title: A phosphoproteomic screen demonstrates differential dependence on HER3 for MAP kinase pathway activation by distinct PIK3CA mutations.
abstract: The PIK3CA gene encodes for the p110 alpha isoform of PI3 kinase and is one of the most frequently mutated oncogenes in human cancers. However, the mechanisms by which PIK3CA mutations activate cell signaling are not fully understood. Here we used a phosphoproteomic approach to compare differential phosphorylation patterns between human breast epithelial cells and two isogenic somatic cell knock in derivatives, each harboring a distinct PIK3CA mutation. We demonstrated differential phosphorylation patterns between isogenic cell lines containing a PIK3CA helical domain mutation (E545K) compared to cells with a PIK3CA kinase domain mutation (H1047R). In particular, the receptor tyrosine kinase, HER3, showed increased phosphorylation at tyrosine 1328 in H1047R cells versus E545K cells. Genetic studies using shRNA demonstrated that H1047R cells have a profound decrease in growth factor independent proliferation upon HER3 knock down, but this effect was attenuated in E545K cells. In addition, HER3 knock down led to reductions in both PI3 kinase and MAP kinase pathway activation in H1047R cells, but in E545K cells only PI3 kinase pathway diminution was observed. These studies demonstrate the power of using paired isogenic cell lines for proteomic analysis to gain new insights into oncogenic signal transduction pathways.
authors.affiliation: The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins University School of Medicine, Baltimore, MD, USA.
authors.forename: Brian G
authors.initials: BG
authors.lastname: Blair
authors.forename: Xinyan
authors.initials: X
authors.lastname: Wu
authors.forename: Muhammad Saddiq
authors.initials: MS
authors.lastname: Zahari
authors.forename: Morassa
authors.initials: M
authors.lastname: Mohseni
authors.forename: Justin
authors.initials: J
authors.lastname: Cidado
authors.forename: Hong Yuen
authors.initials: HY
authors.lastname: Wong
authors.forename: Julia A
authors.initials: JA
authors.lastname: Beaver
authors.forename: Rory L
authors.initials: RL
authors.lastname: Cochran
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Zabransky
authors.forename: Sarah
authors.initials: S
authors.lastname: Croessmann
authors.forename: David
authors.initials: D
authors.lastname: Chu
authors.forename: Patricia Valda
authors.initials: PV
authors.lastname: Toro
authors.forename: Karen
authors.initials: K
authors.lastname: Cravero
authors.forename: Akhilesh
authors.initials: A
authors.lastname: Pandey
authors.forename: Ben Ho
authors.initials: BH
authors.lastname: Park
mesh_terms: D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D058534:Class I Phosphatidylinositol 3-Kinases; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009154:Mutation; D019869:Phosphatidylinositol 3-Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D017434:Protein Structure, Tertiary; D040901:Proteomics; D034622:RNA Interference; D020893:Receptor, ErbB-3 / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction
chemical_list: D019869:Phosphatidylinositol 3-Kinases; D058534:Class I Phosphatidylinositol 3-Kinases; C484760:PIK3CA protein, human; C581292:ERBB3 protein, human; D020893:Receptor, ErbB-3; D020928:Mitogen-Activated Protein Kinases
keywords: Biomedicine; HER3; MAP kinase; PI3 kinase; PIK3CA