Mitochondrial Involvement in Vertebrate Speciation? The Case of Mito-nuclear Genetic Divergence in Chameleons

Compatibility between the nuclear (nDNA) and mitochondrial (mtDNA) genomes is important for organismal health. However, its significance for major evolutionary processes such as speciation is unclear, especially in vertebrates. We previously identified a sharp mtDNA-specific sequence divergence between morphologically indistinguishable chameleon populations (Chamaeleo chamaeleon recticrista) across an ancient Israeli marine barrier (Jezreel Valley). Because mtDNA introgression and gender-based dispersal were ruled out, we hypothesized that mtDNA spatial division was maintained by mito-nuclear functional compensation. Here, we studied RNA-seq generated from each of ten chameleons representing the north and south populations and identified candidate nonsynonymous substitutions (NSSs) matching the mtDNA spatial distribution. The most prominent NSS occurred in 14 nDNA-encoded mitochondrial proteins. Increased chameleon sample size (N = 70) confirmed the geographic differentiation in POLRMT, NDUFA5, ACO1, LYRM4, MARS2, and ACAD9. Structural and functionality evaluation of these NSSs revealed high functionality. Mathematical modeling suggested that this mito-nuclear spatial divergence is consistent with hybrid breakdown. We conclude that our presented evidence and mathematical model underline mito-nuclear interactions as a likely role player in incipient speciation in vertebrates

Gene Expression Patterns of Oxidative Phosphorylation Complex I Subunits Are Organized in Clusters

After the radiation of eukaryotes, the NUO operon, controlling the transcription of the NADH dehydrogenase complex of the oxidative phosphorylation system (OXPHOS complex I), was broken down and genes encoding this protein complex were dispersed across the nuclear genome. Seven genes, however, were retained in the genome of the mitochondrion, the ancient symbiote of eukaryotes. This division, in combination with the three-fold increase in subunit number from bacteria (N = ∼14) to man (N = 45), renders the transcription regulation of OXPHOS complex I a challenge. Recently bioinformatics analysis of the promoter regions of all OXPHOS genes in mammals supported patterns of co-regulation, suggesting that natural selection favored a mechanism facilitating the transcriptional regulatory control of genes encoding subunits of these large protein complexes. Here, using real time PCR of mitochondrial (mtDNA)- and nuclear DNA (nDNA)-encoded transcripts in a panel of 13 different human tissues, we show that the expression pattern of OXPHOS complex I genes is regulated in several clusters. Firstly, all mtDNA-encoded complex I subunits (N = 7) share a similar expression pattern, distinct from all tested nDNA-encoded subunits (N = 10). Secondly, two sub-clusters of nDNA-encoded transcripts with significantly different expression patterns were observed. Thirdly, the expression patterns of two nDNA-encoded genes, NDUFA4 and NDUFA5, notably diverged from the rest of the nDNA-encoded subunits, suggesting a certain degree of tissue specificity. Finally, the expression pattern of the mtDNA-encoded ND4L gene diverged from the rest of the tested mtDNA-encoded transcripts that are regulated by the same promoter, consistent with post-transcriptional regulation. These findings suggest, for the first time, that the regulation of complex I subunits expression in humans is complex rather than reflecting global co-regulation

Complete mitochondrial DNA sequences provide new insights into the Polynesian motif and the peopling of Madagascar

More than a decade of mitochondrial DNA (mtDNA) studies have given the 'Polynesian motif' renowned status as a marker for tracing the late-Holocene expansion of Austronesian speaking populations. Despite considerable research on the Polynesian motif in Oceania, there has been little equivalent work on the western edge of its expansion - leaving major issues unresolved regarding the motif's evolutionary history. This has also led to considerable uncertainty regarding the settlement of Madagascar. In this study, we assess mtDNA variation in 266 individuals from three Malagasy ethnic groups: the Mikea, Vezo, and Merina. Complete mtDNA genome sequencing reveals a new variant of the Polynesian motif in Madagascar; two coding region mutations define a Malagasy-specific sub-branch. This newly defined 'Malagasy motif' occurs at high frequency in all three ethnic groups (13-50%), and its phylogenetic position, geographic distribution, and estimated age all support a recent origin, but without conclusively identifying a specific source region. Nevertheless, the haplotype's limited diversity, similar to those of other mtDNA haplogroups found in our Malagasy groups, best supports a small number of initial settlers arriving to Madagascar through the same migratory process. Finally, the discovery of this lineage provides a set of new polymorphic positions to help localize the Austronesian ancestors of the Malagasy, as well as uncover the origin and evolution of the Polynesian motif itself

