Rapamycin inhibition of interleukin-2-dependent p33cdk2 and p34cdc2 kinase activation in T lymphocytes.
The immunosuppressant rapamycin (RAP) is a potent inhibitor of the entry of interleukin (IL)-2-stimulated T cells into S-phase. Earlier results indicated that RAP treatment arrested the growth of the murine IL-2-dependent T cell line CTLL-2 in late G1-phase. To explore further the interactions of RAP with the cell cycle control machinery in T cells, we examined the effects of RAP treatment on the activation of the cyclin-dependent kinases p34cdc2 and p33cdk2 in G1-phase CTLL-2 cells. Stimulation of factor-deprived cells with IL-2 led to the assembly of high molecular weight complexes containing active p34cdc2 and p33cdk2. The appearance of these complexes was explained, at least in part, by the association of both cyclin-dependent kinases with IL-2-induced cyclin A. RAP treatment profoundly inhibited both cyclin A expression and the appearance of active cyclin A-cyclin-dependent kinase complexes in IL-2-stimulated, late G1-phase CTLL-2 cells. Although p34cdc2 activation was largely dependent on association with cyclin A, a significant proportion of the active p33cdk2 pool was complexed with cyclin E. In contrast to cyclin A, the IL-2-induced accumulation of cyclin E in G1-phase cells was only partially suppressed by RAP, and cyclin E-p33cdk2 complexes were readily detected in drug-treated cells. These cyclin E-cyclin-dependent kinase complexes were nonetheless devoid of histone H1 kinase activity. The inhibitory effects of RAP on the activation of cyclin E- and cyclin A-associated cyclin-dependent kinases suggest that one or both events participate in the regulation of T cell entry into S-phase.
Morice
W G
WG
Department of Immunology, Mayo Clinic, Rochester, Minnesota 55905.
Wiederrecht
G
G
Brunn
G J
GJ
Siekierka
J J
JJ
Abraham
R T
RT
D000595:Amino Acid Sequence; D015152:Blotting, Northern; D016203:CDC2 Protein Kinase / Q000378:metabolism*; D042846:CDC2-CDC28 Kinases; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002850:Chromatography, Gel; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases; D016213:Cyclins / Q000096:biosynthesis / Q000302:isolation & purification; D015342:DNA Probes; D004261:DNA Replication / Q000187:drug effects; D004789:Enzyme Activation; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology*; D007376:Interleukin-2 / Q000494:pharmacology*; D007700:Kinetics; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000276:immunology; D011090:Polyenes / Q000494:pharmacology*; D006656:Protamine Kinase / Q000378:metabolism; D011494:Protein Kinases / Q000302:isolation & purification / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D011994:Recombinant Proteins / Q000494:pharmacology; D020123:Sirolimus; D013601:T-Lymphocytes / Q000187:drug effects* / Q000201:enzymology
D016213:Cyclins; D015342:DNA Probes; D007166:Immunosuppressive Agents; D007376:Interleukin-2; D009842:Oligopeptides; D011090:Polyenes; D011994:Recombinant Proteins; D011494:Protein Kinases; D006656:Protamine Kinase; D017346:Protein Serine-Threonine Kinases; D016203:CDC2 Protein Kinase; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases; D020123:Sirolimus