Multiple signaling pathways mediate interleukin-4-induced 3 beta-hydroxysteroid dehydrogenase/Delta(5)-Delta(4) isomerase type 1 gene expression in human breast cancer cells

The 3 beta-hydroxysteroid dehydrogenase/Delta 5-Delta 4 isomerase (3 beta-H SD) isoenzymes catalyze an essential step in the formation of all classes o f active steroid hormones. We have recently shown that 3 beta-HSD type 1 ge ne expression is specifically induced by interleukin (IL)-4 and IL-13 in br east human cancer cell lines and in normal human mammary epithelial cells i n primary culture. There is evidence that IL-4 stimulates bifurcating signa ling pathways in which the signal transducer and activator of transcription -6 (Stat6)-signal pathway is involved in differentiation and gene regulatio n, whereas insulin receptor substrate (IRS) proteins mediate the mitogenic action of IL-4. In fact, we have shown that State was activated by IL-4 in all cell lines studied where IL-4 induced 3 beta-HSD expression, but not in those that failed to respond to IL-4. The present study was designed to in vestigate the potential contribution of IRS proteins and their downstream t argets to IL-4-induced 3 beta-HSD type 1 gene expression. IL-4 rapidly indu ced IRS-1 and IRS-2 phosphorylation in ZR-75-1 human breast cancer cell lin es. Moreover, insulin-like growth factor (IGF)-I and insulin, which are wel l known to cause IRS-1 and IRS-2 phosphorylation, increased the stimulatory effect of IL-4 on 3 beta-HSD activity. IRS-1 and IRS-2 are adapter molecul es that provide docking sites for different SH2-domain-containing proteins such as the phosphatidylinositol (PI) 3-kinase. In this light, the inhibiti on of IL-4-induced 3 beta-HSD expression by wortmannin and LY294002, two po tent PI 3-kinase inhibitors, indicates the probable involvement of the PI 3 -kinase signaling molecules in this response to IL-4. Furthermore, it has b een suggested that the IRS proteins are part of the signaling complexes tha t lead to activation of the mitogen-activated protein (MAP) kinase by insul in; thus we investigated the potential role of the MAP kinase (MAPK) cascad e in the IL-4 action. In ZR-75-1 cells, both the activation of MAPK by IL-4 and the IL-4-induced 3 beta-HSD activity were completely blocked by PD9805 9, an inhibitor of MAPK activation. Wortmannin also blocked MAPK activation by IL-4, IGF-I, and insulin, suggesting that the MAPK cascade acts as a do wnstream effector of PI 3-kinases. To further understand the cross-talk bet ween signaling pathways involved in IL-4 action, we investigated the possib le involvement of protein kinase C (PKC). The potential role of PKC was sug gested by the observation that the well known PKC activator phorbol-12-myri state-13-acetate (PMA) potentiated the IL-4-induced 3 beta-HSD activity. Ta ken together, these findings suggest the existence of a novel mechanism of gene regulation by IL-4. This mechanism would involved the phosphorylation of IRS-1 and IRS-2, which transduce the IL-4 signal through a PI 3-kinase- and MAPK-dependent signaling pathway. The inability of IGF-I, insulin, and PMA to stimulate 3 beta-HSD expression by themselves in the absence of IL-4 makes obvious the absolute requirement of an IL-4-specific signaling molec ule. Our findings thus suggest that the multiple pathways downstream of IRS -1 and IRS-2 must act in cooperation with the IL-4-specific transcription f actor State to mediate the induction of 3 beta-HSD type 1 gene expression i n ZR-75-1 human breast cancer cells.