A la lumière, l’activité carboxylase de la RuBisCO permet de fixer le CO2 inorganique en matière organique, sous forme de 3-phosphoglycérate (3-PGA), qui sera utilisé pour la biosynthèse de sucres, d’acides organiques et aminés, de la paroi végétale etc. Cependant, elle possède aussi une activité oxygénase qui produit du 3-PGA et du 2-phosphoglycolate. Ce dernier composé étant toxique, il est métabolisé en 3-PGA par le cycle photorespiratoire qui se déroule dans le chloroplaste, le peroxysome et la mitochondrie. Malgré une perte partielle en carbone et en azote, l’importance de la photorespiration pour les plantes est illustré par les phénotypes néfastes que les mutants d’enzymes photorespiratoires présentent dans l’air (comme un retard de croissance, la chlorose, et de la létalité) et qui sont absents en fort CO2. Ceci pourrait refléter des interactions étroites entre la photorespiration et le métabolisme primaire des plantes. Afin de mieux comprendre ces interactions et la mise en place des phénotypes photorespiratoires, des mutants pour la glycolate oxydase (GOX) et la glutamate:glyoxylate aminotransférase ont été caractérisés à travers plusieurs analyses complémentaires: des échanges gazeux, de la fluorescence chlorophyllienne, du marquage des métabolites avec du 13C, des dosages de métabolites, de cofacteurs, et de la RuBisCO. Les résultats montrent que, suite à un transfert de fort CO2 dans l’air, l’inhibition de la photosynthèse observée chez nos mutants est principalement due à un défaut du recyclage du carbone photorespiratoire qui diminue l’activité de la RuBisCO. Cette inhibition photosynthétique a un impact négatif sur la quantité de RuBisCO dans les feuilles de ces mutants par rapport aux plantes contrôles. De plus, lorsque l’inhibition de la photosynthèse est trop importante chez nos mutants photorespiratoires, la carence en carbone déclenche de la sénescence dans leurs feuilles âgées. En parallèle, une comparaison des paramètres cinétiques de la GOX d’A. thaliana (plante en C3) et de Z. mays (plante en C4) associée à la mesure d’effets isotopiques 13C et 2H a révélé que ces enzymes partageaient des paramètres Michaéliens équivalents pour le glycolate, ainsi qu’un mécanisme réactionnel identique mettant en jeu un transfert d’hydrure. / In the light, the RuBisCO carboxylase activity assimilates inorganic CO2 into organic compounds, via the production of 3-phosphoglycerate (3-PGA) that is used for the biosynthesis of sugars, organic and amino acids, plant cell walls etc. However, it also has an oxygenase activity that makes 3-PGA and 2-phosphoglycolate (2-PG). The toxic 2-PG is metabolized to 3-PGA by the photorespiratory cycle, which takes place in chloroplasts, peroxisomes and mitochondria. Despite a partial loss of carbon and nitrogen, the importance of photorespiration for growth can be seen by the negative phenotypes exhibited by photorespiratory enzyme mutants in air (i.e. slow growth, leaf chlorosis, and sometimes lethality), which are not observed under high CO2 conditions. This may reflect the metabolic interactions between photorespiration and plant primary metabolism. To better understand such interactions and the development of photorespiratory phenotypes, mutants for glycolate oxidase (GOX) and glutamate:glyoxylate aminotransferase have been characterized by several complementary methods: analysis of gas exchanges, chlorophyll fluorescence,13C-labeling of metabolites, measurements of metabolites, cofactors and RuBisCO levels. The results show that, after a high CO2-to-air transfer, the inhibition of photosynthesis in the mutants is mainly due to a defect in photorespiratory carbon recycling leading to a decreased RuBisCO activity. The inhibition of carbon assimilation negatively impacts mutant leaf RuBisCO content when compared to wild-type plants. In the mutants, when photosynthetic inhibition is too high, the resulting carbon starvation triggers the onset of senescence in their old leaves. In parallel to this work, a comparison of the kinetic parameters of GOX from A. thaliana (C3 plant) and Z. mays (C4 plant) coupled to measurements of 13C and 2H kinetic isotopic effects showed that these enzymes share similar Michaelian parameters for glycolate, and a similar hydride transfer reaction mechanism.

