The overall goal of this proposal is to characterize the novel tumor suppressor activity of a key metabolic enzyme and determine the mechanisms transforming metabolic effects into regulation of proliferation. FDH (10-formyltetrahydrofolate dehydrogenase) irreversibly converts 10-formyltetrahydrofolate, an essential substrate for de novo purine biosynthesis, to tetrahydrofolate. Through depletion of this substrate, FDH can restrict purine biosynthesis. In turn, this interferes with important downstream cellular processes, including DNA/RNA biosynthesis and DNA repair. Because of this critical metabolic function, down-regulation of FDH in cancer cells was predicted to be pro-survival. Indeed, we have initially made the important observation that FDH is strongly and ubiquitously down-regulated in tumors through the promoter hypermethylation. We have further demonstrated that moderate FDH expression in FDH-deficient cancer cells induces apoptotic cell death. In contrast, non-cancer cells are insensitive to high levels of the enzyme. Therefore, it is proposed that cancer cells silence the FDH gene in order to escape cytotoxicity. Studies of phenotypic effects upon reactivation of normal FDH expression in FDH-deficient tumor cells have further explored JNK1/2 and p53 as key components of FDH-induced apoptotic signaling, and determined DHFR and folate supplementation as proliferation rescue factors. Importantly, a novel pathway linking FDH, through intracellular folate regulation, to control of cell motility, was discovered. The current proposal extends previous studies of antiproliferative mechanisms of FDH, and related folates, to direct interaction with p53 and evaluation of its role in vivo in mouse model. Our central hypothesis is that FDH down-regulation through promoter hypermethylation is one of the important means by which malignancies gain pro-survival advantage over normal cells. We further suggest that FDH exerts its regulatory effects through multiple mechanisms.
The Specific Aims to probe these mechanisms and test our hypothesis are: (1) Investigate the functional interaction of FDH with p53 tumor suppressor protein in cytoplasm. (2) Determine the impact of FDH silencing on tumor initiation/progression using FDH deficient mouse model. Investigation of the critical role of FDH, in cancer cell survival/induction of folate stress at the onset of the disease, will provide important insight into the malignant process itself and link deregulation of key metabolic pathways to cancer disease, as well as establish new targets for diagnostics of the malignant transformation.
Folate, an important and essential part of the human diet, regulates many cellular processes including nucleotide biosynthesis and methylation, while folate deficiency promotes many diseases. This application is focused on a novel tumor suppressor function of FDH, an abundant human enzyme, which is an important regulator of folate pathways. Since FDH can function as a restrictor of excessive proliferation, understanding its role in cellular metabolism will provide better understanding of tumorigenic processes.
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|Prakasam, A; Ghose, S; Oleinik, N V et al. (2014) JNK1/2 regulate Bid by direct phosphorylation at Thr59 in response to ALDH1L1. Cell Death Dis 5:e1358|
|Oleinik, Natalia V; Helke, Kristi L; Kistner-Griffin, Emily et al. (2014) Rho GTPases RhoA and Rac1 mediate effects of dietary folate on metastatic potential of A549 cancer cells through the control of cofilin phosphorylation. J Biol Chem 289:26383-94|
|DebRoy, Suchandra; Kramarenko, Inga I; Ghose, Sampa et al. (2013) A novel tumor suppressor function of glycine N-methyltransferase is independent of its catalytic activity but requires nuclear localization. PLoS One 8:e70062|
|Strickland, Kyle C; Krupenko, Natalia I; Krupenko, Sergey A (2013) Molecular mechanisms underlying the potentially adverse effects of folate. Clin Chem Lab Med 51:607-16|
|Hoeferlin, L Alexis; Fekry, Baharan; Ogretmen, Besim et al. (2013) Folate stress induces apoptosis via p53-dependent de novo ceramide synthesis and up-regulation of ceramide synthase 6. J Biol Chem 288:12880-90|
|Oleinik, Natalia V; Krupenko, Natalia I; Krupenko, Sergey A (2011) Epigenetic Silencing of ALDH1L1, a Metabolic Regulator of Cellular Proliferation, in Cancers. Genes Cancer 2:130-9|
|Hoeferlin, L Alexis; Oleinik, Natalia V; Krupenko, Natalia I et al. (2011) Activation of p21-Dependent G1/G2 Arrest in the Absence of DNA Damage as an Antiapoptotic Response to Metabolic Stress. Genes Cancer 2:889-99|
|Oleinik, N V; Krupenko, N I; Krupenko, S A (2010) ALDH1L1 inhibits cell motility via dephosphorylation of cofilin by PP1 and PP2A. Oncogene 29:6233-44|
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