The goal of this project is to elucidate basic mechanisms of eukaryotic translation initiation using structural and other biochemical methods. Eukaryotic translation initiation is highly regulated by elements of the untranslated regions of mRNAs, 5'UTR and 3'UTR, by the cellular concentration of initiation factors, by the action of regulatory proteins and by signaling pathways that are initiated by external messages or cellular events. Dysregulation of translation initiation by elevated levels of initiation factors is found in many forms of cancer. Thus, correcting for out-of-balance initiation with small molecule agents is a promising new route for cancer therapy. The proposed research is focused on the mechanisms by which the small ribosomal particle is recruited to mRNA, mediated by the initiation factors eIF4E, eIF4G and the regulatory phosphoprotein 4EBP-1. The second aspect is focused on scanning of the pre-initiation complex to the AUG initiation codon, aided by the interplay between the initiation factors eIF4A, eIF4G and eIF4H. The third topic is on elucidating processes by which the factors eIF1, eIF1A, eIF5 and eIF22 help recruiting the initiator Met-tRNAMet to the ribosome and warrant the correct placement. A significant aspect is to discover and characterize inhibitors of initiation with the goal of developing anti-cancer agents with broad specificity. The research will pursue three specific aims: 1. Define the regulation of eIF4E by tumor-suppressive proteins and small molecules 2. Structure of the eIF4G/eIF4A/eIF4H complex and approaches for inhibition 3. Control of start-codon selection by interactions of eIF5, eIF1, eIF1A and eIF2

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Using structural methods we will study mechanisms of eukaryotic translation initiation. We will investigate routes for inhibiting this process to develop novel anti-tumor agents. We also will study the complex of the scaffold protein eIF4G with the RNA helicase eIF4A and the interaction of factors eIF5, eIF1 and eIF1A, which support start codon recognition by interacting with eIF2.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Research Project (R01)
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Macromolecular Structure and Function C Study Section (MSFC)
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Knowlton, John R
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Harvard University
Schools of Medicine
United States
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Edmonds, Katherine A; Wagner, Gerhard (2015) (1)H, (13)C, and (15)N backbone and sidechain chemical shift assignments for the HEAT2 domain of human eIF4GI. Biomol NMR Assign 9:157-60
Luna, Rafael E; Akabayov, Sabine R; Ziarek, Joshua J et al. (2014) Examining weak protein-protein interactions in start codon recognition via NMR spectroscopy. FEBS J 281:1965-73
Ivanov, Pavel; O'Day, Elizabeth; Emara, Mohamed M et al. (2014) G-quadruplex structures contribute to the neuroprotective effects of angiogenin-induced tRNA fragments. Proc Natl Acad Sci U S A 111:18201-6
Papadopoulos, Evangelos; Jenni, Simon; Kabha, Eihab et al. (2014) Structure of the eukaryotic translation initiation factor eIF4E in complex with 4EGI-1 reveals an allosteric mechanism for dissociating eIF4G. Proc Natl Acad Sci U S A 111:E3187-95
Yi, Tingfang; Kabha, Eihab; Papadopoulos, Evangelos et al. (2014) 4EGI-1 targets breast cancer stem cells by selective inhibition of translation that persists in CSC maintenance, proliferation and metastasis. Oncotarget 5:6028-37
Akabayov, Sabine R; Akabayov, Barak; Wagner, Gerhard (2014) Human translation initiation factor eIF4G1 possesses a low-affinity ATP binding site facing the ATP-binding cleft of eIF4A in the eIF4G/eIF4A complex. Biochemistry 53:6422-5
Yi, Tingfang; Zhai, Bo; Yu, Yonghao et al. (2014) Quantitative phosphoproteomic analysis reveals system-wide signaling pathways downstream of SDF-1/CXCR4 in breast cancer stem cells. Proc Natl Acad Sci U S A 111:E2182-90
Akabayov, Sabine R; Wagner, Gerhard (2014) Backbone resonance assignment of the HEAT1-domain of the human eukaryotic translation initiation factor 4GI. Biomol NMR Assign 8:89-91
Martin-Marcos, Pilar; Nanda, Jagpreet S; Luna, Rafael E et al. (2014) Enhanced eIF1 binding to the 40S ribosome impedes conformational rearrangements of the preinitiation complex and elevates initiation accuracy. RNA 20:150-67
Yi, Tingfang; Papadopoulos, Evangelos; Hagner, Patrick R et al. (2013) Hypoxia-inducible factor-1ýý (HIF-1ýý) promotes cap-dependent translation of selective mRNAs through up-regulating initiation factor eIF4E1 in breast cancer cells under hypoxia conditions. J Biol Chem 288:18732-42

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