Le virus Epstein-Barr (EBV), un membre de la famille des Herpesviridae, est associé à la mononucléose infectieuse et à différents types de cancers comme le lymphome de Burkitt, les lymphomes post-transplantation ou encore le carcinome du nasopharynx. Ce virus est capable de persister à vie dans l’organisme en combinant des phases de latence et des phases de multiplication active. L’autophagie est un processus cellulaire primordial qui conduit à la dégradation et au recyclage de protéines à longue durée de vie et d’organites endommagés ou vieillissants. Elle contribue non seulement à maintenir l’homéostasie cellulaire mais aussi à s’adapter aux conditions environnementales. Souvent décrite comme un mécanisme antiviral, l’autophagie est contrecarrée par de nombreux virus. Elle peut également être détournée à leur profit. Il a été démontré que l’EBV est capable de stimuler l’autophagie durant le cycle lytique et d’échapper à la dégradation dans les autolysosomes en bloquant la maturation des autophagosomes. Le but de cette étude était d’identifier des protéines virales impliquées dans la modulation du processus autophagique par l’EBV. Nous avons démontré que l’expression ectopique de BHRF1, une protéine transmembranaire de 17kDa orthologue de la protéine cellulaire Bcl-2, module l’autophagie.Alors que Bcl-2 est une protéine anti-autophagique, nous avons établi par différentes approches que l’expression de BHRF1 conduit à l’accumulation d’autophagosomes. De plus, en utilisant une sonde tandem bifluorescente LC3 (mRFP-GFP-LC3) pour étudier le flux autophagique, nous avons montré que BHRF1 stimule l’autophagie. BHRF1 est engagée dans un complexe avec Beclin1, une protéine de la machinerie autophagique. Nous avons établi que BHRF1 est localisée au niveau des membranes mitochondriales et du réticulum endoplasmique (RE). L’expression de BHRF1 est associée à une réorganisation du réseau mitochondrial conduisant à la formation d’agrégats mitochondriaux juxta-nucléaires. Considérant l’importance des microtubules dans l’autophagie et le transport des mitochondries, nous avons exploré la dynamique des microtubules et les modifications post-traductionnelles de la tubuline après expression de BHRF1. Nous avons observé un recrutement d’acétyl-tubuline autour des mito-aggresomes associé à un réseau intact de microtubules. Nos résultats ont montré que le réseau de microtubules et l’hyper-acétylation de l’alpha-tubuline sont nécessaires pour former les mito-aggrésomes induits par BHRF1. Par différentes approches, nous avons démontré le rôle de BHRF1 dans l’induction de la mitophagie, un processus qui entraine la clairance des mitochondries endommagées par autophagie. Considérant le rôle des mitochondries endommagées dans l’induction de l’apoptose, nous suggérons que le rôle anti-apoptotique de BHRF1 pourrait être associé à l’induction de la mitophagie. / Epstein-Barr virus (EBV), a member of the Herpesviridae family, is associated with infectious mononucleosis and with several types of cancers including Burkitt’s lymphoma, post-transplant B-cell lymphoma disease and nasopharyngeal carcinoma. This virus is able to establish persistent infection and to undergo lytic cycle after reactivation. Autophagy is a critical cellular process leading to degradation of long lasting proteins and damaged or aging organelles. It contributes not only to maintain cell homeostasis but also to the adaptation to environmental stresses. Sometimes, autophagy is described as an antiviral mechanism, and viruses have evolved multiple strategies to subvert it or to hijack it to their own profit. It has been reported that EBV is able to stimulate autophagy during lytic cycle and then to escape degradation within autolysosomes by blocking autophagosomes maturation. The aim of my study was to identify EBV viral proteins involved in this modulation. Among the numerous viral proteins encoded by EBV, we have identified BHRF1, a transmembrane protein homolog of cellular protein Bcl-2, which was able to modulate autophagy by ectopic expression.Whereas Bcl-2 is an anti-autophagic protein, we demonstrated by different approaches that BHRF1 expression leads to accumulation of autophagosomes. Moreover, using tandem-fluorescent-tagged LC3 (mRFP-GFP-LC3), which is based on different pH stability of GFP and mRFP fluorescent proteins, for monitoring autophagic flux, we clearly confirmed that BHRF1 stimulates autophagy. By co-immunoprecipitation we demonstrated that BHRF1 is part of acomplex including Beclin1, a protein of the autophagic machinery. We characterized the subcellular localization of BHRF1, and report that BHRF1 is localized in mitochondria and ER membranes. Expression of BHRF1 leads to a complete reorganization of the mitochondria network to form juxtanuclear mitochondrial aggregates. Based on the importance of microtubules on both autophagy and mitochondria transport, we explored microtubule dynamics and tubulin post-translational modifications after BHRF1 expression. We observed a clustering of acetyl-tubulin around the mito-aggresomes associated with an intact microtubules network. Our results showed that the microtubules network and the hyperacetylation of alpha-tubulin were both required to form BHRF1-induced mito-aggresomes.By different approaches, we demonstrated the role of BHRF1 in the induction of mitophagy, a process which promotes the clearance of impaired mitochondria by autophagy. We hypothesized that the role of BHRF1 to protect against apoptosis and to promote cell survival is related to the induction of selective autophagy.

