Cytochrome Oxidase Subunit III Research Articles

Using genomics and morphometrics to monitor data-poor and commercially exploited octopod populations

Over 150 species of benthic octopods have been described within the ‘catch-all’ Octopus genus (Family: Octopodidae) and yet, many Octopus species harvested by fisheries remain unidentified to species-level due to a lack of distinguishing traits. Within species, there is also limited information on how populations differ genetically and the level of connectivity between populations. Therefore, we sampled octopods from commercial fisheries in southeast Australia, in order to identify the species, examine the phylogeographic relationships among species and the level of population genetic structuring within species, as well as to look for any adaptive genetic variation. The mitochondrial gene, cytochrome oxidase subunit III (COIII), was sequenced in 346 octopods along with single nucleotide polymorphisms using double digest restriction site-associated DNA sequencing (ddRADseq). Morphometric traits were also measured in mature specimens. The southern keeled octopus (‘Octopus’berrima) and pale octopus (‘Octopus’pallidus) were identified using COIII data. For ‘Octopus’berrima, we found that some populations whilst being morphologically similar were genetically distinct. In contrast, ‘Octopus’pallidus populations were both morphologically and genetically distinct across the studied regions. Our results provide key information to better inform conservation and management decisions for developing octopod fisheries in southeast Australia and highlight the importance of genomics tools in the conservation management of commercially and recreationally important species.

An integrative taxonomic approach reveals Octopus insularis as the dominant species in the Veracruz Reef System (southwestern Gulf of Mexico).

The common octopus of the Veracruz Reef System (VRS, southwestern Gulf of Mexico) has historically been considered as Octopus vulgaris, and yet, to date, no study including both morphological and genetic data has tested that assumption. To assess this matter, 52 octopuses were sampled in different reefs within the VRS to determine the taxonomic identity of this commercially valuable species using an integrative taxonomic approach through both morphological and genetic analyses. Morphological and genetic data confirmed that the common octopus of the VRS is not O. vulgaris and determined that it is, in fact, the recently described O. insularis. Morphological measurements, counts, indices, and other characteristics such as specific colour patterns, closely matched what had been reported for O. insularis in Brazil. In addition, sequences from cytochrome oxidase I (COI) and 16S ribosomal RNA (r16S) mitochondrial genes confirmed that the common octopus from the VRS is in the same highly supported clade as O. insularis from Brazil. Genetic distances of both mitochondrial genes as well as of cytochrome oxidase subunit III (COIII) and novel nuclear rhodopsin sequences for the species, also confirmed this finding (0–0.8%). We discuss our findings in the light of the recent reports of octopus species misidentifications involving the members of the ‘O. vulgaris species complex’ and underscore the need for more morphological studies regarding this group to properly address the management of these commercially valuable and similar taxa.

Recent expansion led to the lack of genetic structure of Sargassum aquifolium populations in Southeast Asia

Phylogeographical study of the brown macroalga, Sargassum aquifolium using nuclear internal transcribed spacer 2, plastidal RuBisCo spacer, and mitochondrial cytochrome oxidase subunit-III revealed the populations in Southeast Asia to be homogeneous. On the other hand, genetic differences were detected between populations from Southeast Asia and western Pacific Islands/Guam, suggesting the presence of genetic break between these regions. This further suggests that populations of S. aquifolium may have survived east of Sunda Shelf during the Last Glacial Maximum and recent recolonization led to homogeneity of the populations in the Sunda Shelf region. Recolonization could be facilitated by year-round reproduction of the populations and dispersal of germlings on floating thalli by coastal currents. Restricted current flow across Maluku Sea and directional equatorial current flows could have isolated the Pacific Island and Guam populations from those of Southeast Asia. Our results support the presence of multiple refugia as the source of different lineages of S. aquifolium populations with a lack of secondary contact in the post-glacial dispersal between Southeast Asia and western Pacific as the mechanisms behind the phylogeographical patterns observed.

Homogeneous population of the brown alga Sargassum polycystum in Southeast Asia: possible role of recent expansion and asexual propagation.

