CRITICAL PROLINE RESIDUES OF THE CYTOPLASMIC DOMAIN OF THE IL-5 RECEPTOR-ALPHA CHAIN AND ITS FUNCTION IN IL-5-MEDIATED ACTIVATION OF JAK KINASE AND STAT5

The high-affinity receptor (R) for IL-5 consists of a unique alpha cha in (IL-5R alpha) and a beta chain (beta c) that is shared with the rec eptors for IL-3 and granulocyte macrophage colony stimulating factor ( GM-CSF). We defined two regions of IL-5R alpha for the IL-5-induced pr oliferative response, the expression of nuclear proto-oncogenes, and t he tyrosine phosphorylation of cellular proteins including beta c, SH2 /SH3-containing proteins and JAK2 kinase. In the studies described her e, we demonstrate that IL-S, IL-3 or GM-CSF stimulation induces the ty rosine phosphorylation of JAK2, and to a lesser extent JAK1, and of ST AT5. Mutational analysis revealed that one of the proline residues, pa rticularly Pro352 and Pro355, in the membrane-proximal proline-rich se quence (Pro352-Pro353-X-Pro355) of the cytoplasmic domain of IL-5R alp ha is required for cell proliferation, and for both JAK1 and JAK2 acti vation. In addition, transfectants expressing chimeric receptors which consist of the extracellular domain of IL-5R alpha and the cytoplasmi c domain of beta c responded to IL-5 for proliferation and tyrosine ph osphorylation of JAK1. Intriguingly, electrophoretic mobility shift as say analysis revealed that STAT5 was activated in cells showing either JAK1 or JAK2 tyrosine phosphorylation. These results indicate that ac tivation of JAK1, JAK2 and STAT5 is critical to coupling IL-5-induced tyrosine phosphorylation and ultimately mitogenesis, and that Pro352 a nd Pro355 in the proline-rich sequence appear to play more essential r oles in cell growth and in both JAK1/STAT5 and JAK2/STAT5 activation t han Pro353 does.