cell motility.

Abstract

PTP-PEST is a cytosolic ubiquitous protein tyrosine phosphatase (PTP) that contains, in addition to its catalytic domain, several protein-protein interaction domains that allow it to interface with several signaling pathways. Among others, PTP-PEST is a key regulator of cellular motility and cytoskeleton dynamics. The complexity of the PTP-PEST interactome underscores the necessity to identify its interacting partners and physiological substrates in order to further understand its role in focal adhesion complex turnover and actin organization. Using a modified yeast substrate trapping two-hybrid system, we identified a cytosolic adaptor protein named Src kinase-associated phosphoprotein 55 homologue (SKAP-Hom) as a novel substrate of PTP-PEST. To confirm PTP-PEST interaction with SKAP-Hom, in vitro pull down assays were performed demonstrating that the PTP catalytic domain and Proline-rich 1 (P1) domain are respectively binding to the SKAP-Hom Y260 and Y297 residues and its SH3 domain. Subsequently, we generated and rescued SKAP-Hom-deficient mouse embryonic fibroblasts (MEFs) with WT SKAP-Hom, SKAP-Hom tyrosine mutants (Y260F, Y260F/Y297F), or SKAP-Hom SH3 domain mutant (W335K). Given the role of PTP-PEST, wound-healing and trans-well migration assays were performed using the generated lines. Indeed, SKAP-Hom-deficient MEFs showed a defect in migration compared with WT-rescued MEFs. Interestingly, the SH3 domain mutant-rescued MEFs showed an enhanced cell migration corresponding potentially with higher tyrosine phosphorylation levels of SKAP-Hom. These findings suggest a novel role of SKAP-Hom and its phosphorylation in the regulation of cellular motility. Moreover, these results open new avenues by which PTP-PEST regulates cellular migration, a hallmark of metastasis.

• Authors: Emily Ayoub, Anita Hall, Adam M Scott, Mélanie J Chagnon, Géraldine Miquel, Maxime Hallé, Masaharu Noda, Andreas Bikfalvi, Michel L Tremblay
• Journal: The Journal of biological chemistry (J Biol Chem) Vol. 288 Issue 36 Pg. 25739-25748 (Sep 06 2013) ISSN: 1083-351X [Electronic] United States
• PMID: 23897807 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Intracellular Signaling Peptides and Proteins
• src kinase associated phosphoprotein 2
• Protein Tyrosine Phosphatase, Non-Receptor Type 12
• Ptpn12 protein, mouse

Topics

• Amino Acid Substitution
• Animals
• Cell Movement (physiology)
• Cells, Cultured
• Embryo, Mammalian (cytology, metabolism)
• Fibroblasts (cytology, metabolism)
• Intracellular Signaling Peptides and Proteins (genetics, metabolism)
• Mice, Knockout
• Mutation, Missense
• Protein Tyrosine Phosphatase, Non-Receptor Type 12 (genetics, metabolism)
• Two-Hybrid System Techniques
• src Homology Domains