Roles of Jak Kinases in Human GM-CSF Receptor
The IL-3 and GM-CSF (hGMR) receptors consist of two subunits, α and β, both of which are members of the cytokine receptor superfamily. Chemical cross-linking and immunoprecipitation revealed that the α and β subunits associate following stimulation with GM-CSF, but the β subunit forms a homodimer even in the absence of the ligand. We analyzed the mechanism of c-fos mRNA activation by GM-CSF using several hGMRβ subunit mutants. In addition to boxl region, a membrane distal region (a.a. 544-589) of hGMRβ is required for c-fos activation. Only one tyrosine residue (Tyr577) exists within the region 544–589, and substitution of Tyr577 to phenylalanine in GMRβ 589 resulted in the loss of c-fos activation. In contrast, the same substitution in a wild type receptor did not affect GM-CSF-induced activities such as c-fos mRNA induction and proliferation but abolished Shc phosphorylation. These results suggest that the activation of She is not essential for c-fos activation and several tyrosine residues coordinate to activate c-fos activation.
It is well documented that IL-3 or GM-CSF activates JAK in various cells. However the role of JAK2 in IL-3/GM-CSF functions is largely unknown. We examined the role of JAK2 in GM-CSF-induced signaling pathways. Dominant negative JAK2 (∆JAK2) lacking the C-terminus kinase domain, suppressed IL3/GM-CSF induced c-fos activation, c-myc activation and proliferation, suggesting that JAK2 is involved in both signaling pathways. Several tyrosine residues are known to be phosphorylated by GM-CSF within this region and JAK2 expressed transiently in COS7 cells phosphorylated certain tyrosine residues within hGMRß. JAK2 also phosphorylated PTP1D in COS7 cells. PTP1D and Shc are phosphorylated by IL-3/GM-CSF in BA/F3 cells, but these phosphorylation events were inhibited by expression of ∆JAK2. Taken together, these results indicate that JAK2 is a primary kinase regulating all the known activities of GM-CSF. JAK2 mediates GM-CSF induced c-fos activation through receptor phosphorylation and Shc/PTP1D activation.
KeywordsTyrosine Residue COS7 Cell Wild Type Receptor Membrane Proximal Region Tyrosine Kinase Inhibitor Genistein
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