Contents lists available at ScienceDirect Mitochondrion journal homepage: www.elsevier.com/locate/mito Mitochondrial ND5 mutation mediated elevated ROS regulates apoptotic pathway epigenetically in a P53 dependent manner for generating pro- cancerous phenotypes Rajnish Kumar Singh 1 , Sunil Saini, Dileep Verma, Ponnusamy Kalaiarasan, Rameshwar N.K. Bamezai ⁎ National Centre of Applied Human Genetics, School of life Sciences, Jawaharlal Nehru University, New Delhi 110067, India ARTICLE INFO Keywords: mtND5 Mitochondrial DNA mutation Reactive oxygen species Apoptotic pathway Differential methylation ABSTRACT We have previously observed concomitant events of mutations in mitochondrial and nuclear genes, along with elevated reactive oxygen species (ROS) and differential methylation within the promoters of nuclear genes in tumors and in vitro experiments of tumorigenesis. These observations have made it pertinent to replicate and understand the role of acquired mitochondrial condition in tuning a cell to accomplish a pro-cancerous state. Using a codon optimized vector system for exogenous over-expression and mitochondrial localization; we have characterized here the role of over-expressed wild type mtND5 and one of its non-synonymous somatic mutation, ND5:P265H. The ectopically over-expressed ND5:P265H in mitochondria resulted in a reduced Complex I activity, generation of higher ADP/ATP ratio, reactive oxygen species (ROS) and carbonylation of proteins as compared to mock-transfected cells. Cells over-expressing mtND5 variant produced both peroxide as well as super-oxide ROS; the generation of which was dependent on the functional status of P53; modulating epigenetically the expression of key apoptosis pathway genes. The pro-cancerous phenotypes, of anchorage dependent and independent growth; increased glucose uptake and lactate production, were selectively observed only in P53 non-functional cells over-expressing mutant ND5:P265H. We propose that somatic mutation in mtND5 resulting in down-regulated complex I enzyme activity, elevated ROS and up-regulation of a set of nuclear anti-apoptotic genes epigenetically in the P53 dysfunctional cellular background, has provided a unique understanding of the molecular mechanism of mitochondrial mutation; and the concomitant existence of somatically acquired mitochondrial and nuclear p53 mutations, in cancer progression and promotion. 1. Introduction Mutations in mitochondrial DNA prevail in human populations with a high over-representation in diseases and are proposed to be under purifying selection (Ye et al., 2014). Amongst 13 polypeptides synthe- sized within mitochondria, NADH ubiquinone Oxidoreductase subunit 5 (ND5 or MT-ND5, EC 1.6.5.3) is encoded by the largest gene located in the guanine rich heavy chain of mitochondrial DNA (mtDNA co- ordinates: 12337–14148). ND5 protein is the second largest subunit of mitochondrial electron transport chain Complex I (1812 bp long, 603 AA, 67 Kda protein) after NDUSF1 (2184 bp long, 727 AA, 75 Kda protein), and also a probable hydrophobic protein fragment component of Complex I (Anderson et al., 1981; Hirst, 2013; Mimaki et al., 2012). The subunit is considered as a hot-spot for multiple atypical deletions and somatic mutations, which are associated with neuro-gastrointestinal-encephalo-myopathy (Nishigaki et al., 2004) and several complex disorders, such as Leber's hereditary optic neuropathy (Liu et al., 2011; Mayorov et al., 2005), MELAS syndrome (Naini et al., 2005; Zhao et al., 2011), Leigh syndrome (Petruzzella et al., 2003; Shanske et al., 2008; Taylor et al., 2002) and various type of cancers (Gochhait et al., 2008; Larman et al., 2012; Parrella et al., 2001; Shen et al., 2011; Yin et al., 2010). The proposed role of deletions or mutations in the mitochondrial DNA coded genes in pathogenesis has always been considered to be due to altered ROS related metabolism (Lu et al., 2009; Sharma et al., 2011). Unlike mutations within nuclear DNA coded genes, only a handful of studies till date have been conducted to understand and validate the functional role of mitochon- drial mutations prevalent in different cancers. Most of the earlier http://dx.doi.org/10.1016/j.mito.2017.05.001 Received 11 April 2017; Received in revised form 2 May 2017; Accepted 8 May 2017 ⁎ Corresponding author. 1 Current address: Department of Microbiology and Tumor Virology Program of the Abramson Cancer Center, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania – 19,104. E-mail address: bamezai@mail.jnu.ac.in (R.N.K. Bamezai). Mitochondrion xxx (xxxx) xxx–xxx 1567-7249/ © 2017 Published by Elsevier B.V. Please cite this article as: Singh, R.K., Mitochondrion (2017), http://dx.doi.org/10.1016/j.mito.2017.05.001