Identification of novel targets of immunosuppressive agents by cDNA-based microarray analysis.
The immunosuppressive agents cyclosporin A (CsA) and tacrolimus (FK506) bind to unrelated intracellular immunophilin receptors, cyclophilin (CyP) and FK506-binding protein (FKBP), respectively. The complexes of CsA-CyP and of FK506-FKBP both bind to and inhibit the activity of the calcium/calmodulin-dependent serine/threonine phosphatase calcineurin. We used cDNA microarray analysis to characterize early human peripheral blood T cell transcriptional responses following antigen receptor stimulation in the absence or presence of CsA or FK506, hoping to identify novel targets dependent upon calcineurin or immunophilins or, perhaps, specific targets of either CyP or FKBP inhibitable by one drug alone. The array data failed to identify genes uniquely sensitive to only one drug, suggesting that transcriptionally regulated, immunophilin-dependent but calcineurin-independent targets fell below the limits of detection in this system. In contrast, transcript profiling identified and mRNA and protein analysis confirmed novel as well as known genes reproducibly induced or inhibited by both immunosuppressive agents. In this context, we show that transcriptional activation of Stat5a and repression of the cytokine interleukin-16 are regulated by T cell receptor engagement and dependent upon drug-immunophilin complexes and, presumably, calcineurin activity.
Cristillo
Anthony D
AD
Laboratory of Lymphocyte Biology, NHLBI, National Institutes of Health, Bethesda, Maryland 20892, USA.
Bierer
Barbara E
BE
D001483:Base Sequence; D016572:Cyclosporine / Q000494:pharmacology*; D017931:DNA Primers; D018076:DNA, Complementary; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology*; D019410:Interleukin-16 / Q000235:genetics / Q000378:metabolism; D008894:Milk Proteins; D005810:Multigene Family; D020411:Oligonucleotide Array Sequence Analysis; D012333:RNA, Messenger / Q000235:genetics; D050799:STAT5 Transcription Factor; D013601:T-Lymphocytes / Q000187:drug effects* / Q000378:metabolism; D016559:Tacrolimus / Q000494:pharmacology*; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism; D025521:Tumor Suppressor Proteins
D017931:DNA Primers; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D007166:Immunosuppressive Agents; D019410:Interleukin-16; D008894:Milk Proteins; D012333:RNA, Messenger; D050799:STAT5 Transcription Factor; C494092:STAT5A protein, human; D015534:Trans-Activators; D025521:Tumor Suppressor Proteins; D016572:Cyclosporine; D016559:Tacrolimus