Amino acid-dependent control of p70(s6k) - Involvement of tRNA aminoacylation in the regulation

In human T-lymphoblastoid cells, downstream signaling events of mammalian t arget of rapamycin (mTOR), including the activity of p70(s6k) and phosphory lation of eukaryotic initiation factor 4E-binding protein 1, were dependent on amino acid concentration in the culture media, whereas other growth-rel ated protein kinases were not. Amino acid-induced p70(s6k) activation was c ompletely inhibited by rapamycin but only partially inhibited by wortmannin . Moreover, amino acid concentration similarly affected the p70(s6k) activi ty, which was dependent on a rapamycin-resistant mutant (S2035I) of mTOR. T hese data indicate that mTOR is required for amino acid-dependent activatio n of p70(s6k). The mechanism by which amino acids regulate p70(s6k) activit y was further explored: 1) amino acid alcohols, which inhibit aminoacylatio n of tRNA by their competitive binding to tRNA synthetases, suppressed p70( s6k) activity; 2) suppression of p70(s6k) by amino acid depletion was block ed by cycloheximide or puromycin, which inhibit utilization of aminoacylate d tRNA in cells; and 3) in cells having a temperature-sensitive mutant of h istidyl tRNA synthetase, p70(s6k) was suppressed by a transition of cells t o a nonpermissible temperature, which was partially restored by addition of high concentrations of histidine. These results indicate that suppression of tRNA aminoacylation is able to inhibit p70(s6k) activity. Deacylated tRN A may be a factor negatively regulating p70(s6k).