Southeast Asia has been known as one of the biodiversity hotspots in the world. Repeated glacial cycles during Pleistocene were believed to cause isolation of marine taxa in refugia, resulting in diversification among lineages. Recently, ocean current was also found to be another factor affecting gene flow by restricting larval dispersal in animals. Macroalgae are unique in having mode of reproduction that differs from that of animals. Our study on the phylogeographical pattern of the brown macroalga Sargassum polycystum using nuclear Internal Transcribed Spacer 2 (ITS2), plastidal RuBisCO spacer (Rub spacer) and mitochondrial cytochrome oxidase subunit-III (Cox3) as molecular markers revealed genetic homogeneity across 27 sites in Southeast Asia and western Pacific, in sharp contrast to that revealed from most animal studies. Our data suggested that S. polycystum persisted in single refugium during Pleistocene in a panmixia pattern. Expansion occurred more recently after the Last Glacial Maximum and recolonization of the newly flooded Sunda Shelf could have involved asexual propagation of the species. High dispersal ability through floating fronds carrying developing germlings may also contribute to the low genetic diversity of the species.

10.1007/s11177-008-2018-y

The nucleotide sequences of mitochondrial genome fragments containing the genes coding for cytochrome oxidase subunit III (COIII) and 12S ribosomal RNA of common European viper and Nikolsky’s viper from various habitats (Saratov, Samara, and Penza oblasts; Chuvash Republic; and the Republic of Mordovia) were determined. According to the sequencing data, all samples clustered into two groups except for a number of individuals carrying single mutations in the genes in question. One group comprised V. nikolskii from Saratov oblast and the other, V. berus from the Chuvash Republic, Republic of Mordovia, and Samara and Penza oblasts. These results comply with the available data on the karyotypes of the studied vipers of this region. Further genetic studies of V. nikolskii and V. berus from various parts of this area are necessary.

Genetic divergence of Vipera berus and Vipera nikolskii (Reptilia: Viperidae, Vipera) populations in lower Volga and adjacent territories assessed according to the sequences of cytochrome oxidase III and 12S ribosome RNA genes

The nucleotide sequences of mitochondrial genome fragments containing the genes coding for cytochrome oxidase subunit III (COIII) and 12S ribosomal RNA of common European viper and Nikolsky's viper from various habitats (Saratov, Samara, and Penza oblasts; Chuvash Republic; and the Republic of Mordovia) were determined. According to the sequencing data, all samples clustered into two groups except for a number of individuals carrying single mutations in the genes in question. One group comprised V. nikolskii from Saratov oblast and the other, V. berus from the Chuvash Republic, Republic of Mordovia, and Samara and Penza oblasts. These results comply with the available data on the karyotypes of the studied vipers of this region. Further genetic studies of V. nikolskii and V. berus from various parts of this area are necessary.

Mitochondrial dysfunction and ultrastructural damage in the hippocampus of pilocarpine-induced epileptic rat

Mitochondrial dysfunction has been implicated as a contributing factor in epileptic seizures. Present studies were carried out to decipher seizure-dependent changes in mitochondrial function and ultrastructure in the chronic condition of temporal lobe epilepsy (TLE) induced by pilocarpine in rat hippocampus. Enzyme assay revealed significant depression of the activity of mitochondrial- and nuclear-encoded cytochrome oxidase (COX). Conversely, the activity of nuclear-encoded succinate dehydrogenase (SDH) remained unchanged. Discernible mitochondrial ultrastructural damage, varying from swelling to disruption of membrane, was observed in the hippocampus. Quantitative real-time PCR and Western blotting showed the expression of mitochondrial-encoded COX subunit III (COXIII) dropped significantly during the chronic seizure activity; the corresponding expression of COX subunit IV (COXIV) displayed no significant change. Most likely, our results suggest that dysfunction of mitochondrial COX respiratory enzyme and mitochondrial ultrastructural damage in the hippocampus are associated with prolonged seizure during experimental TLE and mitochondria are more vulnerable to epilepsy.

