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Animal Mitochondria: Evolution, Function, and Disease M. Tao1,§, C.-P. You2,§, R.-R. Zhao1,§, S.-J. Liu*,1, Z.-H. Zhang1, C. Zhang1 and Y. Liu1 1

Key Laboratory of Protein Chemistry and Fish Developmental Biology of Education Ministry of China, College of Life Sciences, Hunan Normal University, Changsha, Hunan 410081, China 2

Central Laboratory, Linyi People’s Hospital, Linyi, Shandong 276000, China Abstract: Mitochondria are sub-cellular organelles responsible for producing the majority of cellular energy through the process of oxidative phosphorylation (OXPHOS), and are found in nearly all eukaryotic cells. Mitochondria have a unique genetic system, mitochondrial DNA (mtDNA), which is a small, self-replicating and diverse genome. In the past 30 years, mtDNA has made significant contribution to molecular ecology and phylogeography. Mitochondria also represent a unique system of mitochondrial–nuclear genomic cooperation. Additionally, mitochondrial dysfunction can be fatal. In this paper, we review several aspects of mitochondria, including evolution and the origin of mitochondria, energy supply and the central role of mitochondria in apoptosis, and mitochondrial dysfunction. It is shown that mitochondria play a critical role in many aspects of life.

Keywords: Apoptosis, mitochondria, mtDNA, OXPHOS.

INTRODUCTION Mitochondria were originally observed in living cells using light microscopy in the 1840s [1] and Carl Benda firstly named them mitochondria [2]. In the 1930s, Hans Krebs first studied the formulation of the urea and tricarboxylic acid cycle (TCA). In the 1950s, mitochondria were found to be the site of respiration and oxidative phosphorylation (OXPHOS) [3]. Margit M.K. Nass and Sylvan Nass first discovered DNA in the mitochondria in the 1960s [4]. In 1981, Anderson et al. published the sequence and organization of the human mitochondrial genome [5]. This was the first mitochondrial genome to be sequenced and it was 16,569bp long. Since then, over the next 30 years, the mitochondrial genes had been widely used as pivotal tool in phylogeography, evolution, population genetics, and phylogenetic study. The mitochondrial DNA (mtDNA) mutations which initially discovered in mitochondrial myopathy and Leber’s hereditary optic atrophy were reported in 1988, indicating a role of mitochondrial dysfunction in rare metabolic disorders [6, 7]. Now, we know that mitochondria are tiny (membrane-enclosed) organelles inside nearly all eukaryotic cells, which comprise ≈1500 proteins; they generate almost all the energy for the life in the form of ATP; they have their own genetic material, mitochondrial DNA (mtDNA); they are dynamic and constantly undergo fission and fusion, depending on different tissues and conditions; they represent a unique system of mitochondrial–nuclear genomic *Address correspondence to this author at the Key Laboratory of Protein Chemistry & Developmental Biology of State Education Ministry of China, College of Life Sciences, Hunan Normal University, Changsha, Hunan 410081, China; Tel: +86-73188873010; Fax: +86-73188873074; E-mail: [email protected]

cooperation. Nucleus-encoded proteins assemble with or bind to mitochondrion-encoded protein peptides in OXPHOS complexes, and nuclear proteins are involved in mtDNA replication, transcription and translation [8]. The mitochondria are involved in the whole progress of life including energy, cell signaling, cell differentiation, cell cycle and growth, aging and death.

ORIGIN HYPOTHESES The integral part that mitochondria play in many aspects of eukaryote biology might well reflect their role in the origin of eukaryotes themselves. The hypothesis pertaining to an endosymbiotic origin of the mitochondria was widely accepted. The confidence in this theory was then strengthened by the discovery of mitochondrial genomes and the results of phylogenetic reconstructions with sequences for rRNA as well as for a few proteins. There is strong evidence to support the hypothesis that mitochondria were once free-living alpha-proteobacterial [9-11], however, the exact position of the mitochondria within the alphaproteobacteria is still argued [12]. One study has pointed outside the Rickettsiales to Rhodospirillum [13], but most have placed the mitochondrial ancestor within or basal to the Rickettsiales, some specifically within the Rickettsiaceae [14], and others, as a sister to the combined Rickettsiaceae and Anaplasmataceae [11]. Following the acquisition of mitochondria, there have been complex patterns of gene retention, gene transfer to nucleus, and redirection of non-alphaproteobacterial proteins to the mitochondrial proteome [15]. The acquisition of the mitochondrion during eukaryotic evolution may have been a central catalyst allowing further development of the many unique features of eukaryotic cells.

