**3. Mitochondrial-derived peptides**

Vertebrate mitogenomes have the potential of encoding several short peptides (mitochondrial-derived peptides (MPDs)) [26–28]. The best characterized peptides among the MDPs are MOTS-c and humanin (HN). Genes coding for MOTS-c and HN are found as small open reading frames within the mitochondrial small subunit (mtSSU) and large subunit (mtLSU) ribosomal DNA, respectively [29, 30]. Studies in mammals indicate that MDPs are circulating signaling molecules with a number of proposed roles. While HN is involved in cellular stress resistance, apoptosis, and metabolism [29, 31–34], MOTS-c apparently represents an MDP hormone that regulates metabolic homeostasis and insulin sensitivity [30, 35].

The Atlantic cod open reading frames encoding MOTS-c and HN were identified at the exact same locations as in human, within the domain 3'M and domain IV of the mtLSU rRNA and mtSSU rRNA, respectively (**Figure 2A** and **B**). Comparative analysis revealed MOTS-c and HN to be invariant among Atlantic cod specimens [18] and well conserved between Atlantic cod and human (**Figure 2C**). Here, 8 of 16 amino acid residues in MOTS-c and 13 of 21 amino acid residues of HN were shared. Furthermore, when comparing gadiform species representing seven diverse families, we noted 10 of 16 and 15 of 21 amino acid residues to be shared in MOTS-c and HN, respectively (**Figure 2C**). The conserved features seen between gadiform species and human suggest related MDP functions.

**Figure 2.** Putative mitochondrial-derived peptides in Atlantic cod. (A) Secondary structure diagram of the Atlantic cod mtSSU rRNA domain 3'M coding for the putative MOTS-c peptide (red letters). (B) Secondary structure diagram of the Atlantic cod mtLSU rRNA domain IV coding for the putative HN peptide (red letters). (C) Alignment of MDP (MOTS-c and HN) sequences from seven gadiform species representing different families (gm, *Gadus morhua*, Gadidae, HG514359; Ll, *Lota lota*, Lotidae, AP004412; mm, *Merluccius merluccius*, Merlucciidae, FR751402; Ec, *Enchelyopus cimbrius*, Phycidae, AJ315624 and FJ215015; tm, *Trachyrincus murragi*, Macrouridae, AP008990; Bn, *Bregmaceros nectabanus*, Bregmacerotidae, AP004409; Pj, *Physiculus japonicus*, Moridae, AP004409) and human (Hs, *Homo sapiens*, NC\_012920). Stars above and below the alignment represent conserved residues among gadiforms and between gadiforms and human, respectively.

Expanding the Coding Potential of Vertebrate Mitochondrial Genomes: Lesson Learned…

http://dx.doi.org/10.5772/intechopen.75883

99

Expanding the Coding Potential of Vertebrate Mitochondrial Genomes: Lesson Learned… http://dx.doi.org/10.5772/intechopen.75883 99

were specific to NA and NC cod, respectively. From the dataset we determined the mitochondrial substitution rate to be 14 times higher compared to that of the nuclear genome [16, 17]. More recently we analyzed 156 Atlantic cod mitogenomes at the individual level [18], including 32 specimens previously reported by Carr and Marshall [19]. We found 1034 SNPs in total among the sequences, which were not evenly distributed throughout the mitogenome. The ND2 gene (Complex I) and the COII gene (Complex IV) were the least and most conserved, respectively, among the protein-coding genes. Furthermore, rRNA and tRNA genes showed a significantly lower density of overall SNPs per site compared to protein-coding genes. Thus, the Atlantic cod mitogenome follows a similar pattern of conservation as seen for other vertebrates like zebrafish and human [20–23] and corroborates the observation that mutation rate constrains in vertebrate mitogenomes appear linked to the position of genes in relation to OriH and OriH [24, 25].

The noncoding regions of the Atlantic cod mitogenome showed a mosaic pattern of sequence conservation. Whereas the OriL and the central domain of CR were almost invariant among specimens, the T–P spacer and 5′ domain of CR contain significant sequence variation [7, 10, 13, 18]. The 74-bp T–P spacer was found to contain 16 variable sites and 26 haplotypes among 225 specimens assessed, including a 29-bp sequence duplication in three individuals [10]. Similarly, the 5′ domain of CR was the most variable region within the mitochondrial genome (more than three times that of average substitution rate). The elevated sequence variation was due to hot-spot substitution sites, homopolymeric heterogeneity, and the HTR array [18].

Vertebrate mitogenomes have the potential of encoding several short peptides (mitochondrial-derived peptides (MPDs)) [26–28]. The best characterized peptides among the MDPs are MOTS-c and humanin (HN). Genes coding for MOTS-c and HN are found as small open reading frames within the mitochondrial small subunit (mtSSU) and large subunit (mtLSU) ribosomal DNA, respectively [29, 30]. Studies in mammals indicate that MDPs are circulating signaling molecules with a number of proposed roles. While HN is involved in cellular stress resistance, apoptosis, and metabolism [29, 31–34], MOTS-c apparently represents an MDP

The Atlantic cod open reading frames encoding MOTS-c and HN were identified at the exact same locations as in human, within the domain 3'M and domain IV of the mtLSU rRNA and mtSSU rRNA, respectively (**Figure 2A** and **B**). Comparative analysis revealed MOTS-c and HN to be invariant among Atlantic cod specimens [18] and well conserved between Atlantic cod and human (**Figure 2C**). Here, 8 of 16 amino acid residues in MOTS-c and 13 of 21 amino acid residues of HN were shared. Furthermore, when comparing gadiform species representing seven diverse families, we noted 10 of 16 and 15 of 21 amino acid residues to be shared in MOTS-c and HN, respectively (**Figure 2C**). The conserved features seen between gadiform

hormone that regulates metabolic homeostasis and insulin sensitivity [30, 35].

**3. Mitochondrial-derived peptides**

98 Mitochondrial DNA - New Insights

species and human suggest related MDP functions.

**Figure 2.** Putative mitochondrial-derived peptides in Atlantic cod. (A) Secondary structure diagram of the Atlantic cod mtSSU rRNA domain 3'M coding for the putative MOTS-c peptide (red letters). (B) Secondary structure diagram of the Atlantic cod mtLSU rRNA domain IV coding for the putative HN peptide (red letters). (C) Alignment of MDP (MOTS-c and HN) sequences from seven gadiform species representing different families (gm, *Gadus morhua*, Gadidae, HG514359; Ll, *Lota lota*, Lotidae, AP004412; mm, *Merluccius merluccius*, Merlucciidae, FR751402; Ec, *Enchelyopus cimbrius*, Phycidae, AJ315624 and FJ215015; tm, *Trachyrincus murragi*, Macrouridae, AP008990; Bn, *Bregmaceros nectabanus*, Bregmacerotidae, AP004409; Pj, *Physiculus japonicus*, Moridae, AP004409) and human (Hs, *Homo sapiens*, NC\_012920). Stars above and below the alignment represent conserved residues among gadiforms and between gadiforms and human, respectively.
