We report that the erythropoietin receptor cytosolic juxtamembrane region i
s conformationally rigid and contains a hydrophobic motif, composed of resi
dues L-253, I-257, and W-258, that is crucial for Janus kinase 2 (JAK2) act
ivation and receptor signaling. Alanine insertion mutagenesis shows that th
e orientation of this motif and not its distance from the membrane bilayer
is critical. Intragenic complementation studies suggest that L-253 is conta
ined within an alpha helix functionally continuous to the transmembrane alp
ha helix. The alpha -helical orientation of L-253 is required not for JAK2
activation but for activated JAK2 to induce phosphorylation of the erythrop
oietin receptor. This motif is highly conserved among cytokine receptors an
d couples ligand-induced conformational changes in the receptor to intracel
lular activation of JAK2.