Regulation of OPA1-mediated mitochondrial fusion by leucine zipper/EF-hand-containing transmembrane protein-1 plays a role in apoptosis Longzhen Piao a, 1 , Yuwen Li a , Soung Jung Kim b , Kyung-Cheol Sohn c , Keum-Jin Yang a , Kyeong Ah Park a , Hee Sun Byun a , Minho Won a , Janghee Hong a , Gang Min Hur a , Jeong Ho Seok a , Minho Shong b , Ragna Sack d , Derek P. Brazil e , Brian A. Hemmings d , Jongsun Park a, ⁎ a Department of Pharmacology, Daejeon Regional Cancer Center, Cancer Research Institute, Research Institute for Medical Sciences, College of Medicine, Chungnam National University, Taejeon, 301-131, Republic of Korea b Department of Internal Medicine, College of Medicine, Chungnam National University, Taejeon, 301-131, Republic of Korea c Department of Dermatology, College of Medicine, Chungnam National University, Taejeon, 301-131, Republic of Korea d Friedrich Miescher Institute for Biomedical Research, Maulbeerstrasse 66, CH-4058 Basel, Switzerland e UCD Diabetes Research Centre, UCD Conway Institute, University College Dublin, Belfield, Dublin 4, Ireland abstract article info Article history: Received 30 September 2008 Received in revised form 18 December 2008 Accepted 5 January 2009 Available online 9 January 2009 Keywords: CTMP Mitochondria Apoptosis Wolf–Hirschhorn syndrome LETM11 Carboxyl-terminal modulator protein (CTMP) is a tumor suppressor-like binding partner of Protein kinase B (PKB/Akt) that negative regulates this kinase. In the course of our recent work, we identified that CTMP is consistently associated with leucine zipper/EF-hand-containing transmembrane-1 (LETM1). Here, we report that adenovirus-LETM1 increased the sensitivity of HeLa cells to apoptosis, induced by either staurosporine or actinomycin D. As shown previously, LETM1 localized to the inner mitochondrial membrane. Electron- microscopy analysis of adenovirus-LETM1 transduced cells revealed that mitochondrial cristae were swollen in these cells, a phenotype similar to that observed in optic atrophy type-1 (OPA1)-ablated cells. OPA1 cleavage was increased in LETM1-overexpressing cells, and this phenotype was reversed by overexpression of OPA1 variant-7, a cleavage resistant form of OPA1. Taken together, these data suggest that LETM1 is a novel binding partner for CTMP that may play an important role in mitochondrial fragmentation via OPA1- cleavage. © 2009 Elsevier Inc. All rights reserved. 1. Introduction Carboxyl-terminal modulator protein (CTMP) is a recently identi- fied binding partner for Protein kinase B (PKB/Akt) [1]. Overexpression of CTMP results in inactivation of PKB in cultured cells [2], in a K-ras- induced lung cancer mouse model [3] and in v-Akt transformed cells transplanted to mice [1]. A role for CTMP-mediated inhibition of PKB was supported by a recent report demonstrating that the expression level of CTMP was inhibited by hypermethylation of it's promoter in malignant glioblastomas where PKB activity is frequently elevated [4]. CTMP has been detected at the mitochondria in HeLa cells (Hemmings et al., unpublished data) and becomes phosphorylated upon treatment of cells with pervanadate [1], indicating that CTMP is regulated by post-translational modification. Mitochondria are essential organelles in most eukaryotic cells. Their most well-known biochemical functions are in intermediary metabolism and respiration, which result in the generation of adenosine triphosphate (ATP) through oxidative phosphorylation. In addition, they play important roles in apoptosis, cell signaling, iron metabolism, and steroidogenesis [5]. Alterations in mitochondrial function are responsible for a range of inherited and acquired human diseases and are implicated in the aging process [6]. Recent studies indicate that the functions of mitochondria are coordinated with their dynamic behavior [7]. Mito- chondria frequently fuse and divide, and the balance of these processes determines overall mitochondrial morphology. When mitochondrial fusion is reduced, mitochondria fragment due to ongoing fission; conversely, mitochondria are long and overly interconnected when this balance shifts towards fusion [5]. OPA1 is the human homologue of yeast dynamin-related GTP-binding protein Mgm1, which is involved in mitochondrial fusion [8]. Data from OPA1 mutant patients highlight a potential role for OPA1 in mitochondrial maintenance [9]. In such patients, mitochondrial DNA content is lower, oxidative phosphorylation in the calf muscle is defective, and the structure of the mitochondrial network in monocytes is altered compared to normal control subjects [9]. OPA1 knockdown studies provide evidence to support a functional role for this protein in mitochondrial morphology maintenance [8,10]. Cellular Signalling 21 (2009) 767–777 ⁎ Corresponding author. Department of Pharmacology, College of Medicine, Chungnam National University, 6 Munhwa-Dong, Jung-Gu, Taejeon, 301-131, Republic of Korea. Tel.: +82 42 580 8252; fax: +82 42 585 6627. E-mail address: insulin@cnu.ac.kr (J. Park). 1 Current address: Department of Endocrinology, Yanbian University Hospital, Yanji 133000, China. 0898-6568/$ – see front matter © 2009 Elsevier Inc. All rights reserved. doi:10.1016/j.cellsig.2009.01.020 Contents lists available at ScienceDirect Cellular Signalling journal homepage: www.elsevier.com/locate/cellsig