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Mitochondrial DNA (mtDNA)

Inheritance

MtDNA was formally identified and localized in the 1960s [3, 16, 17], but in the 1950s, researchers found some mitochondrial characteristics in yeast that were inherited from a cytoplasmic fashion. In 1972, Dawid & Blackler provided the first evidence for uniparental, the maternal inheritance of mtDNA in animals [18]. Then, the concept of strict maternal inheritance in animals gained popularity and gained further support in the 1980s [17].

In nearly all eukaryotes, the theory that mitochondrial genes are inherited uniparentally through the maternal germline has been widely accepted, but the mechanism for this phenomenon is not well understood. The most popular explanation is simple dilution of paternal mtDNA. The number of mitochondria in paternal gametes is less than that in the maternal gametes, which makes paternal mtDNA prone to loss by drift. For example, in mammalian female gametocytes, mtDNA copies can outnumber 3 4 their male counterparts to the order of 10 to 10 . Furthermore, the mitochondrial bottleneck and the suspension of mtDNA replication in the fertilized oocyte amplify the effects of genetic drift of paternal lineages. Recent studies reported other mechanisms in different species. For example, elimination of sperm mitochondria within the egg cytoplasm has been shown in mice and hamsters [19]. By using highly sensitive SYBR Green I vital staining to visualize mitochondrial

In animals, mtDNA is typically a single circular chromosome that is approximately 16 kb long (Fig. 1). For example, there are 16 569 base pairs in the human mitochondrial genome. MtDNA has 37 genes, including: (i) 13 protein-coding genes for subunits of respiratory complexes I, III, IV and V; (ii) two rRNAs of the mitochondrial ribosome; (iii) 22 tRNAs necessary for protein translation. MtDNA has some unique characteristics: maternal inheritance, high mutation rate, and nonrecombination.

Fig. (1). The circular map of a typical animal mtDNA. MtDNA has 37 genes, including: (i) 13 protein-coding genes for subunits of respiratory complexes I, III, IV and V，including：ND1-6, ND4L, cytochrome b, cytochrome c, oxidase 1-3, ATP6 and ATP 8 ; (ii) two rRNAs ：12SrRNA and 16SrRNA; (ⅲ) 22 tRNAs necessary for protein translation, including: tRNA-Phe (P), tRNA-Val (V), 2 tRNA-Leu (L), tRNA-Ile (I), tRNA-Gln (Q), tRNA-Met (M), tRNA-Trp (W), tRNA-Ala (A), tRNA-Asn (N), tRNA-Cys (C), tRNA-Tyr (Y), 2 tRNA-Ser (S), tRNA-Asp (D), tRNA-Lys (K), tRNA-Gly (G), tRNA-Arg (R), tRNA-His (H), tRNA-Glu (E), tRNA-Thr (T), and tRNA-Pro (P).

