title: Protein interaction network of the mammalian Hippo pathway reveals mechanisms of kinase-phosphatase interactions.
abstract: The Hippo pathway regulates organ size and tissue homeostasis in response to multiple stimuli, including cell density and mechanotransduction. Pharmacological inhibition of phosphatases can also stimulate Hippo signaling in cell culture. We defined the Hippo protein-protein interaction network with and without inhibition of serine and threonine phosphatases by okadaic acid. We identified 749 protein interactions, including 599 previously unrecognized interactions, and demonstrated that several interactions with serine and threonine phosphatases were phosphorylation-dependent. Mutation of the T-loop of MST2 (mammalian STE20-like protein kinase 2), which prevented autophosphorylation, disrupted its association with STRIPAK (striatin-interacting phosphatase and kinase complex). Deletion of the amino-terminal forkhead-associated domain of SLMAP (sarcolemmal membrane-associated protein), a component of the STRIPAK complex, prevented its association with MST1 and MST2. Phosphatase inhibition produced temporally distinct changes in proteins that interacted with MOB1A and MOB1B (Mps one binder kinase activator-like 1A and 1B) and promoted interactions with upstream Hippo pathway proteins, such as MST1 and MST2, and with the trimeric protein phosphatase 6 complex (PP6). Mutation of three basic amino acids that are part of a phospho-serine- and phospho-threonine-binding domain in human MOB1B prevented its interaction with MST1 and PP6 in cells treated with okadaic acid. Collectively, our results indicated that changes in phosphorylation orchestrate interactions between kinases and phosphatases in Hippo signaling, providing a putative mechanism for pathway regulation.
authors.affiliation: 1Centre for Systems Biology, Lunenfeld-Tanenbaum Research Institute at Mount Sinai Hospital, Toronto, Ontario M5G 1X5, Canada.
authors.forename: Amber L
authors.initials: AL
authors.lastname: Couzens
authors.forename: James D R
authors.initials: JD
authors.lastname: Knight
authors.forename: Michelle J
authors.initials: MJ
authors.lastname: Kean
authors.forename: Guoci
authors.initials: G
authors.lastname: Teo
authors.forename: Alexander
authors.initials: A
authors.lastname: Weiss
authors.forename: Wade H
authors.initials: WH
authors.lastname: Dunham
authors.forename: Zhen-Yuan
authors.initials: ZY
authors.lastname: Lin
authors.forename: Richard D
authors.initials: RD
authors.lastname: Bagshaw
authors.forename: Frank
authors.initials: F
authors.lastname: Sicheri
authors.forename: Tony
authors.initials: T
authors.lastname: Pawson
authors.forename: Jeffrey L
authors.initials: JL
authors.lastname: Wrana
authors.forename: Hyungwon
authors.initials: H
authors.lastname: Choi
authors.forename: Anne-Claude
authors.initials: AC
authors.lastname: Gingras
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D015153:Blotting, Western; D016000:Cluster Analysis; D057809:HEK293 Cells; D006367:HeLa Cells; D000090683:Hippo Signaling Pathway; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D010749:Phosphoprotein Phosphatases / Q000235:genetics / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D025941:Protein Interaction Mapping / Q000379:methods; D060066:Protein Interaction Maps; D017346:Protein Serine-Threonine Kinases / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D000090742:Serine-Threonine Kinase 3; D015398:Signal Transduction; D000099317:Hippo Kinases
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D047908:Intracellular Signaling Peptides and Proteins; D008156:Luciferases; D008164:Luminescent Proteins; D008565:Membrane Proteins; D010749:Phosphoprotein Phosphatases; D010744:Phosphoric Monoester Hydrolases; D017346:Protein Serine-Threonine Kinases; D000090742:Serine-Threonine Kinase 3; C496256:MOB1A protein, human; C496255:MOB1B protein, human; C109742:SLMAP protein, human; C095566:STK4 protein, human; D000099317:Hippo Kinases; C097835:STK3 protein, human; C106079:protein phosphatase 6