**3. The possible roles of metabolic states that can alter transcription profiles**

which encode mitochondria-localizing poly(ADP-ribose) (PAR) degrading enzyme [42] and an anti-viral RNA-binding protein PARP13 [43, 44], respectively. These findings suggest that the expression of genes encoding single-strand DNA break repair factors is commonly regu-

The promoter activities of the human *WRN* and *TERT* genes, both encoding telomere maintenance factors, positively respond to both 2-deoxy-d-glucose (2DG) [45] and *trans*-resveratrol (Rsv) [46], which are caloric restriction (CR) mimetic drugs that have been shown to prolong the life span of several organisms [47]. The natural compound Rsv upregulates the expression of the *HELB* gene [46, 48], which encodes DNA replication and DNA double strand break repair-helicase HELB (HDHB) [49–52]. Moreover, the 5′-regulatory regions of the genes that encode DNA repair factors, such as *XPB*, *RB1*, *RTEL1*, *ATR*, *TP53*, and *CDKN1A (p21)*, contain GGAA duplications near the TSS [53]. Several of the DNA repair factors are localized in the

**2.4. The surveillance of regulatory regions adjacent to the TSSs of human mitochondrial** 

The surveillance of a human genomic DNA database suggested putative TPA-responsive elements in the 5′-upstream regions of the *MRPL32*, *NDUFB3*, *NDUFS3*, *SDHB*, and *SDHAF2* genes contain GGAA duplication [54]. The duplicated GGAA-motifs are present in the upstream regions of human genes encoding mitochondrial ribosomal proteins and enzymes or components that function in the TCA cycle and oxidative phosphorylation (OXPHOS) [54]. Mitochondrial dysfunction is thought to cause either cellular senescence or oncogenesis [55–58]. Remarkably, TCA cycle enzymes, fumarate hydratase (FH), and succinate dehydrogenase (SDH) have been suggested as tumor suppressors [59]. Hence, mutations of the TCA cycle factor-encoding genes give rise to abnormal mitochondrial respiration, which is one of the characteristics of tumors and cancer [60, 61]. Mutations of the *IDH1* and *IDH2* genes have been identified in human brain cancer cells [62, 63]. A recent study demonstrated that the mutation of IDH2 could lead to the generation of sarcoma [64]. Duplicated GGAA-motifs are contained in the upstream region of the *NAMPT* (*NmPRT*), encoding a nicotinamide phosphoribosyltransferase that catalyzes the first rate-limiting step of (nicotinamide adenine dinu-

synthesis from nicotinamide [65–67]. Depending on the NAD+

could modulate the TCA cycle, poly(ADP-ribosyl)ation, and sirtuin-mediated de-acetylation [66, 67]. The duplicated GGAA-motifs are present near the TSSs of the human TCA cycle enzyme-encoding *ACLY*, *ACO2*, *CS*, *FH*, *IDH1*, *IDH3A*, *IDH3B*, *SDHAF2*, *SDHB*, *SDHD*, and

A duplicated GGAA-motif is present in the bidirectional promoter of the *PDHX* [54], which encodes one of the components of the PDH enzyme to metabolize pyruvate to acetyl-CoA. Aberrant pyruvate metabolism is thought to play a prominent role in cancer [69]. The

boxylase that converts malate into pyruvate), is found in pancreatic ductal adenocarcinoma [70]. The GGAA-duplication is not only found near the TSSs of the human *ME2* gene, but

genomic deletion of *ME2*, which encodes malic enzyme 2 (an NAD+

level, NAMPT


mitochondria and may also regulate the mitochondrial functions [53].

lated by the duplicated GGAA-motifs.

110 Mitochondrial Diseases

**function-associated genes**

cleotides) NAD+

*SUCLG1* genes [68].

Recently, a study using a CAP-SELEX analysis showed that different transcription factors, such as FOXO1 and ETS family proteins, are mediated by a DNA that contains a GGAAcore motif [71]. As described above, a number of promoters or regulatory regions of human genes that encode immune response-/DNA repair-/mitochondrial function-associated proteins contain overlapping or duplicated GGAA-containing motifs. Thus, the alteration of the profile of the GGAA-motif-binding proteins or their associated protein factors may allow for the control of appropriate cellular responses against viral infection, DNA damage, and oxidative/nutrient/metabolic stress. Importantly, the DNA damage responses affect the transcriptional state [72] through oxidative stress, which is mainly produced by the mitochondria [73, 74]. NF-κB- and p53-dependent transcription, which regulates the expression of the ISGs and DNA repair factor-encoding genes, is also affected by oxidative stress [75]. Thus, metabolites that are mainly produced by respiration or mitochondrial functions may influence the transcription control system [76, 77].

