title: Phosphorylation of serine 106 in Asef2 regulates cell migration and adhesion turnover.
abstract: Asef2, a 652-amino acid protein, is a guanine nucleotide exchange factor (GEF) that regulates cell migration and other processes via activation of Rho family GTPases, including Rac. Binding of the tumor suppressor adenomatous polyposis coli (APC) to Asef2 is known to induce its GEF activity; however, little is currently known about other modes of Asef2 regulation. Here, we investigated the role of phosphorylation in regulating Asef2 activity and function. Using high-resolution mass spectrometry (MS) and tandem mass spectrometry (MS/MS), we obtained complete coverage of all phosphorylatable residues and identified six phosphorylation sites. One of these, serine 106 (S106), was particularly intriguing as a potential regulator of Asef2 activity because of its location within the APC-binding domain. Interestingly, mutation of this serine to alanine (S106A), a non-phosphorylatable analogue, greatly diminished the ability of Asef2 to activate Rac, while a phosphomimetic mutation (serine to aspartic acid, S106D) enhanced Rac activation. Furthermore, expression of these mutants in HT1080 cells demonstrated that phosphorylation of S106 is critical for Asef2-promoted migration and for cell-matrix adhesion assembly and disassembly (adhesion turnover), which is a process that facilitates efficient migration. Collectively, our results show that phosphorylation of S106 modulates Asef2 GEF activity and Asef2-mediated cell migration and adhesion turnover.
authors.affiliation: Department of Biological Sciences and Vanderbilt Kennedy Center for Research on Human Development, ‡Department of Chemistry, §Vanderbilt Institute for Chemical Biology (VICB), ∥Vanderbilt Institute for Integrative Biosystems Research and Education (VIIBRE), ⊥Mass Spectrometry Research Center, #Department of Biochemistry, and ●Department of Cancer Biology, Vanderbilt University , Nashville, Tennessee 37235, United States.
authors.forename: J Corey
authors.initials: JC
authors.lastname: Evans
authors.forename: Kelly M
authors.initials: KM
authors.lastname: Hines
authors.forename: Jay G
authors.initials: JG
authors.lastname: Forsythe
authors.forename: Begum
authors.initials: B
authors.lastname: Erdogan
authors.forename: Mingjian
authors.initials: M
authors.lastname: Shi
authors.forename: Salisha
authors.initials: S
authors.lastname: Hill
authors.forename: Kristie L
authors.initials: KL
authors.lastname: Rose
authors.forename: John A
authors.initials: JA
authors.lastname: McLean
authors.forename: Donna J
authors.initials: DJ
authors.lastname: Webb
mesh_terms: D000595:Amino Acid Sequence; D002448:Cell Adhesion; D045744:Cell Line, Tumor; D002465:Cell Movement; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D012694:Serine / Q000737:chemistry
chemical_list: D020662:Guanine Nucleotide Exchange Factors; C525704:SPATA13 protein, human; D012694:Serine