title: New insights into FAK phosphorylation based on a FAT domain-defective mutation.
abstract: Mounting evidence suggests that the FAK N-terminal (FERM) domain controls FAK phosphorylation and function; however, little is known regarding the role of the C terminal (FAT) domain in FAK regulation. We identified a patient-derived FAK mutant, in which a 27-amino acid segment was deleted from the C-terminal FAT domain (named FAK-Del33). When FAK-Del33 was overexpressed in specific tumor cell lines, Y397 phosphorylation increased compared with that observed in cells expressing FAK-WT. Here, we attempt to unveil the mechanism of this increased phosphorylation. Using cell biology experiments, we show that FAK-Del33 is incapable of co-localizing with paxillin, and has constitutively high Y397 phosphorylation. With a kinase-dead mutation, it showed phosphorylation of FAK-Del33 has enhanced through auto-phosphorylation. It was also demonstrated that phosphorylation of FAK-Del33 is not Src dependent or enhanced intermolecular interactions, and that the hyperphosphorylation can be lowered using increasing amounts of transfected FERM domain. This result suggests that Del33 mutation disrupting of FAT's structural integrity and paxillin binding capacity leads to incapable of targeting Focal adhesions, but has gained the capacity for auto-phosphorylation in cis.
authors.affiliation: Department of Clinical Laboratory, Ruijin North Hospital, Ruijin Hospital, Shanghai JiaoTong University School of Medicine, Shanghai, P. R. China.
authors.forename: Xuqian
authors.initials: X
authors.lastname: Fang
authors.affiliation: Faculty of Medical Laboratory Science, Shanghai JiaoTong University School of Medicine, Shanghai, P. R. China.
authors.forename: Xiangfan
authors.initials: X
authors.lastname: Liu
authors.affiliation: Department of Biochemistry and Molecular Biology, Shanghai JiaoTong University School of Medicine, Shanghai, P. R. China.
authors.forename: Ling
authors.initials: L
authors.lastname: Yao
authors.affiliation: Department of Clinical Laboratory, Ruijin North Hospital, Ruijin Hospital, Shanghai JiaoTong University School of Medicine, Shanghai, P. R. China.
authors.forename: Changqiang
authors.initials: C
authors.lastname: Chen
authors.affiliation: Department of Clinical Laboratory, Ruijin North Hospital, Ruijin Hospital, Shanghai JiaoTong University School of Medicine, Shanghai, P. R. China.
authors.forename: Jiafei
authors.initials: J
authors.lastname: Lin
authors.affiliation: Faculty of Medical Laboratory Science, Shanghai JiaoTong University School of Medicine, Shanghai, P. R. China.
authors.forename: Peihua
authors.initials: P
authors.lastname: Ni
authors.affiliation: Department of Biochemistry and Molecular Biology, Shanghai JiaoTong University School of Medicine, Shanghai, P. R. China; Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York, United States of America.
authors.forename: Xinmin
authors.initials: X
authors.lastname: Zheng
authors.affiliation: Department of Clinical Laboratory, Ruijin North Hospital, Ruijin Hospital, Shanghai JiaoTong University School of Medicine, Shanghai, P. R. China.
authors.forename: Qishi
authors.initials: Q
authors.lastname: Fan
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002448:Cell Adhesion; D002461:Cell Line, Transformed; D045744:Cell Line, Tumor; D051416:Focal Adhesion Protein-Tyrosine Kinases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D051419:Paxillin / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs / Q000235:genetics*; D017433:Protein Structure, Secondary; D021381:Protein Transport; D016415:Sequence Alignment; D019061:src-Family Kinases / Q000378:metabolism
chemical_list: D051419:Paxillin; D051416:Focal Adhesion Protein-Tyrosine Kinases; D019061:src-Family Kinases