DNA topoisomerases participate in fragility of the oncogene RET

Fragile site breakage was previously shown to result in rearrangement of the RET oncogene, resembling the rearrangements found in thyroid cancer. Common fragile sites are specific regions of the genome with a high susceptibility to DNA breakage under conditions that partially inhibit DNA replication, and often coincide with genes deleted, amplified, or rearranged in cancer. While a substantial amount of work has been performed investigating DNA repair and cell cycle checkpoint proteins vital for maintaining stability at fragile sites, little is known about the initial events leading to DNA breakage at these sites. The purpose of this study was to investigate these initial events through the detection of aphidicolin (APH)-induced DNA breakage within the RET oncogene, in which 144 APHinduced DNA breakpoints were mapped on the nucleotide level in human thyroid cells within intron 11 of RET, the breakpoint cluster region found in patients. These breakpoints were located at or near DNA topoisomerase I and/or II predicted cleavage sites, as well as at DNA secondary structural features recognized and preferentially cleaved by DNA topoisomerases I and II. Co-treatment of thyroid cells with APH and the topoisomerase catalytic inhibitors, betulinic acid and merbarone, significantly decreased APH-induced fragile site breakage within RET intron 11 and within the common fragile site FRA3B. These data demonstrate that DNA topoisomerases I and II are involved in initiating APH-induced common fragile site breakage at RET, and may engage the recognition of DNA secondary structures formed during perturbed DNA replication

Updated Three-Stage Model for the Peopling of the Americas

Background: We re-assess support for our three stage model for the peopling of the Americas in light of a recent report that identified nine non-Native American mitochondrial genome sequences that should not have been included in our initial analysis. Removal of these sequences results in the elimination of an early (i.e.,40,000 years ago) expansion signal we had proposed for the proto-Amerind population. Methodology/Findings: Bayesian skyline plot analysis of a new dataset of Native American mitochondrial coding genomes confirms the absence of an early expansion signal for the proto-Amerind population and allows us to reduce the variation around our estimate of the New World founder population size. In addition, genetic variants that define New World founder haplogroups are used to estimate the amount of time required between divergence of proto-Amerinds from the Asian gene pool and expansion into the New World. Conclusions/Significance: The period of population isolation required for the generation of New World mitochondrial founder haplogroup-defining genetic variants makes the existence of three stages of colonization a logical conclusion. Thus, our three stage model remains an important and useful working hypothesis for researchers interested in the peopling of th

Origin and Post-Glacial Dispersal of Mitochondrial DNA Haplogroups C and D in Northern Asia

More than a half of the northern Asian pool of human mitochondrial DNA (mtDNA) is fragmented into a number of subclades of haplogroups C and D, two of the most frequent haplogroups throughout northern, eastern, central Asia and America. While there has been considerable recent progress in studying mitochondrial variation in eastern Asia and America at the complete genome resolution, little comparable data is available for regions such as southern Siberia – the area where most of northern Asian haplogroups, including C and D, likely diversified. This gap in our knowledge causes a serious barrier for progress in understanding the demographic pre-history of northern Eurasia in general. Here we describe the phylogeography of haplogroups C and D in the populations of northern and eastern Asia. We have analyzed 770 samples from haplogroups C and D (174 and 596, respectively) at high resolution, including 182 novel complete mtDNA sequences representing haplogroups C and D (83 and 99, respectively). The present-day variation of haplogroups C and D suggests that these mtDNA clades expanded before the Last Glacial Maximum (LGM), with their oldest lineages being present in the eastern Asia. Unlike in eastern Asia, most of the northern Asian variants of haplogroups C and D began the expansion after the LGM, thus pointing to post-glacial re-colonization of northern Asia. Our results show that both haplogroups were involved in migrations, from eastern Asia and southern Siberia to eastern and northeastern Europe, likely during the middle Holocene

The role of DNA damage response pathways in chromosome fragility in Fragile X syndrome

FRAXA is one of a number of fragile sites in human chromosomes that are induced by agents like fluorodeoxyuridine (FdU) that affect intracellular thymidylate levels. FRAXA coincides with a >200 CGG•CCG repeat tract in the 5′ UTR of the FMR1 gene, and alleles prone to fragility are associated with Fragile X (FX) syndrome, one of the leading genetic causes of intellectual disability. Using siRNA depletion, we show that ATR is involved in protecting the genome against FdU-induced chromosome fragility. We also show that FdU increases the number of γ-H2AX foci seen in both normal and patient cells and increases the frequency with which the FMR1 gene colocalizes with these foci in patient cells. In the presence of FdU and KU55933, an ATM inhibitor, the incidence of chromosome fragility is reduced, suggesting that ATM contributes to FdU-induced chromosome fragility. Since both ATR and ATM are involved in preventing aphidicolin-sensitive fragile sites, our data suggest that the lesions responsible for aphidicolin-induced and FdU-induced fragile sites differ. FRAXA also displays a second form of chromosome fragility in absence of FdU, which our data suggest is normally prevented by an ATM-dependent process