A phylogenetic analysis of the emberizid sparrows based on three mitochondrial genes

Previous molecular phylogenetic studies have examined the taxonomic relationships among a number of typical emberizid sparrow genera. To help clarify these relationships, we sequenced a 1673 base pair fragment for the complete sequence of three mitochondrial genes: adenosine triphosphatase ( Atp8 and Atp6) and cytochrome oxidase subunit III (COIII) for 38 sparrow species, along with Passerina amoena (Cardinalidae) and Piranga ludoviciana (Thraupidae) which were selected as the outgroups. Our analysis confirms the monophyly of traditional genera such as Junco, Melospiza, and Zonotrichia. Although Calcarius and Plectrophenax are often thought to be putative emberizids, all our analyses placed these genera basal to all other sparrows examined. As observed with Calcarius, Spizella did not form a monophyletic group, with S. arborea being the sister-taxon to Passerella iliaca. Our analyses also suggest that Aimophila ruficeps is probably more closely related to the “brown towhees” ( Pipilo aberti, P. crissalis, and P. fuscus) than its putative congeners. The genus Ammodramus was also not monophyletic, since it appears that Passerculus sandwichensis is more closely related to A. henslowii and A. leconteii then either one is related to its congener A. savannarum. Finally, our analyses exhibited other unsuspected associations, such as the sister-taxon relationships between Amphispiza bilineata and the Chondestes grammacus/ Calamospiza melanocorys clade, and Amphispiza belli and Pooecetes gramineus.

Sources of Incongruence among Mammalian Mitochondrial Sequences: COII, COIII, and ND6 Genes Are Main Contributors

To investigate the origins of incongruence among mammalian mitochondrial protein-coding genes, we compiled a matrix that included 13 protein-coding-genes for 41 mammals from 14 different orders. This matrix was examined for congruence using different partitioning strategies. The incongruence length difference test showed significant incongruence among the 13 gene partitions used simultaneously, and the result was not affected by third codon or transversion weighting. In the pair-wise comparisons, significant incongruence was detected between NADH:ubiquinone oxidoreductase subunit 6 gene (ND6), cytochrome oxidase subunit II (COII), or cytochrome oxidase subunit III (COIII) gene partitioned individually against the rest of the genes. Omission of any of the 14 mammalian orders alone or in combinations from the matrix did not result in a statistically significant improvement of congruence, suggesting that taxonomic sampling will not improve congruence among the data sets. However, omission of the ND6, COII, and COIII significantly improved congruence in our data matrix. Possible origins of unusual phylogenetic properties of the three genes are discussed.

Ventricular Mitochondrial Gene Expression During Development and Following Embryonic Ethanol Exposure

The effects of chronic embryonic ethanol exposure were evaluated in chick ventricular muscle. Ethanol treatments were administered on embryonic days 11, 13, 15, and 17 and chicks were sacrificed at various time points following treatments. Fluctuations in embryonic blood ethanol levels were examined following treatments. Developmental increases in the activities of mitochondrial enzymes, cytochrome oxidase (CO) and citrate synthase (CS), were observed. Ethanol exposure resulted in a depression in CO activity, but not CS activity. Since, a maximal depression in CO activity was seen with ethanol treatments of 75 mg/100 g, this dosing paradigm was adopted for subsequent experiments. A tissue-specific effect of ethanol was demonstrated as CO activity was unchanged in atrial, liver, pectoralis, and brain tissues. The role of mitochondrial DNA replication and transcription during the developmental up-regulation and ethanol-induced down-regulation of CO activity was evaluated using a cDNA probe for cytochrome oxidase subunit III (COIII). The relative levels of COIII mRNA and mitochondrial DNA (cpm/mg protein) decreased by 3-fold and 4-fold, respectively, across the developmental time course, while CO activity increased by 3.5-fold. Therefore, increases in mitochondrial DNA and mitochondrial mRNA transcripts are unlikely to be responsible for the developmentally-regulated increases in CO activity. Similarly, embryonic ethanol exposure failed to elicit alterations in COIII mRNA levels, indicating that the ethanol-induced depression in CO activity was not transcriptionally regulated. However, ventricular mitochondrial DNA concentrations were elevated in ethanol-treated embryos, indicating that ethanol-exposure either directly or indirectly induces mitochondrial DNA replication.

