Distinct regulatory mechanism for p70 S6 kinase beta from that for p70 S6 kinase alpha.
BACKGROUND
A novel ribosomal S6 kinase, termed p70 S6 kinase beta (p70beta), has a highly homologous amino acid sequence to that of p70/p85 S6 kinase (p70alpha). This includes the critical phosphorylation sites, Thr252, Ser394 and Thr412 in p70alpha1, which correspond to Thr241, Ser383 and Thr401 in p70beta1, respectively. However, the regulatory mechanism for p70beta remains to be elucidated.


RESULTS
We report here the expression and the mechanism of in vivo regulation of p70beta. Two isoforms, p70beta1 and p70beta2, were expressed in a variety of tissues at a different level. p70beta1 was mainly targeted to the nucleus, whereas p70beta2 dispersed throughout the cytoplasm including nucleoplasm. The kinase activity of p70beta1 was less sensitive to the inhibition induced by rapamycin, wortmannin and amino acid withdrawal than that of p70alpha. The portion of p70beta 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 p70beta1 was indispensable for the kinase activity. In contrast, a p70beta1 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 p70beta1 with mTOR and 3-phosphoinositide-dependent protein kinase 1 (PDK1) in vitro synergistically activated their kinase activities.


CONCLUSIONS
These results indicate that p70beta 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 p70beta may be less sensitive to dephosphorylation mediated by putative phosphatases activated by rapamycin, amino acid withdrawal, and probably wortmannin.
Minami
T
T
Biosignal Research Center, Kobe University, Kobe 657-8501, Japan.
Hara
K
K
Oshiro
N
N
Ueoku
S
S
Yoshino
K
K
Tokunaga
C
C
Shirai
Y
Y
Saito
N
N
Gout
I
I
Yonezawa
K
K
D000730:Androstadienes / Q000494:pharmacology; D015153:Blotting, Western; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D019893:Ribosomal Protein S6 Kinases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020123:Sirolimus / Q000494:pharmacology; D000077191:Wortmannin
D000730:Androstadienes; D004791:Enzyme Inhibitors; D007527:Isoenzymes; D019893:Ribosomal Protein S6 Kinases; D020123:Sirolimus; D000077191:Wortmannin