Mitochondrial Pseudogenes in the Nuclear Genomes of Drosophila

Mitochondrial pseudogenes in nuclear chromosomes (numts) have been detected in the genomes of a diverse range of eukaryotic species. However, the numt content of different genomes and their properties is not uniform, and study of these differences provides insight into the mechanisms and dynamics of genome evolution in different organisms. In the genus Drosophila, numts have previously only been identified on a genome-wide scale in the melanogaster subgroup. The present study extends the identification to 11 species of the Drosophila genus. We identify a total of 302 numts and show that the numt complement is highly variable in Drosophilids, ranging from just 4 in D. melanogaster to 67 in D. willistoni, broadly correlating with genome size. Many numts have undergone large-scale rearrangements in the nucleus, including interruptions, inversions, deletions and duplications of sequence of variable size. Estimating the age of the numts in the nucleus by phylogenetic tree reconstruction reveals the vast majority of numts to be recent gains, 90% having arisen on terminal branches of the species tree. By identifying paralogs and counting duplications among the extant numts we estimate that 23% of extant numts arose through post-insertion duplications. We estimate genus average rates of insertion of 0.75 per million years, and a duplication rate of 0.010 duplications per numt per million years

Parental diabetes status reveals association of mitochondrial DNA haplogroup J1 with type 2 diabetes

<p>Abstract</p> <p>Background</p> <p>Although mitochondrial dysfunction is consistently manifested in patients with Type 2 Diabetes mellitus (T2DM), the association of mitochondrial DNA (mtDNA) sequence variants with T2DM varies among populations. These differences might stem from differing environmental influences among populations. However, other potentially important considerations emanate from the very nature of mitochondrial genetics, namely the notable high degree of partitioning in the distribution of human mtDNA variants among populations, as well as the interaction of mtDNA and nuclear DNA-encoded factors working in concert to govern mitochondrial function. We hypothesized that association of mtDNA genetic variants with T2DM could be revealed while controlling for the effect of additional inherited factors, reflected in family history information.</p> <p>Methods</p> <p>To test this hypothesis we set out to investigate whether mtDNA genetic variants will be differentially associated with T2DM depending on the diabetes status of the parents. To this end, association of mtDNA genetic backgrounds (haplogroups) with T2DM was assessed in 1055 Jewish patients with and without T2DM parents ('DP' and 'HP', respectively).</p> <p>Results</p> <p>Haplogroup J1 was found to be 2.4 fold under-represented in the 'HP' patients (p = 0.0035). These results are consistent with a previous observation made in Finnish T2DM patients. Moreover, assessing the haplogroup distribution in 'DP' versus 'HP' patients having diabetic siblings revealed that haplogroup J1 was virtually absent in the 'HP' group.</p> <p>Conclusion</p> <p>These results imply the involvement of inherited factors, which modulate the susceptibility of haplogroup J1 to T2DM.</p

Stable Expression of Antibiotic-Resistant Gene ble from Streptoalloteichus hindustanus in the Mitochondria of Chlamydomonas reinhardtii

The mitochondrial expression of exogenous antibiotic resistance genes has not been demonstrated successfully to date, which has limited the development of antibiotic resistance genes as selectable markers for mitochondrial site-directed transformation in Chlamydomonas reinhardtii. In this work, the plasmid pBSLPNCB was constructed by inserting the gene ble of Streptoalloteichus hindustanus (Sh ble), encoding a small (14-kilodalton) protective protein into the site between TERMINVREP-Left repeats and the cob gene in a fragment of mitochondrial DNA (mtDNA) of C. reinhardtii. The fusion DNA-construct, which contained TERMINVREP-Left, Sh ble, cob, and partial nd4 sequence, were introduced into the mitochondria of the respiratory deficient dum-1 mutant CC-2654 of C. reinhardtii by biolistic particle delivery system. A large number of transformants were obtained after eight weeks in the dark. Subsequent subculture of the transformants on the selection TAP media containing 3 ìg/mL Zeomycin for 12 months resulted in genetically modified transgenic algae MT-Bs. Sequencing and Southern analyses on the mitochondrial genome of the different MT-B lines revealed that Sh ble gene had been integrated into the mitochondrial genome of C. reinhardtii. Both Western blot, using the anti-BLE monoclonal antibody, and Zeomycin tolerance analysis confirmed the presence of BLE protein in the transgenic algal cells. It indicates that the Sh ble gene can be stably expressed in the mitochondria of C. reinhardtii