Specific cleavage of pre-edited mRNAs in trypanosome mitochondrial extracts.

RNA editing in Trypanosoma brucei is a posttranscriptional processing event that results in the addition and deletion of uridine residues within several mitochondrial mRNAs. We have examined reactions involving pre-edited precursor RNAs in vitro. In this study, we report specific cleavage of pre-edited cytochrome b (CYb), cytochrome oxidase subunit II (COII), and cytochrome oxidase subunit III (COIII) mRNAs when incubated with T. brucei mitochondrial extracts. The pre-edited CYb RNA was cleaved near the 3'-most uridine addition sites, within the region where editing would be expected to commence. Pre-edited COII mRNA was similarly cleaved adjacent to its small editing domain, while pre-edited COIII RNA was cleaved at multiple sites in the region where uridine addition and deletion occurs in vivo. In contrast, edited versions of CYb, COII, and COIII RNAs were not cleaved within the editing domains. Such differential cleavage of the edited and pre-edited forms of these mRNAs suggests either a direct involvement in RNA editing or involvement in another aspect of mitochondrial gene expression requiring cleavage of pre-edited RNAs.

Specific cleavage of pre-edited mRNAs in trypanosome mitochondrial extracts.

RNA editing in Trypanosoma brucei is a posttranscriptional processing event that results in the addition and deletion of uridine residues within several mitochondrial mRNAs. We have examined reactions involving pre-edited precursor RNAs in vitro. In this study, we report specific cleavage of pre-edited cytochrome b (CYb), cytochrome oxidase subunit II (COII), and cytochrome oxidase subunit III (COIII) mRNAs when incubated with T. brucei mitochondrial extracts. The pre-edited CYb RNA was cleaved near the 3'-most uridine addition sites, within the region where editing would be expected to commence. Pre-edited COII mRNA was similarly cleaved adjacent to its small editing domain, while pre-edited COIII RNA was cleaved at multiple sites in the region where uridine addition and deletion occurs in vivo. In contrast, edited versions of CYb, COII, and COIII RNAs were not cleaved within the editing domains. Such differential cleavage of the edited and pre-edited forms of these mRNAs suggests either a direct involvement in RNA editing or involvement in another aspect of mitochondrial gene expression requiring cleavage of pre-edited RNAs.

Identification of negative-strand complements to cytochrome oxidase subunit III RNA in Trypanosoma brucei.

A substantial amount of cytochrome oxidase subunit III (COIII) mRNA continues to be synthesized de novo in Trypanosoma brucei in the presence of actinomycin D, presumably by a DNA-independent transcription process. We describe the identification of negative-strand COIII RNA molecules, characterization of their termini, and the detection of RNA-dependent RNA polymerase activity. Three lines of evidence for the existence of negative-strand COIII RNA are presented: (i) hybridization with oligonucleotide probes with the same polarity as mRNA after preliminary enrichment for putative negative-strand RNA by affinity purification; (ii) cloning and sequencing of negative-strand complements for the unedited, edited, and partially edited COIII RNA; and (iii) exact correspondence of the terminal sequences of the putative negative-strand RNA molecules to the ends of COIII RNA. The presence of negative-strand complements of COIII RNA is consistent with the notion that a significant amount of mRNA in T. brucei is synthesized by an RNA-dependent RNA polymerase with negative-strand RNA as an intermediate template.

Partially edited mRNAs for cytochrome b and subunit III of cytochrome oxidase from leishmania tarentolae mitochondria: RNA editing intermediates

Partially edited mRNAs were selected by the polymerase chain reaction and sequenced. In the case of cytochrome b, 102 out of 106 clones displayed patterns of editing that were consistent with a strictly progressive 3′ to 5′ editing process, as predicted by the guide RNA model of RNA editing. In the case of cytochrome oxidase subunit III (COIII), 177 out of 304 clones displayed strictly progressive 3′ to 5′ patterns of editing. However, the remaining 127 COIII clones displayed unexpected patterns in which upstream editing preceded downstream editing, uridines were inserted at sites not normally edited, and purine residues were deleted. We suggest that many of these RNAs are produced by normal 3′ to 5′ editing of the COIII mRNA with incorrect guide RNA molecules.

