Distinct regulatory mechanism for p70 S6 kinase β from that for p70 S6 kinase α

Abstract

Background: A novel ribosomal S6 kinase, termed p70 S6 kinase β (p70β), has a highly homologous amino acid sequence to that of p70/p85 S6 kinase (p70α). This includes the critical phosphorylation sites, Thr252, Ser394 and Thr412 in p70α1, which correspond to Thr241, Ser383 and Thr401 in p70β1, respectively. However, the regulatory mechanism for p70β remains to be elucidated. Results: We report here the expression and the mechanism of in vivo regulation of p70β. Two isoforms, p70β1 and p70β2, were expressed in a variety of tissues at a different level. p70β1 was mainly targeted to the nucleus, whereas p70β2 dispersed throughout the cytoplasm including nucleoplasm. The kinase activity of p70β1 was less sensitive to the inhibition induced by rapamycin, wortmannin and amino acid withdrawal than that of p70α. The portion of p70β activity inhibited by rapamycin was rescued by the rapamycin-resistant mutant of the mammalian target of rapamycin (mTOR). Mutational analysis revealed that the phosphorylation of Thr241 and Thr401 in p70β1 was indispensable for the kinase activity. In contrast, a p70β1 mutant in which Ser383 was substituted with Gly (S383G) still retained nearly the half maximal activity. Sequential phosphorylation of wild-type and S383G mutant of p70β1 with mTOR and 3-phosphoinositide-dependent protein kinase 1 (PDK1) in vitro synergistically activated their kinase activities. Conclusion: These results indicate that p70β is regulated by the mTOR- and PDK1-signalling pathways through a synergistic interaction between phosphorylated Thr241 and Thr401, while Ser383 plays minor role in their activation mechanism. Activated p70β may be less sensitive to dephosphorylation mediated by putative phosphatases activated by rapamycin, amino acid withdrawal, and probably wortmannin.