Expression of mTOR pathway proteins in human amniotic fluid stem cells.
The discovery of human amniotic fluid stem cells initiated a new and promising stem cell research field. These cells harbor a high proliferative capacity and the potential to differentiate into cells of all three embryonic germ layers. The facts that they do not form tumors in vivo and do not raise the ethical concerns associated with human embryonic stem cells support their role as an optimal tool to study the underlying molecular mechanisms of cell differentiation processes and of their deregulation in human genetic diseases. Deregulation of the protein kinase mammalian target of rapamycin (mTOR) pathway is a hallmark of a wide variety of human genetic diseases. Here we report the establishment of an amniotic fluid stem cell line. We analysed the endogenous expression of the mTOR pathway proteins tuberin, mTOR, raptor, rictor, sin1, mLST8, Akt and p70S6K in human amniotic fluid stem cells. In addition, we studied the endogenous activity of the kinase p70S6K, one of the major targets of the mTOR complex 1 kinase, by analysing the p70S6K T389 phosphorylation status. The activity of the Akt kinase, the major mTOR complex 2 target, was studied by analysing its phosphorylation at S473. In addition, the mTOR inhibitor rapamycin was found to affect the phosphorylation status of p70S6K in amniotic fluid stem cells. Taken together, we provide evidence that the mTOR pathway is fully active in human amniotic fluid stem cells. These data demonstrate that amniotic fluid stem cell lines can be used as new tools to study the molecular and cell biological consequences of natural occurring alterations of the mTOR pathway being responsible for a wide variety of different human genetic diseases.
Siegel
Nicol
N
Medical Genetics, Medical University of Vienna, Währinger Gürtel 18-20, A-1090 Vienna, Austria.
Valli
Alessandro
A
Fuchs
Christiane
C
Rosner
Margit
M
Hengstschläger
Markus
M
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000653:Amniotic Fluid / Q000166:cytology*; D002352:Carrier Proteins / Q000378:metabolism; D002478:Cells, Cultured; D006801:Humans; D015151:Immunoblotting; D007166:Immunosuppressive Agents / Q000494:pharmacology; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism; D000076226:Rapamycin-Insensitive Companion of mTOR Protein; D000076223:Regulatory-Associated Protein of mTOR; D038762:Ribosomal Protein S6 Kinases, 70-kDa / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology*; D020123:Sirolimus / Q000494:pharmacology; D013234:Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D058570:TOR Serine-Threonine Kinases; D000077005:Tuberous Sclerosis Complex 2 Protein; D025521:Tumor Suppressor Proteins / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D007166:Immunosuppressive Agents; C493277:MAPKAP1 protein, human; D011506:Proteins; C492571:RICTOR protein, human; C463659:RPTOR protein, human; D000076226:Rapamycin-Insensitive Companion of mTOR Protein; D000076223:Regulatory-Associated Protein of mTOR; C000624653:TSC2 protein, human; D000077005:Tuberous Sclerosis Complex 2 Protein; D025521:Tumor Suppressor Proteins; D011494:Protein Kinases; C546842:MTOR protein, human; D038762:Ribosomal Protein S6 Kinases, 70-kDa; D058570:TOR Serine-Threonine Kinases; D020123:Sirolimus