**2. Mitochondrial transcripts and noncoding RNAs**

cell processes. However, many other functions have been assigned to this tiny organelle, such as apoptosis, reactive oxidative species (ROS) signaling, inflammation and metastasis. Mitochondria play a central role in apoptosis, principally due to the release of proteins from the mitochondrial inter-membrane space [1], linking mitochondria to cell suicide. Mitochondria also represent a major source of DNA-damaging reactive oxygen species (ROS), mainly as byproducts of oxidative phosphorylation. In comparison to nuclear DNA, mitochondrial DNA (mtDNA) is more susceptible to DNA damage due to the reduced capacity of the cell to repair

In inflammation, mitochondria mainly serve as a source of various signaling molecules, termed damage-associated molecular patterns or DAMPS, which propagate inflammatory signals and

Cancer death is most often due to secondary tumors or metastasis. This process requires a complex series of events, which include epithelial-to-mesenchymal transition, stromal remodeling, invasion and ultimately migration of cancer cells. In this context, mitochondrial ROS play a key role, leading to angiogenesis and metastasis [4] and promoting the migratory plasticity of cells through activation of two essential factors, Src and protein tyrosine kinase 2 [5]. The basic understanding of the dependence of cancer cells on various mitochondrial roles is

Along that line, an expanding group of evidences indicate that the mitochondrial genome is not only responsible for the synthesis of the canonical 13 proteins, 22 tRNAs and two ribosomal RNAs (12S and 16S). Novel and expanding evidence suggest that mtDNA compensates for reduced length by using little known phenomena that potentially increase DNA's protein coding repertoire, such as so called swinger polymerization, that consists of systematic exchanges between nucleotides during DNA or RNA polymerization, producing so-called swinger sequences [6, 7]. These transformations alter gene and mRNA coding properties. Moreover, in human mitochondria systematic deletions of mono- and dinucleotides after each trinucleotide have been reported, producing delRNAs. Recently, an exhaustive analysis of human nanoLc mass spectrometry peptidome data detect numerous tetra- and pentapeptides translated from the human mitogenome, and this peptide subgroup would be the result of the translation of delRNAs. Therefore, non-canonical transcriptions and translations could considerably expand the coding potential of mitochondrial DNA and RNA sequences [8, 9].

Finally, in the field of antisense RNAs and because mitochondrial tRNA mutations are 6.5 times more frequently pathogenic than in other mitochondrial sequences, a potential additional tRNA gene function is that of templating for antisense tRNAs. Most antisense tRNAs probably function routinely in translation and extend the tRNA pool and mutation pathogenicity, probably frequently resulting from a mixture of effects due to sense and antisense tRNA translational activity for many mitochondrial tRNAs [10, 11]. These could link to mitochondrial disorders and cancer.

This small but powerful organelle is also a novel source of noncoding RNAs (ncRNAs), and growing evidence shows that mammalian mitochondria can also import/export ncRNAs, turning this organelle into a pivotal player not only in cellular physiology but also in cancer,

representing potential targets for innovative ncRNA-based treatment strategies [12].

mtDNA, potentially promoting cancer [2].

already endorsing novel therapeutic approaches in cancer.

therefore activate inflammation [3].

180 Mitochondrial DNA - New Insights

Human mitochondrial DNA (mtDNA) is a circular molecule of 16,569 bp in length [13] which encodes a small subset of 13 proteins required for OxPhos, 22 tRNAs and two ribosomal RNAs, 12S rRNA and 16S rRNA, which form part of the small (28S) and large (39S) subunits of the 55S mitoribosome [14]. All other protein components are encoded by nuclear genes and imported into mitochondria from the cytosol.

Replication and transcription of mtDNA is initiated from the D loop, a small noncoding region, and is regulated by nuclear-encoded proteins imported into mitochondria [15]. Mitochondrial RNAs are transcribed as long polycistronic precursors from both strands, termed heavy (H) and light (L) strands [16]. Except for NADH dehydrogenase 6 (ND6), all the 13-mitochondrial proteins are encoded in the H-strand. Additionally, the H-strand encodes 14 of the 22 tRNAs and the 2 rRNAs. The remaining 8 tRNAs are encoded on the L-strand [17]. The precursor transcripts are processed according to the tRNA punctuation model, whereby 22 interspersed tRNAs are excised at their 5' and 3'ends by RNase P [18] and by RNase Z, elaC homology 2 (ELAC2), respectively, releasing simultaneously individual rRNAs and mRNAs [19]. The RNAs then undergo maturation, involving polyadenilation at the 3'eextremitoess of mRNAs and rRNAs, and specific nucleotide modifications and addition of CCA trinucleotides to the 3'eextremities of tRNAs [20]. The data of several groups indicate that 250–300 nuclear-encoded proteins are dedicated exclusively to serve mitochondrial gene expression. This includes RNA polymerase, endonucleases for RNA processing, translation factors, biogenesis factors for the mitochondrial ribosome, aminoacyl-tRNA synthetases, and other auxiliary factors [21, 22].

However, evidence has accumulated supporting the notion that, besides proteins, many types of RNAs transcribed from the nuclear genome are actively delivered to mitochondria. Among these transcripts are different types of noncoding RNAs, such as tRNAs, 5S rRNA, MRP RNA (RMRP) and RNase P RNA (RPPH1) [23], as well as microRNAs (mitomiRs) [24]. The logical explanation is that, despite their critical function, the handfuls of mitochondrial- and nuclearencoded proteins are insufficient to maintain mitochondrial structure or activity.