#### **3.1. The transcription profile may be controlled by the NAD<sup>+</sup> /NADH balance**

We have reported that the promoter regions of the human *TP53*, *HELB*, and telomere maintenance factor-encoding genes respond positively to Rsv [46, 48, 78]. Rsv not only activates SIRT1, which is an NAD+ -dependent deacetylase [79], but also inhibits phosphodiesterase [80]. Importantly, low-dose Rsv activates mitochondrial complex I [81] to upregulate the NAD+ /NADH ratio, to induce the expression of duplicated GGAA-motif-driven genes. The transcription of the bidirectional promoter-driven *BRCA1/NBR2* genes, which contain a duplication of the GGAA-motif, may be regulated by the NAD+ /NADH ratio [82]. Notably, the C terminal-binding protein (CtBP) [83, 84] has a central role in this regulation as a metabolic sensor. Moreover, PARP1 poly(ADP-ribosyl)ates transcription elongation factor NELF to release the paused RNA pol II-dependent transcription [85], suggesting that PARP1 itself contributes to NAD+ -sensitive transcription. Recently, it was reported that nuclear PAR can be utilized by NUDIX5 to supply ATP molecules, which are required for chromatin remodeling [86]. Thus, the accumulation of NAD+ molecules or NAD+ /NADH ratio-sensitive proteins, including GGAA-motif binding TFs, might affect the transcription of ISGs/DNA repair/mitochondrial function-associated genes in response to metabolic stress.

It should be noted that PARP activity is upregulated in tumors and cancer cells [44]. Because the duplicated GGAA-motifs are present in the 5′-upstream regions of the human *PARP* and *PARG* genes [40], subtle changes in the quality/quantity profile of the GGAA-binding TFs may modulate the PAR synthesis/degradation ratio at the transcription level. The accumulation of NAD+ molecules in cells might be transiently caused by mitochondrial dysfunction, which is usually accompanied by insufficient OXPHOS or aberrant respiration [87, 88]. However, when DNA damage eventually activates PARP, NAD+ molecules will be consumed to synthesize PAR polymer in the nuclei or mitochondria. Thus, the decrease in the NAD+ /NADH ratio would not only reduce the activities but also the expression of enzymes that function in the NAD+ -dependent TCA cycle progression. At this point, cells will have to produce ATP in a mitochondria-independent manner. This metabolic change would be observed as the "Warburg effect" in cancer cells [5, 6].

**4.1. The possible functions of poly(ADP-ribosyl)ation on epigenetic regulation**

gene expression regulatory system [101]. More importantly, NAD+

[115]. Taken together, these observations imply that NAD+

**4.2. The DNA methylation of chromosomal DNAs**

 not only plays important roles in DNA repair, mitochondrial functions, and cellular senescence [72, 100], but also affects the modification of chromatin proteins [77] and modulates the

A New Insight into the Development of Novel Anti-Cancer Drugs that Improve the Expression…

, which might accompany the decrease in PARP activity [114]. SIRT1, which depends on

involved in epigenetic regulation, and that it may be altered in line with the aging process.

The methylation of promoter regions of specific genes in human chromosomes can be used as biomarkers in various cancers [116]. The DNA methylation reaction is catalyzed by methyltransferases (DNMTs), which utilize AdoMet as a methyl group donor [117]. A recent study showed that intragenic DNA methylation, which is carried out by Dnmt3b in mouse embryonic stem cells, protects the gene body from the entry of spurious RNA pol II and the initiation of cryptic transcription [118]. The extended data showed that the ETS factor binding regions are sensitive to the knock out of the *Dnmt3b* gene, suggesting that the occupation of the GGAA (TTCC)-motifs by GGAA-motif binding proteins could be epigenetically controlled by methylation. Furthermore, the regulation of demethylation by ten-eleven translocation (TET)-family enzymes [119], the activity of which is reduced by hypoxia, should not be ignored. Hypoxia-induced hyper methylation has been demonstrated to occur on the promoter regions of the DNA repair factor-encoding genes, including *BRCA1*, *FANCD2*, *FANCF*, *POLL*, and *UNG* [120]. Of note, the duplicated GGAA-motifs are contained in these gene promoters [53]. A methylation sensitive SELEX analysis showed that ETS-binding was inhibited by mCpG, though NFAT, which also recognizes the GGAA-core motif sequence and preferentially binds to methylated DNA [121]. The observation suggests that GGAA-motif recognizing TFs could be classified into two groups according to their preference to DNA methylation. The SET protein is an epigenetic regulatory factor that promotes loss of methylation through direct interaction with hypo-acetylated histones [122]. A genome-wide analysis showed that DNA hypermethylation is apparently induced in old male adults, relative to young male

molecule to de-acetylate histone proteins, plays important roles in the aging process

enzyme to synthesize PAR macromolecules, which modify both PARP itself and chromosomal proteins and DNA repair factors [67]. Histones and HMGB (High mobility group box) proteins can be poly(ADP-ribosyl)ated [102–105], suggesting that modifications by such macromolecules on chromosomes affect the epigenetic regulation of the gene expression system. Moreover, poly(ADP-ribosyl)ation on the chromosomal insulator protein CTCF (CCCTC-binding factor) may be involved in epigenetic regulation [106, 107]. Recently, it was shown that CTCF binds directly to PAR to be recruited at DNA lesion sites, indicating that the CTCF also plays a role in the DNA damage response [108]. It has been suggested that poly(ADP-ribosyl)ation affects the methylation patterns in chromosomal DNAs [109, 110]. A recent study showed that the transcriptional regulation of the *EZH2* gene, which encodes the catalytic subunit of the polycomb repressive complex 2 (PRC2), by PARP1 [111], affects the methylation of chromatin proteins [112]. Because the incidence of cancer increases with aging [113], a decline in the cellular level of

is a substrate for the PARP

113

http://dx.doi.org/10.5772/intechopen.71095

and its polymerized form, PAR, are

NAD+

NAD+

the NAD+
