CYCLOSPORINE-A INHIBITS EARLY MESSENGER-RNA EXPRESSION OF G(0) G(1) SWITCH GENE-2 (G0S2) IN CULTURED HUMAN BLOOD MONONUCLEAR-CELLS/

Cyclosporin A (CsA) may achieve its immunosuppressive effects by inhib iting the calcium-and calmodulin-dependent phosphatase calcineurin whi ch is required for activation of target genes by members of the NFAT ( nuclear factor of activated T cells) transcription factor family, Amon g these target genes is the gene encoding interleukin-2 (IL2), a cytok ine facilitating progression through the G(1) phase of the cell cycle, However, IL2 does not reverse CsA inhibition, suggesting that at leas t one other NFAT-sensitive gene may be involved, The human G(0)/G(1) s witch gene, G0S2, has potential NFAT-binding sites in the 5' flank and encodes a small basic potential phosphoprotein of unknown function, U sing a sensitive, reverse transcription-polymerase chain reaction (RT- PCR) assay, G0S2 mRNA levels were assayed in cultured blood mononuclea r cells, Freshly isolated cells contain high levels of G0S2 mRNA which rapidly decline, This ''spontaneous stimulation'' is also noted with some other G0S genes and has been attributed to some aspect of the iso lation procedure, In cells that have been preincubated to lower mRNA l evels, there is a transient increase in G0S2 mRNA, peaking between 1-2 h, in response to Concanavalin-A (ConA), or to the combination of pho rbol ester (TPA), and the calcium ionophore, ionomycin, Both these res ponses' are inhibited by CsA, Our results suggest that G0S2 expression is required to commit cells to enter the G(1) phase of the cell cycle , and that, while not excluding other possible targets, early inhibiti on of G0S2 expression by CsA may be important in achieving immunosuppr ession, G0S2 may be of value as a reporter gene for analyzing the mech anism of action of CsA and its influence on the positive and negative selection of lymphocytes in response to self and not-self antigens.