Animal Mitochondria: Evolution, Function, and Disease

nucleoids, Nishimura et al. [20] found that elimination of sperm mtDNA upon fertilization in Japanese Medaka is achieved by two steps: gradual decrease of mitochondrial nucleoid numbers during spermatogenesis and rapid digestion of sperm mtDNA just after fertilization. In cattle and primates, the elimination process involves ubiquitination of the sperm mitochondria outer membrane [21]. Down-regulation of mitochondrial transcription factors during spermatogenesis in humans suggests a mechanism for reducing mtDNA copy number during spermatogenesis, and has implications for our understanding of maternal transmission of mtDNA [22]. These studies have provided significant information for understanding the maternal inheritance of mtDNA, but they only involved a few animal species. Thus, it will be necessary to compare and contrast various eukaryotes to understand the mechanisms and significance of maternal inheritance. Although mitochondrial genes are transmitted to progeny mainly from the maternal parent, paternal inheritance has been shown to occur at low frequency [23-26]. Paternal inheritance of mtDNA includes two patterns: recombination and paternal leakage. MtDNA recombination is a long-standing topic, but rare cases referring to mtDNA recombination have been documented, and most of them have occurred in the laboratory, such as in fungi [26]. Many studies have shown that mtDNA does not recombine in animals, while there is indisputably mitochondrial recombination in mussels [25]. The presence of enzymes necessary for homologous recombination in human mitochondria [27] also suggests the possibility of recombination in animal mtDNA. The paternal mitochondria are not always eliminated during egg fertilization, which leads to paternal leakage. Most biparental inheritance events result from paternal leakage. These events have been observed in several species including mussels, Drosophila, birds, lizards, mice and humans [28-32]. Mutation It is believed that the mutation rate of mtDNA is 5– 10 times higher than that of nuclear DNA [33]. Low efficiency of DNA repair pathways and a more mutagenic intracellular environment could be responsible for the high mutation rate of mtDNA. HaagLiautard et al. [34] detected a total of 28 point mutations and eight insertion–deletion mutations when they scanned the mitochondrial genome of Drosophila melanogaster that had undergone approximately 200 generations. In the mutational hotspots, such as control region of human mtDNA, the mutation rate was around one mutation every 25–61 generations [33, 35, 36]. Besides, the mtDNA mutation rate seems to vary among animal lineages, and it is also dissimilar in different nucleotide regions [37-39]. The high mutation rate and maternal inheritance have made mtDNA useful in evolution and phylogenetic study. Based on the polymorphism (mainly caused by mutation) of mtDNA sequences, researchers could reconstruct the history of species, trace their origin, and provide information for ecology. However, it has


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recently been suggested that the pervasive nature of direct and indirect selection on mtDNA renders any conclusion derived from it ambiguous [40]. In humans, mtDNA mutations probably lead to disease, because mtDNA encodes essential subunits of the mitochondrial respiratory chain and its defects would result in impaired OXPHOS. To date, more than 250 pathogenic mutations in mtDNA have been described [41].

MITOCHONDRIAL FUNCTION Energy By the process of OXPHOS (Fig. 2), mitochondria can produce ATP. Pyruvates (produced by glycolysis) and fatty acids in the cytoplasm are actively transported across the inner mitochondrial membrane and into the matrix where they are oxidized to form acetyl-CoA and NADH. Through TCA, acetyl-CoA is oxidized into CO2, and NADH and FADH2 are produced, which contain redox energy. The redox energy from NADH and FADH2 can be transferred to oxygen in several steps via the electron transport chain, and in the process, ADP and inorganic phosphate (Pi) are synthesized into ATP. This process of energy transfer and formation of ATP is called OXPHOS. The electron transport chain consists of five complexes in the inner mitochondrial membrane: complexes I–V. In most animals, OXPHOS protein complexes comprise almost 90 subunits, most of which are encoded by the nuclear genome and only 13 proteins which are encoded by mtDNA, including: seven subunits of complex I (ND1–6, ND4L), one of complex III (cytochrome b), three of complex IV (cytochrome c oxidase 1–3), and two of complex V (ATP6 and 8). In complex I (NADH dehydrogenase), two electrons are removed from NADH and transferred to a lipid-soluble carrier, ubiquinone; then they are transferred to cytochrome c in complex III (cytochrome bc1 complex), and ultimately to oxygen in complex IV (cytochrome c oxidase) to form water. Complex V is ATP synthase contained in the inner membrane. Following the transfer of electrons, the protons are pumped into the intermembrane space. The resulting electrochemical proton gradient is used to generate chemical energy in the form of ATP in complex V. Meanwhile, in the process of OXPHOS, mitochondria also produce substantial amounts of reactive oxygen species (ROS), such as the superoxide radical, O2¯, H2O2, and HO¯ [42, 43]. It was estimated that about 2% of the oxygen consumed by isolating mitochondria from mammalian tissues is converted to ROS. These species are natural byproducts of the normal metabolism of oxygen and they can damage proteins, membranes, and DNA, thereby cause oxidative stress in the mitochondria, and may contribute to the decline in mitochondrial function associated with the aging process [43]. Apoptosis Apoptosis has a lot of pathways. There are at least two broad pathways: the extrinsic and intrinsic. Intrinsic

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Fig. (2). Complex I-V and the process of oxidative phosphorylation (OXPHOS).

