BioMol Concepts, Vol. 3 (2012), pp. 107–115 • Copyright © by Walter de Gruyter • Berlin • Boston. DOI 10.1515/bmc-2011-0058 Review Mitochondrial DNA: a blind spot in neuroepigenetics Hari Manev *, Svetlana Dzitoyeva and Hu Chen (7) . These studies of chromatin remodeling comprise protein (e.g., histone) modifi cations and DNA modifi cations, such The Psychiatric Institute , Department of Psychiatry, as the formation of 5-methylcytosine (5mC) and 5-hydroxy- University of Illinois at Chicago, 1601 West Taylor Street, methylcytosine (5hmC). Whereas 5mC DNA modifi cations in M/C912, Chicago, IL 60612 , USA the dinucleotide sequence CpG are generally considered tran- * Corresponding author scriptional repressors [recent data suggest a more complex role e-mail: [email protected] for 5mC DNA, including stimulation of gene expression (8)], the functional role of 5hmC DNA modifi cations is currently Abstract unclear. Since the early days of developmental epigenetics, new ideas have emerged that imply epigenetic mechanisms Neuroepigenetics, which includes nuclear DNA modifi ca- are a bridge between the environment and lasting, sometimes tions, such as 5-methylcytosine and 5-hydroxymethylcyto- heritable genome modifi cations. Hence, both the adaptive and sine and modifi cations of nuclear proteins, such as histones, harmful biological consequences of an individual ’ s bi-direc- is emerging as the leading fi eld in molecular neuroscience. tional interaction with the environment are possibly best con- Historically, a functional role for epigenetic mechanisms, tained in that individual ’ s modifi ed genome – the epigenome. including in neuroepigenetics, has been sought in the area of Compared to a developmental role of epigenetics, consid- the regulation of nuclear transcription. However, one impor- erations of epigenetic mechanisms as a modifi able functional tant compartment of mammalian cell DNA, different from system in postmitotic cells, such as neurons (i.e., neuroepige- nuclear DNA but equally important for physiological and netics) are more recent. In a series of articles published between pathological processes (including in the brain), mitochondrial 1974 and 1977, Boris Vanyushin and colleagues (9) provided DNA has for the most part not had a systematic epigenetic the fi rst direct evidence in support of the 1969 hypothesis (10) characterization. The importance of mitochondria and mito- that DNA methylation of neuronal ncDNA functions as the epi- chondrial DNA (particularly its mutations) in central nervous genetic component of learning and memory. In these experi- system physiology and pathology has long been recognized. ments, rats were trained (conditioned) to associate a light cue Only recently have the mechanisms of mitochondrial DNA with food (a food reinforcement model). Brain samples (the methylation and hydroxymethylation, including the discovery cortex, hippocampus, and cerebellum) were collected from con- of mitochondrial DNA-methyltransferases and the presence trol and conditioned rats. For each brain region, neuronal nuclei and functionality of 5-methylcytosine and 5-hydroxymeth- were separated from the glial nuclei. ncDNA methylation was ylcytosine in mitochondrial DNA (e.g., in modifying the affected by the conditioning model in the cortex and the hip- transcription of mitochondrial genome), been unequivocally pocampus but not in the cerebellum, and only in neuronal but recognized as a part of mammalian mitochondrial physiology. not in glial ncDNA. In both affected brain regions, the learning Here, we summarize for the fi rst time evidence supporting the model caused an increase of 5mC in neuronal ncDNA. This existence of these mechanisms and propose the term ‘ mito- pioneering work received little attention and was soon forgot- chondrial epigenetics ’ to be used when referring to them. ten. The same idea was resurrected 25 years later in a letter to Currently, neuroepigenetics does not include mitochondrial the editor of the Journal of Theoretical Biology (11) . Although epigenetics – a gap that we expect to close in the near future. no data were presented in support of the proposed hypothesis, which asked the question whether there was an epigenetic com- Keywords: amyotrophic lateral sclerosis (ALS); ponent in long-term memory, subsequent research has provided DNA methylation; DNMT1; DNMT3A; epigenetics; a clear positive response to this question (12) . Moreover, recent 5-hydroxy methylcytosine; mitochondria. advancements in the methodologies for the characterization of DNA methylation demonstrated the ease with which neuronal Introduction activity in the adult brain (e.g., in the hippocampus) is capable of modifying its DNA methylation landscape (13) . Moreover, The earliest understanding of the functional