C OMMUNICATION AIMP3/p18 Controls Translational Initiation by Mediating the Delivery of Charged Initiator tRNA to Initiation Complex Taehee Kang 1 †, Nam Hoon Kwon 1 †, Jin Young Lee 1 , Min Chul Park 1 , Eunji Kang 1 , Hyo Hyun Kim 1 , Taek Jin Kang 2 and Sunghoon Kim 1,3 ⁎ 1 Medicinal Bioconvergence Research Center, Seoul National University, Seoul 151–742, Korea 2 Department of Chemical and Biochemical Engineering, Dongguk University-Seoul, Seoul 100–715, Korea 3 WCU Department of Molecular Medicine and Biopharmaceutical Sciences, Graduate School of Convergence Science and Technology, Seoul National University, Suwon 443–270, Korea Received 17 May 2012; received in revised form 11 July 2012; accepted 17 July 2012 Available online 4 August 2012 Edited by R. L. Gonzales Keywords: AIMP3; translation initiation; methionine; initiator tRNA; eukaryotic initiation factor 2 γ subunit (eIF2γ) Aminoacyl-tRNA synthetase-interacting multifunctional proteins (AIMPs) are nonenzymatic scaffolding proteins that comprise multisynthetase complex (MSC) with nine aminoacyl-tRNA synthetases in higher eukary- otes. Among the three AIMPs, AIMP3/p18 is strongly anchored to methionyl-tRNA synthetase (MRS) in the MSC. MRS attaches methionine (Met) to initiator tRNA (tRNA i Met ) and plays an important role in translation initiation. It is known that AIMP3 is dispatched to nucleus or nuclear membrane to induce DNA damage response or senescence; however, the role of AIMP3 in translation as a component of MSC and the meaning of its interaction with MRS are still unclear. Herein, we observed that AIMP3 specifically interacted with Met-tRNA i Met in vitro, while it showed little or reduced interaction with unacylated or lysine- charged tRNA i Met . In addition, AIMP3 discriminates Met-tRNA i Met from Met-charged elongator tRNA based on filter-binding assay. Pull‐down assay revealed that AIMP3 and MRS had noncompetitive interaction with eukaryotic initiation factor 2 (eIF2) γ subunit (eIF2γ), which is in charge of binding with Met-tRNA i Met for the delivery of Met-tRNA i Met to ribosome. AIMP3 recruited active eIF2γ to the MRS–AIMP3 complex, and the level of Met-tRNA i Met bound to eIF2 complex was reduced by AIMP3 knockdown resulting in reduced protein synthesis. All these results suggested the novel function of AIMP3 as a critical mediator of Met-tRNA i Met transfer from MRS to eIF2 complex for the accurate and efficient translation initiation. © 2012 Elsevier Ltd. All rights reserved. *Corresponding author. Medicinal Bioconvergence Research Center, Seoul National University, Seoul 151-742, Korea. E-mail address: sungkim@snu.ac.kr. † T.K. and N.H.K. contributed equally to this work. Abbreviations used: MRS, methionyl-tRNA synthetase; eIF2γ, eIF2 γ subunit; ARS, aminoacyl-tRNA synthetase; MSC, multisynthetase complex; AIMP, ARS-interacting multifunctional protein; eEF, eukaryotic elongation factor; eIF2, eukaryotic initiation factor 2; TC, ternary complex; MEF, mouse embryonic fibroblast; WT, wild type; GST, glutathione S-transferase. http://dx.doi.org/10.1016/j.jmb.2012.07.020 J. Mol. Biol. (2012) 423, 475–481 Contents lists available at www.sciencedirect.com Journal of Molecular Biology journal homepage: http://ees.elsevier.com.jmb 0022-2836/$ - see front matter © 2012 Elsevier Ltd. All rights reserved.