or mitochondrion-dependent apoptosis is the most important pathway. It involves altering mitochondrial membrane permeability, Cytochrome-C (CytoC) release, formation of the apoptosome, and activation of caspase 3. The process of mitochondrial apoptosis is governed by pro- and antiapoptotic Bcl-2 family members [44]. The Bcl-2 family consists of anti- and proapoptotic members that have different Bcl-2 homology (BH) domains. Proapoptotic members can be divided into two groups: (i) the Bax-like group with BH1, BH2 and BH3 domains, including Bax, Bak and Bok; (ii) the BH3-only group with only BH3 domain such as Bad, Bid, Bim, Noxa and Puma. Antiapoptotic members, which contain Bcl-2, Bcl-xL, Bcl-w, and Bcl1, have all the homology domains BH1–4 [45-47]. In response to apoptotic stimuli, BH3-only proteins are activated (Fig. 3). They act upstream in the pathway and inhibit antiapoptotic Bcl-2 proteins by physically interacting with them, while Bax family proteins act downstream. The anti- and proapoptotic protein interaction leads to the formation of the membrane permeability transition pore (PTP). Through PTP, some small proapoptotic molecules such as cytochrome c [48], Smac/Diablo (second mitochondria-derived activator of caspase) [49], Omi/HtrA2 (high temperature requirement protein-A2) [50], apoptosis-inducing factor (AIF) [51], and endonuclease G (EndoG) [52] are released from the mitochondrial intermembrane space into the cytosol. The released Cytochrome-C (CytoC) binds to apoptotic protease activating factor (Apaf)-1, together with dATP and procaspase 9, to form the apoptosome, and thereby activates caspase 9. Caspase 9 activates caspase 3, which subsequently commits the cell to apoptosis. In addition, the inhibitors of apoptosis (IAPs) are inhibited by mitochondrially derived Smac/Diablo and Omi/HtrA2, and AIF and EndoG, causing chromosomal condensation and fragmentation. Mitochondrial Signaling The mitochondria play a critical role in cell regulatory and signaling events, in the responses of cells to a multiplicity of physiological and genetic

stresses, inter-organelle communication, cell proliferation and cell death. Mitochondrial signaling is an information channel between the mitochondrial respiratory chain and the nucleus for the signals transduction regarding the functional state of the mitochondria [53]. Michael J. Goldenthal et al. [54] suggested that the mitochondria act as a dynamic receiver and integrator of numerous translocated signaling proteins including protein kinases and 2+ fluxes and transcription factors, regulatory Ca membrane phospholipids as well the transmission of mitochondrially-generated oxidative stress and energyrelated signaling. Mitochondrial signaling factors include mitochondrial membrane potential (Ψm), mitochondrially-generated reactive oxygen species 2+ (ROS), Ca , free radical NO•, intracellular pH (pHi) and parameters connected with mitochondrial biogenesis. 2+

2+

Ca plays a central role in cellular signaling. Ca is a key regulator not only of multiple cytosolic enzymes, but also of a variety of metabolic pathways occurring within the lumen of intracellular organelles. 2+ Mitochondria play an important role in Ca storage and 2+ 2+ Ca homeostasis. Ca released either from intracellular stores or from outside the cell is taken up by mitochondria and stored transiently, and then released to the cytoplasmic compartment as part of the 2+ intracellular Ca traffic and signaling [55]. Increased 2+ results in elevated rate-limiting mitochondrial Ca activity levels of enzymes of several TCA which cause + increasing NADH/NAD ratios and ultimately leads to increasing mitochondrial ATP synthesis, and also directly modulates mitochondrial ATP synthase activity in cardiomyocytes [54, 56]. Mitochondrial bioenergetic activity generates ROS including superoxide and hydroxyl radicals and hydrogen peroxide (H2O2). Recent studies suggested that increased ROS generation and oxidative stress have a fundamental role in the regulation of cellular signaling and cytoprotection [54, 57]. Oxidative species (e.g. H2O2) can also function as a signal sent from mitochondria to other cellular sites rapidly and reversibly eliciting an array of intracellular cascades leading to different



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Fig. (3). The mitochondrial apoptosis in mammalian cells.

