Preassembly of interleukin 2 (IL-2) receptor subunits on resting Kit 225 K6 T cells and their modulation by IL-2, IL-7, and IL-15: a fluorescence resonance energy transfer study.
Assembly and mutual proximities of alpha, beta, and gamma(c) subunits of the interleukin 2 receptors (IL-2R) in plasma membranes of Kit 225 K6 T lymphoma cells were investigated by fluorescence resonance energy transfer (FRET) using fluorescein isothiocyanate- and Cy3-conjugated monoclonal antibodies (mAbs) that were directed against the IL-2R alpha, IL-2R beta, and gamma(c) subunits of IL-2R. The cell-surface distribution of subunits was analyzed at the nanometer scale (2-10 nm) by FRET on a cell-by-cell basis. The cells were probed in resting phase and after coculture with saturating concentrations of IL-2, IL-7, and IL-15. FRET data from donor- and acceptor-labeled IL-2R beta-alpha, gamma-alpha, and gamma-beta pairs demonstrated close proximity of all subunits to each other in the plasma membrane of resting T cells. These mutual proximities do not appear to represent mAb-induced microaggregation, because FRET measurements with Fab fragments of the mAbs gave similar results. The relative proximities were meaningfully modulated by binding of IL-2, IL-7, and IL-15. Based on FRET analysis the topology of the three subunits at the surface of resting cells can be best described by a "triangular model" in the absence of added interleukins. IL-2 strengthens the bridges between the subunits, making the triangle more compact. IL-7 and IL-15 act in the opposite direction by opening the triangle possibly because they associate their private specific alpha receptors with the beta and/or gamma(c) subunits of the IL-2R complex. These data suggest that IL-2R subunits are already colocalized in resting T cells and do not require cytokine-induced redistribution. This colocalization is significantly modulated by binding of relevant interleukins in a cytokine-specific manner.
Damjanovich
S
S
Metabolism Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
Bene
L
L
Matkó
J
J
Alileche
A
A
Goldman
C K
CK
Sharrow
S
S
Waldmann
T A
TA
D000328:Adult; D002462:Cell Membrane / Q000378:metabolism; D006801:Humans; D019409:Interleukin-15 / Q000494:pharmacology*; D007376:Interleukin-2 / Q000494:pharmacology*; D015851:Interleukin-7 / Q000494:pharmacology*; D011487:Protein Conformation; D015375:Receptors, Interleukin-2 / Q000737:chemistry / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013601:T-Lymphocytes / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D019409:Interleukin-15; D007376:Interleukin-2; D015851:Interleukin-7; D015375:Receptors, Interleukin-2