In most tropical river systems there has been a lack of integrated ecological research to investigate the dynamics and impacts of invasive species on recipient river systems. This is in sharp contrast to temperate river systems. This thesis investigated the nature, extent, and impact of Nile tilapia, Oreochromis niloticus (Linnaeus, 1758), on indigenous congenerics within the Limpopo River basin in northern South Africa. An integrated approach was adopted to gain a better understanding of factors that allow Nile tilapia to be a successful invader and also to gain an insight into its invasion rate and conservation implications within South Africa. Morphometric and genetic variation between Nile tilapia, indigenous congenerics and their associated hybrids were determined. Intermediate meristic characters obscured the identification of hybrid specimens from pure morpho-specimens and species identity was only confirmed through mtDNA analysis. Preliminary evidence points to unidirectional hybridization among Oreochromis congeners in the Limpopo River system. The hypothesis that bigger Nile tilapia males may have a competitive advantage over spawning grounds and in female mate choice is proposed. The trophic ecology of Nile tilapia was investigated using both stomach contents and stable isotope analysis. A high similarity in stomach contents was observed but interspecific differences were revealed in the isotopic composition of diets that suggest fine scale patterns of resource partitioning that could be achieved by the ability of fish to selectively feed on what is immediately available and the ability to perceive the dynamics that determine food resource availability. Ecological niche models were used to determine the potential invasive range of Nile tilapia and revealed broad invasive potential over most river systems in southern Africa that overlapped the natural range of endemic congenerics. It was noted that model performance and the degree of niche conservatism varied significantly with variable selection and spatial extent of study area. This implied that the spatial distribution of suitable and unsuitable environmental variables varied between the native and introduced ranges of Nile tilapia and also indicated the ability of Nile tilapia to survive in conditions incongruent with its native range. The extreme hardiness and adaptive life history characteristics of Nile tilapia have probably predisposed it to be a successful invader in novel systems within southern Africa. Lastly, a qualitative risk assessment method was developed as a potential application to determine the risk of establishment and spread of the invasive Nile tilapia. Results showed that in the absence of quantitative data on ecosystem structure and functioning, habitat suitability analysis in terms of known physiological tolerance limits to minimum water temperature, presence or absence of dams, seasonality of river flows and the presence of indigenous fish species of concern could be adequate for identifying vulnerable river systems. The model developed also provides an objective method that is easy to implement, modify and improve on as new data become available. Furthermore, the model can be applied to highlight areas of uncertainty where future research should be directed. / Thesis (PhD)--University of Pretoria, 2012. / Zoology and Entomology / Unrestricted

The objective of this study was to characterize the mitochondrial genomes of the species within the genus Ceratocystis and investigate the evolutionary process of the ribosomal RNA cistron found within these fungi. Ceratocystis incorporates a number of pathogenic species affecting a variety of hosts, making the study of these fungi economically significant. The fortuitous identification of a Ceratocystis species, C. manginecans, which contained two different internally transcribed spacer sequence types within the ribosomal rRNA cistron, enabled a study of concerted evolution in this fungus. Using this non-model organism we were able to show empirical evidence for unequal crossing over and gene conversion as the ultimate forces acting on this gene region dictating a concerted evolutionary effect. We suggest that this process is true for all eukaryotes. Using the knowledge drawn from previously characterized and annotated mitochondrial genomes of other eukaryotes, the genomes of three Ceratocystis species, namely