physiological end points for the cell (e.g. apoptosis, necrosis, cytoprotection, cell proliferation). Nitric oxide (NO) is also considered as an important intramitochondrial signaling molecule, which modulates mitochondrial respiration by direct binding to cytochrome C oxidase [58]. Furthermore, retrograde regulation is the general term for mitochondrial signaling, and is broadly defined as cellular responses to changes in the functional state of mitochondria. The nucleus possesses the majority of genetic information and controls most aspects of organelles including gene expression, growth, and development. In return, organelles also send signals back to regulate nuclear gene expression, a process defined as retrograde regulation [59]. Mitochondrial Involvement in Cell Differentiation During the last few years, a growing data have suggested that mitochondrion plays a significant role in cell differentiation [60]. Changes in mitochondrial abundance, morphology and functions were observed during the differentiation of different stem cells types, ranging from embryonic stem cells to induced pluripotent stem cells and somatic stem cells [61], and these changes are accompanied by an increase in ATP content and reactive oxygen species (ROS) levels. Studies using murine and human ESCs have revealed that under differentiation, ESCs develop numerous cristae, increase in number, and generate an extensive reticular network of tubular structure [62-64]. In addition, the mtDNA copy number is strictly regulated in order to generate appropriate levels of adenosine triphosphate (ATP) through OXPHOS to undertake their specific functions when stem cells undergo differentiation [65]. Dickinson et al. showed that human neural stem cells (hNSCs) increased their mtDNA content during differentiation and resulted in increased respiratory capacity [65]. These studies established the

importance of normal mitochondrial function in regulating differentiation, and demonstrated that differentiated cells display a more developed mitochondrial network to meet the energy needs.

MITOCHONDRIAL DYSFUNCTION AND DISEASE Mitochondrial Disease In addition to supplying energy for the cell, mitochondria are also involved in multiple cellular activities, e.g., supplying most energy of cells, regulating the process of apoptosis, aging and growth. Thus, the dysfunction of mitochondria could result in a wide range of human mitochondrial diseases. The term ‘mitochondrial disease’ refers to any disorder affecting the respiratory chain and OXPHOS system, a series of five multisubunit enzyme complexes (complexes I–V) embedded in the inner mitochondrial membrane [66]. Mitochondrial diseases are common, with a combined prevalence of one in 5000. Mitochondrial diseases are generally caused by mutations in mtDNA or in nuclear genes that code mitochondrial proteins. Mutations of mtDNA are a primary cause and can be due to inherited sequence changes in the mtDNA genome (eg, deletions, rearrangements, point mutations), secondary to nuclear gene mutations causing, for example, mtDNA multiple deletions or depletion, as result of free-radicalmediated damage or faulty repair. It is reported that mutations causing mitochondrial dysfunction have been found in all 37 mitochondrial encoded genes and more than 80 nuclear genes [66]. Nerve cells in the brain and muscles require a great deal of energy, and thus appear to be particularly damaged when mitochondrial dysfunction occurs, such as the typical mitochondrial diseases that have neuromuscular disease symptoms which are often called mitochondrial myopathy including mitochondrial encephalomyopathy, lactic

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acidosis, stroke-like episodes (MELAS), Myoclonic epilepsy and ragged-red fibers (MERRF), KearnsSayre syndrome (KSS), Chronic progressive external ophthalmoplegia (CPEO) and so on. Wild-type mtDNA can coexist with mutant mtDNA in one cell, which is known as heteroplasmy. These two kinds of mtDNA are distributed randomly with mitochondria into progeny oocytes in oogenesis and then distributed into somatic cells in process of embryogenesis, which causes variable levels of mutant mtDNA in different cells, tissues, and organs. The mitochondrial dysfunction becomes clinically apparent when mutant mtDNA in one cell reaches a certain level (phenotypic threshold) [67, 68]. The mtDNA disorders occur spontaneously and relatively often because mtDNA mutations occur rapidly. mtDNA-related disorders can be divided into two main groups: (i) those due to mutations in genes involved in mitochondrial protein synthesis, for example, KSS, progressive external ophthalmoplegia (PEO), MERRF, and Leber’s hereditary optic neuropathy (LHON), and (ii) those due to mutations in genes encoding individual proteins of the respiratory chain, for example, MELAS, neuropathy, ataxia, retinitis pigmentosa (NARP) or maternally inherited Leigh syndrome (MILS) [69]. Recent studies indicated that there are three important variations including deletion, point mutation, and depletion, which result in mtDNA-related disorders. There are more than 120 different mtDNA deletions that have been identified [70], in which KSS and CPEO are typical examples [71, 72]. KSS is characterized by retinitis pigmentosa, progressive external ophthalmoplegia, cardiomyopathy, deafness, short stature, endocrinopathy, dysphagia, and neurological symptoms. CPEO is characterized by progressive paralysis of the eye muscles. Point mutations are commonly found in protein-coding, rRNA and tRNA genes, and more than half of disease-related point mutations reported are located within mitochondrial tRNA genes, such as m.3243 A→G in the MT-TL1 gene and m.8344 A→G in MT-TK [73, 74]. The syndrome of m.3243 A→G is Mitochondrial Diabetes and Deafness (MIDD) and mitochondrial myopathy, encephalopathy, lactic acidosis and MELAS. The most frequent manifestation of m.8344 A→G in the MT-TK is myoclonic epilepsy with ragged red fibers. Mitochondrial disorders of nuclear DNA (nDNA) origin include OXPHOS disorders, defects in nuclearencoded mitochondrial proteins for mtDNA integrity, and mitochondrial disorders with secondary effects on the OXPHOS system [75]. The OXPHOS system is composed of the four enzyme complexes (complexes IIV) and the ATP synthase complex V. Complex I deficiency accounts for approximately 30% cases of respiratory-chain deficiency in humans. Leigh syndrome (LS) or Leigh-like disease are the most common phenotypes associated with an isolated complex I deficiency. Mutations in nuclear subunits of complex I have been reported in a number of studies [76], such as point mutations of the Nuclear NDUFV1 and NDUFS1 Genes [77]. The subunits of Complex II are entirely encoded by nuclear genes including the

