AKT activity determines sensitivity to mammalian target of rapamycin (mTOR) inhibitors by regulating cyclin D1 and c-myc expression.
Prior work demonstrates that AKT activity regulates sensitivity of cells to G(1) arrest induced by mammalian target of rapamycin (mTOR) inhibitors such as rapamycin and CCI-779. To investigate this, a novel high-throughput microarray polysome analysis was performed to identify genes whose mRNA translational efficiency was differentially affected following mTOR inhibition. The analysis also allowed the assessment of steady-state transcript levels. We identified two transcripts, cyclin D1 and c-myc, which exhibited differential expression in an AKT-dependent manner: High levels of activated AKT resulted in rapamycin-induced down-regulation of expression, whereas low levels resulted in up-regulation of expression. To ectopically express these proteins we exploited the finding that the p27(kip1) mRNA was efficiently translated in the face of mTOR inhibition irrespective of AKT activity. Thus, the p27(kip1) 5'-untranslated region was fused to the cyclin D1 and c-myc coding regions and these constructs were expressed in cells. In transfected cells, expression of cyclin D1 or c-myc was not decreased by rapamycin. Most importantly, this completely converted sensitive cells to a phenotype resistant to G(1) arrest. Furthermore, the AKT-dependent differential expression patterns of these two genes was also observed in a mouse xenograft model following in vivo treatment with CCI-779. These results identify two critical downstream molecular targets whose expression is regulated by AKT activity and whose down-regulation is required for rapamycin/CCI-779 sensitivity.
Department of Medicine, West Los Angeles Veteran's Administration-UCLA Medical Center, University of California School of Medicine, Los Angeles, California 90073, USA. gera@ucla.edu
Joseph F
JF
Gera
Ingo K
IK
Mellinghoff
Yijiang
Y
Shi
Matthew B
MB
Rettig
Chris
C
Tran
Jung-hsin
JH
Hsu
Charles L
CL
Sawyers
Alan K
AK
Lichtenstein
D002460:Cell Line; D019938:Cyclin D1 / Q000096:biosynthesis* / Q000235:genetics; D016193:G1 Phase / Q000187:drug effects / Q000235:genetics; D005786:Gene Expression Regulation / Q000187:drug effects; D016259:Genes, myc; D006801:Humans; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000235:genetics; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics; D051057:Proto-Oncogene Proteins c-akt; D016271:Proto-Oncogene Proteins c-myc / Q000096:biosynthesis* / Q000235:genetics; D015398:Signal Transduction / Q000187:drug effects; D020123:Sirolimus / Q000031:analogs & derivatives* / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases
D047428:Protein Kinase Inhibitors; D011518:Proto-Oncogene Proteins; D016271:Proto-Oncogene Proteins c-myc; D019938:Cyclin D1; C401859:temsirolimus; D011494:Protein Kinases; C546842:MTOR protein, human; C546843:mTOR protein, mouse; C494918:AKT1 protein, human; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D058570:TOR Serine-Threonine Kinases; D020123:Sirolimus