The role of disulfide-linked dimerization in interleukin-3 receptor signaling and biological activity

Cysteine residues 86 and 91 of the beta subunit of the human interleukin (h IL)-3 receptor (h beta c) participate in disulfide-linked receptor subunit heterodimerization. This linkage is essential for receptor tyrosine phospho rylation, since the Cys-86 --> Ala (Mc4) and Cys-91 --> Ala (Mc5) mutations abolished both events. Here, we used these mutants to examine whether disu lfide-linked receptor dimerization affects the biological and biochemical a ctivities of the IL-3 receptor. Murine T cells expressing hIL-3R alpha and Mc4 or Mc5 did not proliferate in hIL-3, whereas cells expressing wild-type h beta c exhibited rapid proliferation. However, a small subpopulation of cells expressing each mutant could be selected for growth in IL-3, and thes e proliferated similarly to cells expressing wild-type h beta c, despite fa iling to undergo IL-3-stimulated h beta e tyrosine phosphorylation. The Mc4 and Mc5 mutations substantially reduced, but did not abrogate, IL-3-mediat ed anti-apoptotic activity in the unselected populations. Moreover, the mut ations abolished IL-3-induced JAK2, STAT, and AKT activation in the unselec ted cells, whereas activation of these molecules in IL-3-selected cells was normal. In contrast, Mc4 and Mc5 showed a limited effect on activation of Erk1 and -2 in unselected cells. These data suggest that whereas disulfide- mediated cross-linking and h beta c tyrosine phosphorylation are normally i mportant for receptor activation, alternative mechanisms can bypass these r equirements.