title: RET Functions as a Dual-Specificity Kinase that Requires Allosteric Inputs from Juxtamembrane Elements.
abstract: Receptor tyrosine kinases exhibit a variety of activation mechanisms despite highly homologous catalytic domains. Such diversity arises through coupling of extracellular ligand-binding portions with highly variable intracellular sequences flanking the tyrosine kinase domain and specific patterns of autophosphorylation sites. Here, we show that the juxtamembrane (JM) segment enhances RET catalytic domain activity through Y687. This phospho-site is also required by the JM region to rescue an otherwise catalytically deficient RET activation-loop mutant lacking tyrosines. Structure-function analyses identified interactions between the JM hinge, αC helix, and an unconventional activation-loop serine phosphorylation site that engages the HRD motif and promotes phospho-tyrosine conformational accessibility and regulatory spine assembly. We demonstrate that this phospho-S909 arises from an intrinsic RET dual-specificity kinase activity and show that an equivalent serine is required for RET signaling in Drosophila. Our findings reveal dual-specificity and allosteric components for the mechanism of RET activation and signaling with direct implications for drug discovery.
authors.affiliation: Structural Biology Laboratory, The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK. Electronic address: ivan.plazamenacho@unibas.ch.
authors.forename: Iván
authors.initials: I
authors.lastname: Plaza-Menacho
authors.affiliation: Protein Analysis and Proteomics, The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK.
authors.forename: Karin
authors.initials: K
authors.lastname: Barnouin
authors.affiliation: The Breast Cancer Now Toby Robins Research Centre, Mary-Jean Mitchell Green Building, Institute of Cancer Research, SW3 6JB London, UK.
authors.forename: Rachael
authors.initials: R
authors.lastname: Barry
authors.affiliation: Protein Production Facility, The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK.
authors.forename: Annabel
authors.initials: A
authors.lastname: Borg
authors.affiliation: The Breast Cancer Now Toby Robins Research Centre, Mary-Jean Mitchell Green Building, Institute of Cancer Research, SW3 6JB London, UK.
authors.forename: Mariam
authors.initials: M
authors.lastname: Orme
authors.affiliation: Structural Biology Laboratory, The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK.
authors.forename: Rakhee
authors.initials: R
authors.lastname: Chauhan
authors.affiliation: Structural Biology Science Technology Platform, The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK.
authors.forename: Stephane
authors.initials: S
authors.lastname: Mouilleron
authors.affiliation: Structural Biology Laboratory, The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK.
authors.forename: Rubén J
authors.initials: RJ
authors.lastname: Martínez-Torres
authors.affiliation: The Breast Cancer Now Toby Robins Research Centre, Mary-Jean Mitchell Green Building, Institute of Cancer Research, SW3 6JB London, UK.
authors.forename: Pascal
authors.initials: P
authors.lastname: Meier
authors.affiliation: Structural Biology Laboratory, The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK; Department of Biological Sciences, Institute of Structural and Molecular Biology, Birkbeck College, Malet Street, WC1E 7HX London, UK. Electronic address: neil.mcdonald@crick.ac.uk.
authors.forename: Neil Q
authors.initials: NQ
authors.lastname: McDonald
mesh_terms: D000494:Allosteric Regulation / Q000235:genetics; D000595:Amino Acid Sequence / Q000235:genetics; D000818:Animals; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D029721:Drosophila Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004331:Drosophila melanogaster; D004789:Enzyme Activation / Q000235:genetics; D010766:Phosphorylation; D051096:Proto-Oncogene Proteins c-ret / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D020794:Receptor Protein-Tyrosine Kinases / Q000737:chemistry* / Q000235:genetics; D012694:Serine / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics; D013329:Structure-Activity Relationship
chemical_list: D029721:Drosophila Proteins; D012694:Serine; D051096:Proto-Oncogene Proteins c-ret; D020794:Receptor Protein-Tyrosine Kinases; C416649:Ret protein, Drosophila
keywords: Drosophila; RTK; dual-specificity; oncogene; phosphorylation; receptor tyrosine kinase; signaling; structure-function