Evidence for the involvement of three distinct signals in the induction of IL-2 gene expression in human T lymphocytes.
The regulation of IL-2 gene expression during T cell activation and proliferation has been investigated in primary cultures of purified human peripheral blood T cells. Prior results indicated that stimulation of T cells by anti-CD28 mAb plus PMA could induce IL-2 expression and T cell proliferation that was entirely resistant to cyclosporine. The present studies examined whether CD28 augments IL-2 expression by a unique pathway or merely acts at a point common to CD3-induced proliferation but distal to the effects of cyclosporine. The induction of maximal IL-2 gene expression required three signals provided by phorbol ester, calcium ionophore, and anti-CD28 mAb. Stimulation of cells by optimal amounts of calcium ionophore and PMA induced IL-2 mRNA that was completely suppressed by cyclosporine. The addition of anti-CD28 to T cells stimulated with PMA plus calcium ionophore induced a 5- to 100-fold increase in IL-2 gene expression and secretion that was resistant to cyclosporine. The CD28 signal was able to increase steady state IL-2 mRNA levels even in cells treated with maximally tolerated amounts of calcium ionophore and PMA. The three-signal requirement did not reflect differential regulation of lymphokine gene expression between the CD4 and CD8 T cell subsets or differences in the kinetics of IL-2 mRNA expression. The signal provided by CD28 is distinct from that of CD3 because although anti-CD28 plus PMA-induced proliferation is resistant to cyclosporine, anti-CD3 or anti-CD3 plus PMA-induced IL-2 expression is sensitive. Thus, these studies show that three biochemically distinct signals are required for maximal IL-2 gene expression. Furthermore, these studies suggest that lymphokine production in T cells is not controlled by an "on/off" switch, but rather, that CD28 regulates a distinct intracellular pathway which modulates the level of IL-2 production on a per cell basis. The observation that CD28 stimulation results in IL-2 concentrations that exceed 1000 U/m1 in tissue culture supernatants suggests that a role in vivo for CD28 might be to amplify immune responses initiated by the CD3/T cell receptor complex. Finally, the observation that CD28 interacts with the signals provided by PMA and calcium ionophore shows that the function of CD28 is not merely to act as a scaffold to stabilize or enhance signalling through the CD3/TCR complex.
June
C H
CH
Naval Medical Research Institute, Bethesda, MD 20814.
Ledbetter
J A
JA
Lindsten
T
T
Thompson
C B
CB
D000911:Antibodies, Monoclonal / Q000502:physiology; D000945:Antigens, Differentiation, T-Lymphocyte / Q000276:immunology; D018106:CD28 Antigens; D003524:Cyclosporins / Q000494:pharmacology; D004987:Ethers / Q000494:pharmacology; D005786:Gene Expression Regulation; D006801:Humans; D007376:Interleukin-2 / Q000235:genetics* / Q000378:metabolism; D015759:Ionomycin; D007476:Ionophores / Q000494:pharmacology*; D008213:Lymphocyte Activation / Q000187:drug effects; D010641:Phenotype; D012333:RNA, Messenger / Q000096:biosynthesis; D015398:Signal Transduction / Q000187:drug effects; D013601:T-Lymphocytes / Q000145:classification / Q000276:immunology / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D000911:Antibodies, Monoclonal; D000945:Antigens, Differentiation, T-Lymphocyte; D018106:CD28 Antigens; D003524:Cyclosporins; D004987:Ethers; D007376:Interleukin-2; D007476:Ionophores; D012333:RNA, Messenger; D015759:Ionomycin; D013755:Tetradecanoylphorbol Acetate