P53 suppresses ribonucleotide reductase via inhibiting mTORC1
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Zhengfu He1, Xing Hu2, Weijin Liu2, Adrienne Dorrance3, Ramiro Garzon3, Peter J. Houghton4 and Changxian Shen3
1Department of Thoracic Surgery, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, Zhejiang Province, China
2College of Biology and Food Engineering, Huaihua University, Huaihua, Hunan Province, China
3Comprehensive Cancer Center, The Ohio State University, Columbus, Ohio, USA
4The Greehey Children’s Cancer Research Institute, The University of Texas Health Science Center at San Antonio, San Antonio, Texas, USA
Changxian Shen, email: [email protected]
Keywords: mammalian target of rapamycin (mTOR), deoxyribonucleotides (dNTPs), ribonucleotide reductase (RNR), p53, nutlin-3
Received: September 27, 2016 Accepted: April 11, 2017 Published: April 26, 2017
Balanced deoxyribonucleotides pools are essential for cell survival and genome stability. Ribonucleotide reductase is the rate-limiting enzyme for the production of deoxyribonucleotides. We report here that p53 suppresses ribonucleotide reductase subunit 1 (RRM1) and 2 (RRM2) via inhibiting mammalian target of rapamycin complex 1 (mTORC1). In vitro, cancer cell lines and mouse embryonic fibroblast cells were treated with different concentrations of pharmacological inhibitors for different times. In vivo, rhabdomyosarcoma Rh30 cell tumor-bearing mice were treated with rapamycin or AZD8055. Protein levels and phosphorylation status were assessed by immunoblotting and mRNA levels were determined by real time RT-PCR. Pharmacological inhibition of mTORC1 with rapamycin, mTOR kinase with AZD8055 or protein kinase B with MK2206 resulted in decrease of RRM1 and RRM2 in Rh30 cells both in vitro and in mouse tumor xenografts. Moreover, eukaryotic translational initiation factor 4E-binding proteins 1 and 2 double knockout mouse embryonic fibroblast cells demonstrated an elevation of RRM1 and RRM2. Furthermore, down-regulation of mTOR-protein kinase B signaling or cyclin dependent kinase 4 led to decrease of RRM1 and RRM2 mRNAs. In addition, TP53 mutant cancer cells had elevation of RRM1 and RRM2, which was reduced by rapamycin. Importantly, human double minute 2 inhibitor nutlin-3 decreased RRM1 and RRM2 in TP53 wild type rhabdomyosarcoma Rh18 but not in TP53 mutated Rh30 cells. Our data demonstrated that mTOR enhances the cap-dependent protein translation and gene transcription of RRM1 and RRM2. Our findings might provide an additional mechanism by which p53 maintains genome stability.
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