Ceratocystis fimbriata, Ceratocystis albifundus and Ceratocystis moniliformis were fully assembled and annotated for comparative analysis. This comparative study addressed the genome size, gene content, tRNA presence as well as intron types and their homing endonucleases found among these three mitochondrial genomes. An interspecies characterization was then undertaken using the mitochondrial genomes of six different C. albifundus isolates from different geographical locations in Africa. Genetic variation and similarities among these isolates supports the previous hypothesis that the origin of this fungus is Southern Africa. It is hoped that the research presented in this thesis will contribute to the improved understanding of the mitochondrial genomes in Ceratocystis species. / Thesis (PhD)--University of Pretoria, 2013. / gm2013 / Genetics / Unrestricted

Introduction: Cryopreservation of human sperm is considered a routine practice in assisted reproduction laboratories. Semen samples are mainly cryopreserved as a back-up for procedures, donor sperm, and validation of samples from human immunodeficiency virus-positive patients. Human immunodeficiency virus semen samples generally result in a low yield of purified spermatozoa after decontamination. These samples need to be cryopreserved for later use. Unlike conventional cryopreservation, vitrification does not use harmful cryoprotectants, thereby potentially reducing sperm damage. Vitrification is not yet common practice for sperm cryopreservation in assisted reproduction. The aim of this study was to establish the feasibility of utilising vitrification as an alternative to current conventional cryopreservation of spermatozoa. Methods: Semen samples were collected from human immunodeficiency virus-negative patients seeking diagnostic assistance from the unit. All samples were processed according to the unit’s standard protocol. For Study 1A (n=10) washed samples were divided and cryopreserved using three different cryopreservation media, and two different freezing protocols. In Study 1B (n=15), washed samples were divided and preserved using cryoprotectant-free vitrification in 100 μl, 300 μl and 500 μl volumes. For Study 2 (n=35) washed samples were split and cryopreserved using cryoprotectant-free vitrification (utilizing the volume that resulted in the highest quality spermatozoa in Study 1B) and conventional slow freezing (using the medium and protocol that resulted in superior quality spermatozoa in Study 1A). Post thawing, motility and kinetic parameters (Studies 1 and 2), viability (Study 1), mitochondrial membrane potential (Study 2), and DNA fragmentation (Study 2) of the two groups were compared. vi Results: Study 1A indicated that cryopreserving spermatozoa using Freezing Medium resulted in the highest quality spermatozoa with regards to motility and viability (p<0.05). Comparing the two preservation protocols, no conclusion could be reached on which protocol yielded superior results (p>0.05). The RBL freezing method is shorter, simpler and requires less equipment, and was therefore deemed the preferred method. Study 1B showed that the larger vitrification volumes (300 μl and 500 μl) yielded better spermatozoa in terms of motility and viability (p<0.05). No significant difference was observed with respect to the 300 μl and 500 μl vitrification volume groups. For practical reasons, 300 μl volumes will provide sufficient sperm for any procedure and, the intermediate volume ensures that more than one straw can be preserved. Study 2 found that cryoprotectant-free vitrification resulted in spermatozoa with significantly higher mitochondrial membrane potential and significantly lower apoptosis post thawing (p<0.05). Discussion: Conventional cryopreservation methods may compromise various sperm parameters and final yield. In this study, cryopreservation and cryoprotectant-free vitrification had equivalent outcomes with respect to sperm motility. However, the latter method yielded superior results in terms of ΔΨ and DNA sperm fragmentation. In conclusion, vitrification is an easy, rapid and more affordable technique that requires no special equipment. Using vitrification for purified sperm samples of patients could potentially result in a better post thaw quality for ART procedures. / Dissertation (MSc)--University of Pretoria, 2013. / gm2014 / Obstetrics and Gynaecology / unrestricted

