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. This interferes with DNA/RNA biosynthesis and DNA repair. Because of this critical metabolic function, down-regulation of FDH in cancer cells would 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.

Public Health Relevance

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.

National Institute of Health (NIH)
National Cancer Institute (NCI)
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Special Emphasis Panel (ZRG1-OBT-S (02))
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Arya, Suresh
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Medical University of South Carolina
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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
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
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
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
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
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
Krupenko, Sergey A (2009) FDH: an aldehyde dehydrogenase fusion enzyme in folate metabolism. Chem Biol Interact 178:84-93
Ghose, Sampa; Oleinik, Natalia V; Krupenko, Natalia I et al. (2009) 10-formyltetrahydrofolate dehydrogenase-induced c-Jun-NH2-kinase pathways diverge at the c-Jun-NH2-kinase substrate level in cells with different p53 status. Mol Cancer Res 7:99-107
Cahoy, John D; Emery, Ben; Kaushal, Amit et al. (2008) A transcriptome database for astrocytes, neurons, and oligodendrocytes: a new resource for understanding brain development and function. J Neurosci 28:264-78
Oleinik, N V; Krupenko, N I; Krupenko, S A (2007) Cooperation between JNK1 and JNK2 in activation of p53 apoptotic pathway. Oncogene 26:7222-30

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