Substitution of pseudokinase domain residue Val-617 by large non-polar amino acids causes activation of JAK2.
Explaining the uniqueness of the acquired somatic JAK2 V617F mutation, which is present in more than 95% of polycythemia vera patients, has been a challenge. The V617F mutation in the pseudokinase domain of JAK2 renders the unmutated kinase domain constitutively active. We have performed random mutagenesis at position 617 of JAK2 and tested each of the 20 possible amino acids for ability to induce constitutive signaling in Ba/F3 cells expressing the erythropoietin receptor. Four JAK2 mutants, V617W, V617M, V617I, and V617L, were able to induce cytokine independence and constitutive downstream signaling. Only V617W induced a level of constitutive activation comparable with V617F. Also, only V617W stabilized tyrosine-phosphorylated suppressor of cytokine signaling 3 (SOCS3), a mechanism by which JAK2 V617F overcomes inhibition by SOCS3. The V617W mutant induced a myeloproliferative disease in mice, mainly characterized by erythrocytosis and megakaryocytic proliferation. Although JAK2 V617W would predictably be pathogenic in humans, the substitution of the Val codon, GTC, by TTG, the codon for Trp, would require three base pair changes, and thus it is unlikely to occur. We discuss how the predicted conformations of the activated JAK2 mutants can lead to better screening assays for novel small molecule inhibitors.
Dusa
Alexandra
A
Ludwig Institute for Cancer Research, B-1200 Brussels, Belgium.
Staerk
Judith
J
Elliott
Joanne
J
Pecquet
Christian
C
Poirel
Hélène A
HA
Johnston
James A
JA
Constantinescu
Stefan N
SN
D000818:Animals; D016026:Bone Marrow Transplantation; D002460:Cell Line; D049109:Cell Proliferation; D004789:Enzyme Activation; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D053614:Janus Kinase 2 / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D009154:Mutation / Q000235:genetics; D019000:Phosphotyrosine / Q000378:metabolism; D050796:STAT3 Transcription Factor / Q000378:metabolism; D015398:Signal Transduction; D000071223:Suppressor of Cytokine Signaling 3 Protein; D050826:Suppressor of Cytokine Signaling Proteins / Q000235:genetics / Q000378:metabolism; D014633:Valine / Q000235:genetics / Q000378:metabolism
C106644:SOCS3 protein, human; D050796:STAT3 Transcription Factor; D000071223:Suppressor of Cytokine Signaling 3 Protein; D050826:Suppressor of Cytokine Signaling Proteins; D019000:Phosphotyrosine; D053614:Janus Kinase 2; D014633:Valine