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The immunosuppressive drugs FK506 and cyclosporin A block T-lymphocyte proliferation by inhibiting calcineurin, a critical signaling molecule for activation. Multiple intracellular receptors (immunophilins) for these drugs that specifically bind either FK506 and rapamycin (FK506-binding proteins [FKBPs]) or cyclosporin A (cyclophilins) have been identified. We report the cloning and characterization of a new 51-kDa member of the FKBP family from murine T cells. The novel immunophilin, FKBP51, is distinct from the previously isolated and sequenced 52-kDa murine FKBP, demonstrating 53% identity overall. Importantly, Western blot (immunoblot) analysis showed that unlike all other FKBPs characterized to date, FKBP51 expression was largely restricted to T cells. Drug binding to recombinant FKBP51 was demonstrated by inhibition of peptidyl prolyl isomerase activity. As judged from peptidyl prolyl isomerase activity, FKBP51 had a slightly higher affinity for rapamycin than for FK520, an FK506 analog. FKBP51, when complexed with FK520, was capable of inhibiting calcineurin phosphatase activity in an in vitro assay system. Inhibition of calcineurin phosphatase activity has been implicated both in the mechanism of immunosuppression and in the observed toxic side effects of FK506 in nonlymphoid cells. Identification of a new FKBP that can mediate calcineurin inhibition and is restricted in its expression to T cells suggests that new immunosuppressive drugs may be identified that, by virtue of their specific interaction with FKBP51, would be targeted in their site of action.Cyclosporin A (CsA), FK506, and rapamycin are potent immunosuppressive drugs that inhibit T-lymphocyte proliferation. The action of these drugs is mediated by intracellular receptors, termed immunophilins, that bind either CsA (cyclophilins) or FK506 and rapamycin (FK506-binding proteins [FKBPs]) (for reviews, see references 43 and 47). Previous studies identified these receptors as abundant, cytosolic proteins possessing an inherent peptidyl prolyl cis-trans isomerase (PPIase; rotamase) activity that is inhibited by drug ligand binding (12,16,46).The first FKBP to be described in detail at the protein (16, 46) and cDNA (25, 53) levels was FKBP12, a ubiquitous immunophilin of 11.8 kDa highly conserved in eukaryotes. More recently, additional members of this family have been identified on the basis of their ability to bind FK506 and rapamycin. Sequence analysis of the cloned genes corresponding to FKBP12.6, FKBP13, FKBP25, and FKBP52 (named to reflect their molecular weights) from mammalian sources reveals extensive conservation of amino acid sequence, in particular in the protein domain responsible for drug ligand binding and PPIase activity (for reviews, see references 15 and 58).Although the actions of FK506 and rapamycin are mediated by the same family of intracellular receptors, these drugs achieve immunosuppression by different mechanisms (5). Rapamycin inhibits a calcium-independent event involved in the proliferative response of T cells to growth ...

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Glucocorticoids and cyclic AMP exert dramatic effects on the proliferation and viability of murine T lymphocytes through unknown mechanisms. To identify gene products which might be involved in glucocorticoid-induced responses in lymphoid cells, we constructed a XcDNA library prepared from murine thymoma WEHI-7TG cells treated for 5 h with glucocorticoids and forskolin. The library was screened with a subtracted cDNA probe enriched for sequences induced by the two drugs, and cDNA clones representing 11 different inducible genes were isolated. The pattern of expression in BALB/c mouse tissues was examined for each cDNA clone. We have identified two clones that hybridized to mRNAs detected exclusively in the thymus. Other clones were identified that demonstrated tissue-specific gene expression in heart, brain, brain and thymus, or lymphoid tissue (spleen and thymus). The kinetics of induction by dexamethasone and forskolin were examined for each gene. The majority of the cDNA clones hybridized to mRNAs that were regulated by glucocorticoids and forskolin, two were regulated only by glucocorticoids, and three hybridized to mRNAs that required both drugs for induction. Inhibition of protein synthesis by cycloheximide resulted in the induction of all mRNAs that were inducible by glucocorticoids. Preliminary sequence analysis of four of the 11 cDNAs suggests that two cDNAs represent previously undescribed genes while two others correspond to the mouse VL30 retrovirus-like element and the mouse homolog of chondroitin sulfate proteoglycan core protein.Glucocorticoids induce cytolysis in susceptible lymphocyte populations, including immature thymocytes and certain leukemias and lymphomas. Steroid-induced cell death is thought to occur through the activation of an endogenous suicide process. Glucocorticoid treatment of murine lymphocytes results in severe alterations in cellular metabolism which include an inhibition of glucose transport (50), increased RNA (14) and protein (44) degradation, a rise in intracellular calcium levels (37), and decreased incorporation of thymidine into DNA (9). Glucocorticoid-induced lymphocytolysis is preceded by cell cycle arrest (28) and extreme morphological changes. These include widespread chromatin condensation, which is associated with extensive DNA fragmentation in both human and murine lymphocytes (22, 58). The DNA cleavage appears to result from the activation of a preexisting calcium-dependent endonuclease (17, 59). Glucocorticoid-induced DNA fragmentation and cell death are prevented when RNA and protein syntheses are inhibited (17). In addition, there is genetic evidence in mouse and human cells for the existence of a locus involved in cell lysis (25,26,63 (21,32,53,62). The possibility thus exists that both glucocorticoids and cAMP regulate the expression of genes involved in the lytic process.Glucocorticoids and cAMP are known to regulate the levels of several proteins in lymphocytes. For example, glucocorticoids induce glucocortin in rat thymus (18), glutamine synthetase in th...