Noncoding RNAs (ncRNAs) are divided in two major groups according to size, as small noncoding RNAs and long noncoding RNAs. Among the small ncRNAs, microRNAs (miRNAs) are the most-studied class in mammals. These RNAs (20–24 nucleotides in length) negatively regulate gene expression through binding with their target mRNA and have been implicated actively in pathogenic processes of many human diseases [24] and, as such, are important regulators of cancer cell metabolism [25]. The observation of association of miRNAs with/inside mitochondria may have important implications in several cellular processes and suggest that the role of mitochondria clearly extends beyond its role in energy metabolism and other cellular processes. The newfound destination of miRNAs indicates novel roles of mitochondria in normal and pathological events [25].

The class of long noncoding RNAs (lncRNAs) has been recently recognized, and is defined as transcripts longer than 200 nucleotides. The size cutoff is arbitrary and many functional lncRNAs are considerably longer than 200 nucleotides, including X-inactive specific transcript (XIST) [26], its antisense form Tsix [27], and Hox transcript antisense intergenic RNA (HOTAIR) [28], which are several kilobases (kb) in length.

sperm. To determine when during spermatogenesis the mitochondrial RNA is localized in the nucleus, ISH of mouse and human testis was carried out. The nuclei of spermatogonia, spermatocytes and round and elongated spermatids were all positively stained. In human spermatocytes, the nuclear staining pattern was fibrillar, suggesting an association of the

Long Noncoding Mitochondrial RNAs (LncmtRNAs) as Targets for Cancer Therapy

http://dx.doi.org/10.5772/intechopen.75453

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These results suggested that the nuclear localization of the 16S mitochondrial rRNA in spermatogenic cells is the result of an intriguing process of translocation of the transcript from the organelle to the nucleus. This hypothesis leads to several important questions; for example, how does the organelle regulate the exit of the 16S mitochondrial rRNA without affecting the number of copies needed to assemble mitoribosomes for normal mitochondrial translation? Or what is the mechanism by which this RNA is exported from mitochondria? At present

However, the extramitochondrial localization of the 16S mitochondrial rRNA described by us is not unique, since the same transcript has been found consistently in the cytoplasm of Drosophila and Xenopus embryos [37, 38]. Moreover, injection of an anti-16S rRNA ribozyme into cleavage embryos of Drosophila demonstrated that the rRNA is actively involved in the generation of

As mentioned above, ISH revealed that this lncRNA is over-expressed in human sperm and precursor cells [36]. These results suggest that human cells might contain a transcript with structural features similar to the mouse RNA. We found that the human RNA is over-expressed in several human proliferating cells but not in resting cells. The structure of this transcript of 2374 nt, which we designated sense noncoding mitochondrial RNA or SncmtRNA, revealed the presence of an inverted repeat (IR) of 815 nt linked to the 5′ end of the 16S mtrRNA. The expression of this transcript can be induced in resting lymphocytes stimulated with phytohaemagglutinin (PHA), together with DNA synthesis and the expression of the proliferation markers proliferating cell nuclear antigen (PCNA), Ki-67 and phosphohistone H3. On the other hand, treatment of DU145 cells with aphidicolin reversibly blocks cell proliferation as well as the expression of the ncmtRNA. These results suggested that the ncmtRNA is a new marker of cell proliferation [40].

Afterwards, we described 2 mitochondrial transcripts (ASncmtRNAs) in human cells containing stem-loop structures similar to that of the previously described SncmtRNA.Regarding expression of the SncmtRNA and the ASncmtRNAs, 3 different phenotypes of human cells can be defined. Normal proliferating cells express both families of transcripts; in striking contrast, tumor cells express the SncmtRNA and down-regulate the ASncmtRNAs. Finally, neither of these transcripts is expressed in nondividing cells. Down-regulation of the ASncmtRNAs was observed in 15 different tumor cell lines and in tumor cells present in 273 cancer biopsies corresponding to 17 different cancer types [41]. SncmtRNA is expressed in all proliferating cells, independently whether we are dealing with a regulated or a dysfunctional cell cycle. The fact that the ASncmtRNAs are always down-regulated in tumor cells suggests that, hypothetically, the ASncmtRNAs might function as a unique mitochondria-encoded tumor suppressor. **Figure 1** shows a panel of ISH for S and ASncmtRNA in non-proliferating tissue (liver), tumor tissue (cervix carcinoma) and normal proliferating tissue (normal cervix epithelium), representing the concept of differential expression mentioned. For in situ hybridization, tissue sections were incubated with hybridization mixture

mitochondrial transcript with the meiotic chromosomes [36].

these questions remain unanswered.

pole cells, progenitors of the germ line [39].

LncRNAs are able to interact with DNA, RNA, and proteins. In doing so, they regulate several processes including chromatin dynamics, gene transcription, splicing, and translation [29]. Their involvement in these processes implicates lncRNAs in various aspects of human physiology and disease, which include cancer. Aberrant lncRNA expression has been associated to various cancer types [30]. Moreover, deregulated lncRNA expression patterns can modulate several hallmarks of cancer [31], including sustained growth signaling, repressed growth inhibition, apoptosis evasion, stimulated proliferation, and the promotion of angiogenesis [32].