Orientador: Maria Tercília Vilela de Azeredo Oliveira / Resumo: A subordem Heteroptera possui uma enorme diversidade de insetos distribuída em sete infraordens, dentre as quais utilizamos Gerromorpha, Nepomorpha, Cimicomorpha e Pentatomomorpha. As espécies que foram estudadas da primeira infraordem, representando os insetos semi-aquáticos, são Limnogonus aduncus e Mesovelia mulsanti, da segunda infraordem, representando os aquáticos, são Belostoma anurum, Martarega sp. e Buenoa unguis, da terceira infraordem Zelus sp. e Teleonemia sp. e da última infraordem, representando juntas os terrestres, são Largus sp., Stenocoris sp., Zicca pulchra e Dysdercus sp. para descrever as ultraestruturas de suas espermátides e posterior comparação. Durante o processo de espermiogênese em geral, ocorre uma série de modificações nas espermátides antes dessas serem transformadas em espermatozoide, inclusive nos insetos. Neste trabalho verificamos por meio da Microscopia Eletrônica de Transmissão, as modificações ocorridas nas espermátides das infraordens anteriormente mencionadas. Na maioria dos insetos com hábitat aquático foi identificada uma maior variação morfológica com relação aos derivados mitocondriais, apresentando-se tanto com simetria reniforme, com simetria e padrão diferente, quanto com assimetria. Com relação aos terrestres, foi identificado predominantemente um padrão morfológico reniforme com uma área de cada derivado mitocondrial menor que a do axonema. Somente a espécie B. unguis, de hábitat aquático, evidenciou um padrão atípico, assimétri... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The suborder Heteroptera has an enormous diversity of insects distributed in seven infraorders, among which Nepomorpha, Gerromorpha, Pentatomomorpha e Cimicomorpha, The species that have been studied from the first infraorder, representing semi-aquatic insects, are Limnogonus aduncus and Mesovelia mulsanti, from the second order, representing the aquatic ones, Belostoma anurum, Martarega sp. and Buenoa unguis, from the third infraorder Zelus sp. and Teleonemia sp. and the last infraorder, representing terrestrials together, are Largus sp., Stenocoris sp., Zicca pulchra and Dysdercus sp. to describe the ultrastructures of their spermatides and further comparison. During the spermiogenesis process in general, a series of changes in spermatids occur before they are transformed into sperm, including in insects In this work we verified through Transmission Electron Microscopy, the changes occurred in the spermatids of the abovementioned infraorders. In the majority of insects with aquatic habitat, a greater morphological variation was identified in relation to mitochondrial derivatives, presenting both with reniform symmetry, with different symmetry and pattern, and with asymmetry. With respect to terrestrials, a predominantly reniform morphological pattern was identified with an area of each mitochondrial derivative smaller than the axoneme. Only the species B. unguis, of aquatic habitat, showed an atypical, asymmetric pattern of mitochondrial derivatives, in relation to that com... (Complete abstract click electronic access below) / Doutor

Despite recent advances in the understanding of the molecular bases of hematological malignancies, the specific mechanisms on how they originate and why some subtypes are more prevalent than others still remain to be elucidated. These two important aspects have been even more difficult to analyze when dealing with individuals under immune suppression because other factors must be considered. Questions still remain as to why individuals with AIDS tend to develop lymphoproliferative disorders differently from those observed in individuals under iatrogenic immunosuppressive therapy. Most of lymphomas occurring in transplant recipients are B-cell neoplasias typically associated with Epstein-Barr virus (EBV) infection. In contrast, only about 50% of lymphomas of patients with AIDS are associated with lymphotrophic herpesviruses such as EBV and Kaposi's sarcoma-associated herpesvirus (KSHV). No known infectious agent has been detected in the remaining 50% of AIDS-associated lymphomas, suggesting the involvement of novel viruses or unique molecular mechanisms. Since most oncogenic viruses persist as episomal circular viral genomes in the nuclei of tumor cells, we developed a method to visualize and identify covalently closed circular DNA (cccDNA) in lymphoma samples. Although this study revealed no novel viruses, we identified concatemers of the mitochondrial genome in all lymphoma samples tested. We further studied in detail one AIDS-associated lymphoma (denominated EL) whose mitochondrial DNA primarily consisted of tandem head-to-tail genome duplications. Insertion of cytosine residues was noted in the EL mitochondrial genome sequence near the origin of replication. EL cells responded weakly to Fas-apoptotic stimulus, displayed reduced mitochondrial activity and mass, and produced higher levels of reactive oxygen species (ROS) than control cells. Screening of several other AIDS-associated lymphomas and established lymphoma cell lines revealed a different kind of mitochondrial genome concatemers consisting of interlinks of DNA monomer molecules. Concatemers were not detected in normal T-lymphocytes suggesting an association with neoplastic transformation. This dissertation describes the two distinct types of mitochondrial genome concatemers identified in transformed lymphoid cells and presents a detailed analysis of their structure and implications in cellular homeostasis.

