DIFFERENTIAL REGULATION AND TRANSCRIPTIONAL CONTROL OF IMMEDIATE-EARLY GENE-EXPRESSION IN FORSKOLIN-TREATED WEHI7.2 THYMOMA CELLS

Agents that increase intracellular cAMP are frequently growth inhibito ry for lymphocytes and induce apoptosis in cortical thymocytes by regu lating gene expression. In the present study, immediate early gene exp ression was examined in WEHI7.2 thymoma cells undergoing cAMP-mediated apoptosis. Temporal differences in c-fos, junB, and inducible cAMP ea rly repressor (ICER) steady-state mRNA levels were observed after fors kolin exposure. Maximal induction of c-fos and junB occurred within 1 h, returning to basal levels by 3.5 h. In contrast, a 1.5-h time lag w as observed before ICER transcript levels increased, reaching maximal levels after 3.5 h. This rise in expression, correlating with the decr ease in c-fos and junB levels, preceded apoptotic DNA fragmentation by 1.5 h. Transient expression of ICER promoter constructs demonstrated that cAMP responsiveness occurred through cAMP-autoregulatory response element (CARE)3/4, two of the four proposed response elements in the ICER promoter. In contrast to the cAMP-responsive cell line JEG-3, CAR E1/2 was not functional for cAMP-activated transcription in WEHI7.2 ce lls. An observed differential binding pattern of WEHI and JEG nuclear extracts to these elements may account for the cell-specific differenc es in expression patterns. To determine the role of endogenous ICER in regulating gene expression, cells were treated with two sequential do ses of forskolin after which ICER and c-fos mRNA levels were measured. The high levels of cAMP-induced ICER expression dramatically reduced a second induction of c-fos. These data suggest that ICER expression m ay function as an antioncogene to attenuate the expression of certain protooncogenes, thereby preventing transformation and oncogenesis due to continuous overexpression. Moreover, inhibition of growth-stimulato ry genes may be required for the activation of the cell death machiner y in specific cells.