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 (hIL)-3 receptor (hbetac) participate in disulfide-linked receptor subunit heterodimerization. This linkage is essential for receptor tyrosine phosphorylation, since the Cys-86 --> Ala (Mc4) and Cys-91 --> Ala (Mc5) mutations abolished both events. Here, we used these mutants to examine whether disulfide-linked receptor dimerization affects the biological and biochemical activities of the IL-3 receptor. Murine T cells expressing hIL-3Ralpha and Mc4 or Mc5 did not proliferate in hIL-3, whereas cells expressing wild-type hbetac exhibited rapid proliferation. However, a small subpopulation of cells expressing each mutant could be selected for growth in IL-3, and these proliferated similarly to cells expressing wild-type hbetac, despite failing to undergo IL-3-stimulated hbetac tyrosine phosphorylation. The Mc4 and Mc5 mutations substantially reduced, but did not abrogate, IL-3-mediated anti-apoptotic activity in the unselected populations. Moreover, the mutations abolished IL-3-induced JAK2, STAT, and AKT activation in the unselected 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 hbetac tyrosine phosphorylation are normally important for receptor activation, alternative mechanisms can bypass these requirements.
Le
F
F
Hanson Centre for Cancer Research, The Institute of Medical and Veterinary Science, Adelaide, South Australia 5000, Australia.
Stomski
F
F
Woodcock
J M
JM
Lopez
A F
AF
Gonda
T J
TJ
D000818:Animals; D002460:Cell Line; D003545:Cysteine / Q000235:genetics / Q000378:metabolism*; D019281:Dimerization; D004220:Disulfides / Q000378:metabolism*; D006801:Humans; D053614:Janus Kinase 2; D020935:MAP Kinase Signaling System; D051379:Mice; D016296:Mutagenesis; D017346:Protein Serine-Threonine Kinases; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D016185:Receptors, Interleukin-3 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D014157:Transcription Factors / Q000378:metabolism
D004220:Disulfides; D011518:Proto-Oncogene Proteins; D016185:Receptors, Interleukin-3; D014157:Transcription Factors; D011505:Protein-Tyrosine Kinases; C507924:JAK2 protein, human; C507926:Jak2 protein, mouse; D053614:Janus Kinase 2; C494918:AKT1 protein, human; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D003545:Cysteine