Carboxyl-terminal region conserved among phosphoinositide-kinase-related kinases is indispensable for mTOR function in vivo and in vitro.
BACKGROUND
The mammalian target of rapamycin (mTOR) belongs to the family of phosphoinositide (PI)-kinase-related kinases that includes the ataxia-telangiectasia gene product (ATM). mTOR plays a critical role in controlling translational effectors such as p70 S6 kinase alpha (p70 alpha) and eukaryotic initiation factor 4E binding protein 1 (4EBP1).


RESULTS
We show that the C-terminal region of mTOR, which is highly conserved among the PI-kinase-related kinases, plays a critical role in the mTOR protein kinase activity. Deletion of the C-terminal residues did not adversely affect the expression of mTOR, but caused a nearly complete loss of the mTOR protein kinase activity toward both 4EBP1 and p70 alpha in vitro. These deletions also abolished the ability of a rapamycin-resistant mTOR mutant to rescue the activity of p70 alpha from inhibition induced by rapamycin in vivo. Furthermore, replacement of Trp2545, a conserved residue in the C-terminal region throughout the PI-kinase-related kinase family, abolished the function of the mTOR kinase, both in vivo and in vitro. However, substitution of 32 C-terminal residues of mTOR with those of ATM did not restore the mTOR function.


CONCLUSIONS
These findings define an indispensable role for the noncatalytic C-terminal region of mTOR and indicate that, although this highly conserved region may be important throughout the PI-kinase-related kinase family, it is not functionally interchangeable within the family.
Takahashi
T
T
Biosignal Research Center, Kobe University, Kobe 657-8501, Japan.
Hara
K
K
Inoue
H
H
Kawa
Y
Y
Tokunaga
C
C
Hidayat
S
S
Yoshino
K
K
Kuroda
Y
Y
Yonezawa
K
K
D000595:Amino Acid Sequence; D002460:Cell Line; D017124:Conserved Sequence; D039561:Eukaryotic Initiation Factor-4E; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010448:Peptide Initiation Factors / Q000378:metabolism; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000737:chemistry / Q000378:metabolism*; D011494:Protein Kinases; D019893:Ribosomal Protein S6 Kinases / Q000378:metabolism*; D016415:Sequence Alignment; D017384:Sequence Deletion; D020123:Sirolimus / Q000378:metabolism / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases
D039561:Eukaryotic Initiation Factor-4E; D010448:Peptide Initiation Factors; D011494:Protein Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); C546842:MTOR protein, human; D019893:Ribosomal Protein S6 Kinases; D058570:TOR Serine-Threonine Kinases; D020123:Sirolimus