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flavoprotein (SDHA) and iron-sulfur (SDHB). Missense mutations in SDHA gene have been found in two families with autosomal recessive LS and in a family with a late-onset neurodegenerative disease [75, 78, 79]. Mutations in BCS1-L involved in the complex III in early-onset tubulopathy, hepatopathy and encephalopathy [80]. Complex IV (cytochrome oxidase) defects can be caused by nuclear genes of cytochrome oxidase subunits, or cytochrome oxidase assembly and maintenance proteins. The SURF1 gene defects were observed in the large majority of LS-COX (Complex IV) patients [81]. Furthermore, the factors involved in mammalian mtDNA maintenance are all encoded by nuclear genes. Some human diseases were resulted from disturbed mtDNA maintenance mechanisms, such as PEO and mitochondrial DNA depletion syndrome. Mitochondrial disorders often cause problems in many different organ systems, including the nervous, visual, renal (kidneys), digestive and circulatory systems. Despite the fact that mitochondrial diseases can be so variable and affect so many organ systems, a few symptoms are common to many of these disorders. These symptoms include muscle weakness, muscle cramps, extreme fatigue, gastrointestinal problems, droopy eyelids, eye muscle paralysis, retinal degeneration with visual loss, seizures, ataxia and learning delays. The diagnosis of mitochondrial diseases is complex because of its clinical and genetic heterogeneity. The typical diagnostic criteria are heavily dependent on muscle biopsy investigations which include histopathology, electron microscopy, respiratory chain enzymology, and molecular analysis of mitochondrial genes and/or nuclear genes [82]. Various ‘mitochondrial cocktails’ with antioxidant, vitamin or nutrient supplementation are often given to patients with mitochondrial disorders to prevent worsening of symptoms during times of illness and physiologic stress [83]. Some experts also suggested a therapeutic trial of coenzyme Q10 supplementation, along with other antioxidants [84]. The effects of these therapies were limited in mitochondrial disorders [82, 83], but curative treatments for mitochondrial disorders are currently not available. Mitochondria and Cancer Normal cells generate most of their ATP through mitochondrial OXPHOS using glucose, fatty acids, and other metabolic intermediates as energy sources. Besides OXPHOS, cells also produce ATP through glycolysis, which takes place in the cytosol and does not need oxygen. Earlier in the 20th century, Warburg found that cancer cells produce most of their ATP through glycolysis (Warburg effect), even in the presence of oxygen, and he further hypothesized that such a metabolic alteration was attributable to defects in cancer cell OXPHOS [85]. In contrast, recent studies have found that not all areas of all tumors have the phenomenon of the Warburg effect. It has been reported that glioblastoma multiforme, astrocytoma, and certain forms of hepatomas utilize both glycolysis and OXPHOS to an equal extent for energy production,


