The correlations between prolactin (PRL) and breast cancer risk have been well established. One of the roles of PRL is to stimulate cell cycle progression by altering levels of cell cycle regulators. One target for PRL action is cyclin D1. PRL positively regulates the activity of the cyclin D1 promoter, and JAK2 is an important component of PRL signal transduction to cyclin D1. On the other side, increased expression of serine/threonine kinase PAK1 in breast cancer cells also stimulates cyclin D1 promoter activity. Since JAK2 directly phosphorylates and activates PAK1, we were interested in determining whether JAK2-dependent phosphorylation of PAK1 is involved in PRL-dependent regulation of cyclin D1 transcription. We have demonstrated here that overexpression of PAK1 dramatically increased cyclin D1 promoter activity in response to PRL, and that the three tyrosines that are phosphorylated by JAK2 are required for maximal cyclin D1 promoter activity in response to PRL.
We sought to understand the mechanism by which PRL-induced tyrosyl phosphorylation of PAK1 stimulates cyclin D1 transcription. Overexpression of the PAK1 mutant lacking the three nuclear localization signals decreases the magnitude of PAK1-dependent stimulation of cyclin D1 transcription as compared to wild type PAK1, suggesting that nuclear localization of PAK1 stimulates cyclin D1 promoter activity. We wanted to determine whether the stimulation of cyclin D1 promoter activity by PRL-activated PAK1 is to some extent due to the translocation of PAK1 to the nucleus in response to PRL. First, we have demonstrated that JAK2-dependent tyrosyl phosphorylated PAK1 translocates to the nucleus, which may provide a mechanism by which JAK2-dependent tyrosyl phosphorylated PAK1 enhances cyclin D1 transcription. Next, we have demonstrated that Nck is a negative regulator of nuclear translocation of PAK1 upon JAK2 activation and PAK1-dependent stimulation of cyclin D1 transcription, providing further evidence that nuclear translocation of PAK1 may be a mechanism by which PAK1 regulates cyclin D1 transcription.