The human c-Fes tyrosine kinase binds tubulin and microtubules through separate domains and promotes microtubule assembly.
The c-Fes protein-tyrosine kinase (Fes) has been implicated in the differentiation of vascular endothelial, myeloid hematopoietic, and neuronal cells, promoting substantial morphological changes in these cell types. The mechanism by which Fes promotes morphological aspects of cellular differentiation is unknown. Using COS-7 cells as a model system, we observed that Fes strongly colocalizes with microtubules in vivo when activated via coiled-coil mutation or by coexpression with an active Src family kinase. In contrast, wild-type Fes showed a diffuse cytoplasmic localization in this system, which correlated with undetectable kinase activity. Coimmunoprecipitation and immunofluorescence microscopy showed that the N-terminal Fes/CIP4 homology (FCH) domain is involved in Fes interaction with soluble unpolymerized tubulin. However, the FCH domain was not required for colocalization with polymerized microtubules in vivo. In contrast, a functional SH2 domain was essential for microtubule localization of Fes, consistent with the strong tyrosine phosphorylation of purified tubulin by Fes in vitro. Using a microtubule nucleation assay, we observed that purified c-Fes also catalyzed extensive tubulin polymerization in vitro. Taken together, these results identify c-Fes as a regulator of the tubulin cytoskeleton that may contribute to Fes-induced morphological changes in myeloid hematopoietic and neuronal cells.
Department of Molecular Genetics and Biochemistry, University of Pittsburgh School of Medicine, E1240 Biomedical Science Tower, Pittsburgh, PA 15261, USA.
Charles E
CE
Laurent
Frank J
FJ
Delfino
Haiyun Y
HY
Cheng
Thomas E
TE
Smithgall
D000818:Animals; D001704:Biopolymers / Q000737:chemistry / Q000378:metabolism; D019556:COS Cells; D002384:Catalysis; D002455:Cell Division; D002460:Cell Line; D002522:Chlorocebus aethiops; D003703:Demecolcine / Q000494:pharmacology; D004789:Enzyme Activation; D006801:Humans; D007930:Leucine / Q000235:genetics / Q000378:metabolism; D008239:Lysine / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008870:Microtubules / Q000737:chemistry / Q000378:metabolism*; D009154:Mutation / Q000235:genetics; D010766:Phosphorylation; D011485:Protein Binding / Q000187:drug effects; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011505:Protein-Tyrosine Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051578:Proto-Oncogene Proteins c-fes; D008941:Spindle Apparatus / Q000378:metabolism; D014404:Tubulin / Q000737:chemistry / Q000378:metabolism*; D018909:src Homology Domains / Q000235:genetics
D001704:Biopolymers; D011518:Proto-Oncogene Proteins; D014404:Tubulin; D011505:Protein-Tyrosine Kinases; C496645:FES protein, human; C496646:Fes protein, mouse; D051578:Proto-Oncogene Proteins c-fes; D007930:Leucine; D008239:Lysine; D003703:Demecolcine