role of epigenetic the same mechanisms appear to be involved not only in brain mechanisms relates to developmental genome regulation, e.g., physiology but also in the pathobiological mechanism of neu- silencing of gene expression involved in cell differentiation ropsychiatric disorders, e.g., schizophrenia (14 – 16) . and in maintaining cell phenotypes during cell proliferation However, one important compartment of mammalian cell (1 – 6) . Cellular chromatin, a structure composed of nuclear DNA, different from ncDNA but equally important for physi- DNA (ncDNA) and nuclear proteins (including histones), has ological and pathological processes (including in the brain), been the primary target of epigenetic research, which brought mitochondrial DNA (mtDNA) has for the most part escaped about major new concepts on the functionality of the genome a systematic epigenetic characterization. The organization of 108 H. Manev et al. mtDNA is different from the structural organization of ncDNA In postmitotic cells, such as neurons, mtDNA is prone to (17) ; most notably, mitochondria do not contain histones. mutations and brain cells are known to have a higher degree Hence, epigenetic mechanisms, such as histone modifi ca- of heteroplasmy (the presence of more than one type of tions, which are important for ncDNA, may not be applicable mtDNA within a cell) than cells in rapidly dividing tissues. to mtDNA, stressing the importance of the mechanisms of Generally, mtDNA mutations have been proposed as the 5mC and 5hmC formation for mtDNA. source of regional ancient variants that evolutionarily permit- ted humans to adapt to differences in their energetic environ- ments and as a cause of disease. The latter includes deleterious Mitochondrial DNA germline line mutations causing mitochondrial diseases, and somatic mutations that accumulate with age and may cause mtDNA was discovered and fi rst visualized through electron aging-associated disorders (31) . Both mtDNA mutations and microscopy by Margit Nass-Edelson and Sylvan Nass in 1963 deregulation of mtDNA gene expression have been recog- (18, 19) . On the basis of the observed ultrastructural similar- nized as the basis for a number of human disorders (24) . ity of this intracellular organelle and the bacterial cells, the idea of endosymbiosis developed, which suggests that dur- ing evolution, bacterial cells became engulfed and modi- mtDNA cytosine methylation fi ed to become eukaryotic organelles, such as mitochondria (20) . Since the time of these original observations, enormous Although as long ago as the late 1940s (32, 33) , numerous progress has been made in understanding the structure and studies had confi rmed the presence of a substantial amount function of mtDNA and about its role in human pathology, of 5mC in vertebrate ncDNA [including in the brain (34) ], including neuropsychiatric disorders. Extensive reviews have no signifi cant functional role was assigned to this DNA summarized these achievements [e.g., (21 – 24) ]. modifi cation for quite some time. A uniquely powerful argu- Briefl y, mammalian mitochondria contain multiple copies ment for dismissing the important functionality of vertebrate of a maternally inherited genome, a 16,295 – 16,826 bp (25) DNA methylation, which is particularly abundant during circular double-stranded mtDNA that encodes 13 polypep- development, was raised in 1985 (6) . It was pointed out that tides, 2 ribosomal RNAs, and 22 transfer RNAs. The proteins Drosophila (fruit fl y) had no proven DNA methylation, but encoded in mtDNA are all members of the oxidative phospho- nevertheless, like vertebrates, is capable of accomplishing rylation (OXPHOS) complexes. Initially, it was believed that sophisticated developmental pathways. The argument was mtDNA is naked and thus vulnerable to damage. However, then made that if Drosophila could accomplish its compli- recent studies have established that mtDNA is protein coated cated differentiation without DNA methylation, how could and packaged into aggregates called nucleoids or mitochro- vertebrate development use DNA methylation as an impor- mosome. Mitochondrial transcription factor A (TFAM) is tant gene regulator ? In spite of this conceptual obstacle, the the most abundant component of the nucleoid and, similar to current understanding of the functional role of DNA methyla- the action of histones on ncDNA, it plays a major role in the tion in the regulation of gene expression has evolved dramati- packaging and compaction of mtDNA (26 – 28) . Current data cally. In vertebrate ncDNA, a methyl group is added to the 5 indicate that, contrary to the previous belief, most nucleoids position of the base cytosine to generate 5mC by the action contain a single copy of mtDNA, supporting the proposal that of three