Hsp90 and a tyrosine embedded in the Hsp90 recognition loop are required for the Fer tyrosine kinase activity.
Hsp90 is a key regulator of tyrosine kinases activity and is therefore considered as a promising target for intervention with deregulated signaling pathways in malignant cells. Here we describe a novel Hsp90 client - the intracellular tyrosine kinase, Fer, which is subjected to a unique regulatory regime by this chaperone. Inhibition of Hsp90 activity led to proteasomal degradation of the Fer enzyme. However, circumventing the dependence of Fer accumulation on Hsp90, revealed the dependence of the Fer kinase activity and its ability to phosphorylate Stat3 on the chaperone, expressing the necessity of Hsp90 for its function. Mutation analysis unveiled a tyrosine (Tyr(616)) embedded in the Hsp90 recognition loop, which is required for the kinase activity of Fer. Replacement of this tyrosine by phenylalanine (Y616F) disabled the auto-phosphorylation activity of Fer and abolished its ability to phosphorylate Stat3. Notably, surrounding the replaced Y616F with subtle mutations restored the auto and trans-phosphorylation activities of Fer suggesting that Y(616) is not itself an essential auto-phosphorylation site of the kinase. Taken together, our results portray Hsp90 and its recognition loop as novel positive regulators of the Fer tyrosine kinase stability and activity.
Hikri
Elad
E
The Mina & Everard Goodman Faculty of Life Sciences, Bar Ilan University, Ramat Gan 52900, Israel.
Shpungin
Sally
S
Nir
Uri
U
D000230:Adenocarcinoma / Q000473:pathology; D020816:Amino Acid Motifs; D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites; D001943:Breast Neoplasms / Q000473:pathology; D002384:Catalysis; D002460:Cell Line / Q000378:metabolism; D045744:Cell Line, Tumor / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism; D005260:Female; D018841:HSP90 Heat-Shock Proteins / Q000235:genetics / Q000502:physiology*; D006801:Humans; D008297:Male; D051379:Mice; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D017354:Point Mutation; D011471:Prostatic Neoplasms / Q000473:pathology; D011499:Protein Processing, Post-Translational; D011505:Protein-Tyrosine Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013329:Structure-Activity Relationship; D014443:Tyrosine / Q000378:metabolism
D018841:HSP90 Heat-Shock Proteins; D011993:Recombinant Fusion Proteins; C052602:proto-oncogene protein c-fes-fps; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases