title: Uncoupling JAK2 V617F activation from cytokine-induced signalling by modulation of JH2 αC helix.
abstract: The mechanisms by which JAK2 is activated by the prevalent pseudokinase (JH2) V617F mutation in blood cancers remain elusive. Via structure-guided mutagenesis and transcriptional and functional assays, we identify a community of residues from the JH2 helix αC, SH2-JH2 linker and JH1 kinase domain that mediate V617F-induced activation. This circuit is broken by altering the charge of residues along the solvent-exposed face of the JH2 αC, which is predicted to interact with the SH2-JH2 linker and JH1. Mutations that remove negative charges or add positive charges, such as E596A/R, do not alter the JH2 V617F fold, as shown by the crystal structure of JH2 V617F E596A. Instead, they prevent kinase domain activation via modulation of the C-terminal residues of the SH2-JH2 linker. These results suggest strategies for selective V617F JAK2 inhibition, with preservation of wild-type function.
authors.affiliation: Ludwig Institute for Cancer Research, 1200 Brussels, Belgium de Duve Institute, Université catholique de Louvain, 1200 Brussels, Belgium.
authors.forename: Emilie
authors.initials: E
authors.lastname: Leroy
authors.affiliation: Ludwig Institute for Cancer Research, 1200 Brussels, Belgium de Duve Institute, Université catholique de Louvain, 1200 Brussels, Belgium.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Dusa
authors.affiliation: Ludwig Institute for Cancer Research, 1200 Brussels, Belgium.
authors.forename: Didier
authors.initials: D
authors.lastname: Colau
authors.affiliation: Small Molecule Discovery Program, Ludwig Institute for Cancer Research, La Jolla, CA 92093, U.S.A.
authors.forename: Amir
authors.initials: A
authors.lastname: Motamedi
authors.affiliation: Ludwig Institute for Cancer Research, 1200 Brussels, Belgium de Duve Institute, Université catholique de Louvain, 1200 Brussels, Belgium.
authors.forename: Xavier
authors.initials: X
authors.lastname: Cahu
authors.affiliation: Ludwig Institute for Cancer Research, 1200 Brussels, Belgium de Duve Institute, Université catholique de Louvain, 1200 Brussels, Belgium.
authors.forename: Céline
authors.initials: C
authors.lastname: Mouton
authors.affiliation: University of Texas Southwestern Medical Center, Dallas, TX 5323, U.S.A.
authors.forename: Lily J
authors.initials: LJ
authors.lastname: Huang
authors.affiliation: Small Molecule Discovery Program, Ludwig Institute for Cancer Research, La Jolla, CA 92093, U.S.A.
authors.forename: Andrew K
authors.initials: AK
authors.lastname: Shiau
authors.affiliation: Ludwig Institute for Cancer Research, 1200 Brussels, Belgium de Duve Institute, Université catholique de Louvain, 1200 Brussels, Belgium stefan.constantinescu@bru.licr.org.
authors.forename: Stefan N
authors.initials: SN
authors.lastname: Constantinescu
mesh_terms: D015153:Blotting, Western; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects / Q000235:genetics; D016207:Cytokines / Q000494:pharmacology*; D005434:Flow Cytometry; D006801:Humans; D053614:Janus Kinase 2 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D009154:Mutation / Q000235:genetics; D010766:Phosphorylation / Q000187:drug effects; D000072417:Protein Domains / Q000235:genetics / Q000502:physiology
chemical_list: D016207:Cytokines; C507924:JAK2 protein, human; D053614:Janus Kinase 2
keywords: JAK-STAT signalling; JAK2 V617F; inhibitor; kinase; myeloproliferative neoplasm (MPN)