Extensive editing of the cytochrome c oxidase III transcript in Trypanosoma brucei

The gene for cytochrome oxidase subunit III (COIII) is not detected in T. brucei by nucleotide sequence analysis or by cross-hybridization studies. We have identified the COIII transcript in the T. brucei mitochondrion by RNA and cDNA sequencing. Its nucleotide and predicted amino acid sequences are very similar to those from the COIII genes in related species. A mitochondrial DNA sequence that matches the COIII transcript, except for the absence of numerous thymidines, is located upstream of the apocytochrome b gene, the position of the COIII gene in related species. We conclude that this sequence is the COIII gene and that over 50% of the T. brucei COIII transcript sequence is created by RNA editing.

Comparison of the maxicircle (mitochondrial) genomes of Leishmania tarentolae and Trypanosoma brucei at the level of nucleotide sequence.

The entire 16.7-kilobase (kb) transcribed region of the Leishmania tarentolae maxicircle was compared to the entire 15-kb transcribed region of the Trypanosoma brucei maxicircle at the nucleotide sequence level by dot matrix analysis and by alignments of individual genes. The L. tarentolae NADH dehydrogenase subunit 1 (ND1) gene was identified in a newly obtained 2.9-kb sequence. All but two regions which flank the cytochrome b gene are highly conserved in both species. One 3.1-kb region in L. tarentolae that contains the cytochrome oxidase subunit III (COIII) gene and several open reading frames corresponds to a 2-kb sequence in T. brucei with limited sequence homology that lacks the COIII gene. Another 0.6-kb region that comprises an unidentified open reading frame (open reading frame 12) in L. tarentolae is substituted by a nonhomologous 0.4-kb open reading frame in T. brucei. A short intergenic region between the ND1 gene and the maxicircle unidentified reading frame 1 gene shows limited sequence homology, and the regions between the ND4 and ND5 genes and between the COI and ND4 genes are not conserved. All of the intergenic regions share G + C richness and a similar pattern of G versus C strand bias. 1.8 kb of the L. tarentolae divergent region (DV) and around 3 kb of the T. brucei DV were also obtained. The T. brucei DV sequences were not homologous to the L. tarentolae DV sequence but were organized in a similar fashion with tandem repeats of varying complexity.

Cytochrome oxidase subunit III gene in Neurospora crassa mitochondria. Location and sequence.

We have located and sequenced the gene for cytochrome oxidase subunit III (CoIII) in Neurospora crassa mitochondria. The CoIII gene is located downstream from the small rRNA gene within a cluster of tRNA genes and is coded by the same strand as the tRNA and the rRNA genes. Like the tRNA and the rRNA genes, the CoIII gene is also flanked by the GC-rich palindromic DNA sequences which are highly conserved in N. crassa mitochondria. The CoIII coding sequence predicts a protein 269 amino acids long including 8 tryptophan residues. All 8 tryptophan residues are coded for by UGA. This supports our previous conclusion based on the anticodon sequence of N. crassa mitochondrial tryptophan tRNA and provides evidence for the notion that use of UGA as a codon for tryptophan rather than chain termination may be a feature common to most mitochondrial protein synthesis systems. The close correspondence between the amino acid composition of N. crassa CoIII and that of the protein predicted by the CoIII gene sequence suggests that unlike in mammalian mitochondria, AUA is a codon for isoleucine and not for methionine in N. crassa mitochondria. The N. crassa CoIII sequence shows strong homologies to the corresponding yeast and human proteins (53 and 47%, respectively). The overall hydrophobic character of the protein is consistent with suggestions that most of CoIII is embedded in the mitochondrial inner membrane.