Mammalian target of rapamycin and S6 kinase 1 positively regulate 6-thioguanine-induced autophagy.
DNA mismatch repair (MMR) ensures the fidelity of DNA replication and is required for activation of cell cycle arrest and apoptosis in response to certain classes of DNA damage. We recently reported that MMR is also implicated in initiation of an autophagic response after MMR processing of 6-thioguanine (6-TG). It is now generally believed that autophagy is negatively controlled by mammalian target of rapamycin (mTOR) activity. To determine whether mTOR is involved in 6-TG-induced autophagy, we used rapamycin, a potential anticancer agent, to inhibit mTOR activity. Surprisingly, we find that rapamycin cotreatment inhibits 6-TG-induced autophagy in MMR-proficient human colorectal cancer HCT116 (MLH1(+)) and HT29 cells as measured by LC3 immunoblotting, GFP-LC3 relocalization, and acridine orange staining. Consistently, short interfering RNA silencing of the 70-kDa ribosomal S6 kinase 1 (S6K1), the downstream effector of mTOR, markedly reduces 6-TG-induced autophagy. Furthermore, we show that inhibition of mTOR by rapamycin induces the activation of Akt as shown by increased Akt phosphorylation at Ser(473) and the inhibition of 6-TG-induced apoptosis and cell death. Activated Akt is a well-known inhibitor of autophagy. In conclusion, our data indicate that mTOR-S6K1 positively regulates autophagy after MMR processing of 6-TG probably through its negative feedback inhibition of Akt.
Zeng
Xuehuo
X
Department of Radiation Oncology, Case Western Reserve University, Case Comprehensive Cancer Center/University Hospitals Case Medical Center, Cleveland, OH 44106-6068, USA.
Kinsella
Timothy J
TJ
D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D001343:Autophagy / Q000187:drug effects* / Q000502:physiology; D053843:DNA Mismatch Repair; D045325:HCT116 Cells; D019073:HT29 Cells; D006801:Humans; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D019893:Ribosomal Protein S6 Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D020123:Sirolimus / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases; D013866:Thioguanine / Q000494:pharmacology*
D000964:Antimetabolites, Antineoplastic; D019869:Phosphatidylinositol 3-Kinases; D011494:Protein Kinases; D051057:Proto-Oncogene Proteins c-akt; D034741:RNA, Small Interfering; D019893:Ribosomal Protein S6 Kinases; D020123:Sirolimus; D058570:TOR Serine-Threonine Kinases; D013866:Thioguanine; C546842:MTOR protein, human