whereas bone sarcoma, lung carcinoma, breast cancer, skin melanoma, cervical, ovarian and uterine carcinomas, all primarily use OXPHOS for the generation of ATP [86]. To date, the Warburg effect has been observed in many types of tumors, and it has been considered as a basis for clinical diagnosis of tumors [87], but the mechanism for the metabolic shift is still unclear. OXPHOS dysfunction is mostly caused by mutation of mtDNA and nucleus-encoded mitochondrial protein genes. Many studies have identified mutations in mtDNA and nuclear-encoded OXPHOS genes in human cancers, such as carcinomas of the breast, stomach, liver, prostate, kidney, bladder, head and neck, and lung [88, 89]. Polyak et al. detected mutations in seven out of ten cancer cell lines [90]. Maximo et al. have found a significant association between mutations in complex I genes and malignancy [89]. Abu-Amero et al. identified seven somatic mutations in 19 thyroid tumor samples [91]. Shidara et al. [92] have suggested that the pathogenic mtDNA mutations promote tumors by preventing apoptosis. Some tumor-associated mutations have also been found in nuclear genes encoding mitochondrial proteins [93, 94]. The mechanisms by which mtDNA and nuclear DNA mutations can lead to or promote tumorigenesis are not fully understood. It has been suggested that the outcome of such mechanisms would be enhanced glycolysis. Mitochondria and Aging The most famous explanation for aging is the mitochondrial theory of aging, which was first proposed by Harman in 1972. According to this theory, aging is due to the accumulation of damage wrought by free radicals on the mtDNA and function. Mitochondria produce roughly 90% of the energy needs of the cell through the process of OXPHOS in the inner mitochondrial membrane. The respiratory chain of the OXPHOS system is also the primary source of ROS and free radicals, which can cause damage to the membrane, proteins and DNA. Approximately 2% of the molecular oxygen consumed during respiration is converted into ROS. In the past 20 years, many studies have revealed that there are a lot of alterations in mitochondria and mtDNA, including decline in mitochondrial OXPHOS function [95, 96], disorganization of mitochondrial structure [97, 98], increased mitochondrial production of ROS [99, 100] and accumulation of mtDNA mutations [101-103]. These alterations cause mitochondrial dysfunction with lowered ATP production, cellular energy depletion and death, resulting in aging. In addition, dysfunction, disorganization of structure, mtDNA mutation, and increased ROS lead to mitochondrial damage, which then produces ROS and causes mutation. It is suggested that mitochondrial dysfunction promotes further dysfunction, resulting in a vicious cycle. Among all the age-related alterations of mitochondria, two are thought to be the most important. Some studies have


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supported the view that ROS production and mtDNA mutation play the causative role in the aging process and determination of the lifespan of animals [43, 104]. Although mitochondria play an important role in the aging process, there is no good way to increase lifespan by mitochondrial control. Caloric restriction without malnutrition is the only reported technique to modulate energy metabolism in the mitochondria and prevent or attenuate most of these age-associated mitochondrial changes, thereby slowing down the aging process and extending lifespan in various species [105, 106]. However, the underlying mechanism how the caloric restriction slows down the aging process awaits further study.

CONCLUDING REMARKS Mitochondria are involved in many aspects of eukaryotic biology including energy production, signal transduction and apoptosis control. Abnormal functions of mitochondria are linked to various human diseases such as cancer and aging. After more than 100 years’ study on this cellular organelle, we have comprehensive understanding on its structure and functions. Nevertheless, there are still questions left unanswered, especially its functional mechanisms in signaling and cancer. For example, how mtDNA and nuclear DNA mutations lead to or promote tumorigenesis is not fully understood. Although many diseases are derived from or perhaps are caused by mtDNA mutations, the pathogenesis of these diseases remains poorly understood and there are no good therapeutic methods to treat them. More biochemical and physiological studies of mitochondrial functions will bring novel information on these aspects in the future.

CONFLICT OF INTEREST The authors confirm that this article content has no conflicts of interest.

ACKNOWLEDGEMENTS This work was supported by the National Special Fund for Scientific Research in Public Benefits (Grant No. 200903046), the National Natural Science Foundation of China (Grant No. 30930071 and Grant No. 31001105), Major international cooperation projects of the National Natural Science Foundation of China (Grant No. 31210103918), the Doctoral Fund of Ministry of Education of China (Grant No. 20114306130001), the National High Technology Research and Development Program of China (Grant No. 2011AA100403), and Cooperative innovation center of engineering and new products for developmental biology.

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Received: August 08, 2013

Revised: October 18, 2013

Accepted: October 29, 2013

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