I developed and tested a protocol for determining the sex of individual pinnipeds using the sex-chromosome specific genes ZFX and ZFY. I screened a total of 368 seals (168 crabeater, Lobodon carcinophagus; 159 Weddell, Leptonychotes weddellii; and 41 Ross, Ommatophoca rossii) of known or unknown sex and compared the molecular sex to the sex assigned at the time of collection in the Ross and Amundsen seas, Antarctica. Discrepancies ranged from 0.0% - 6.7% among species. It is unclear, however, if mis-assignment of sex occurred in situ or in the laboratory. It also is possible, however, that the assigned morphological and molecular sex both are correct, owing perhaps to developmental effects of environmental pollution. I sequenced a portion (ca 475 bp) of the mitochondrial control region of Weddell seals (N = 181); crabeater seals (N = 143); and Ross seals (N = 41). I resolved 251 haplotypes with a haplotype diversity of 0.98 to 0.99. Bayesian estimates of Θ from the program LAMARC ranged from 0.075 for Weddell seals to 0.576 for crabeater seals. I used the values of theta to estimate female effective population sizes (NEF), which were 40,700 to 63,000 for Weddell seals, 44,400 to 97,800 for Ross seals, and 358,500 to 531,900 for crabeater seals. Weddell seals and crabeater seals had significant, unimodal mean pairwise difference mismatch distributions (p = 0.56 and 0.36, respectively), suggesting that their populations expanded suddenly around 731,000 years ago (Weddell seals) and around 1.6 million years ago (crabeater seals). Both of these expansions occurred during times of intensified glaciations and may have been fostered by expanding pack ice habitat. Autosomal microsatellite based NEs were 147,850 for L. Weddellii, 344,950 for O. rossii, and 939,600 for L. carcinophagus. I screened one X-linked microsatellite (Lw18), which yielded a larger NE estimate for O. rossii than the other two species. Microsatellite NE estimates are compared with previously published mitochondrial NE estimates and this comparison indicates that the Ross seal may have a serially monogamous system of mating. I find no sign of a recent, sustained genetic bottleneck in any of the three species.

Siberia is an ideal region for exploring population histories from a molecular anthropological perspective given the diverse human populations, in terms of linguistic affiliation and lifestyle, currently inhabiting this geographically large region. As such, this thesis explores new methodologies for the investigation of the genetic histories of Siberian populations. While previous genetic work in this area of the world was able to provide detailed insights into paternal histories based on Y chromosomal data, it was not as successful on the maternal side. There existed difficulties in exploring the complex maternal demographic histories due to high levels of sequence identity between individuals in different populations when using only a very small region of the mitochondrial DNA (mtDNA), known as the hypervariable region I (HV1). This realization led to the initial focus of this dissertation which was to identify and test improved methods of sequencing entire mtDNA genomes. This was necessary because the mtDNA genomes that were published for human Siberian populations and across the globe prior to the work described here were chosen based on specific sub-sample selection criteria that introduced an ascertainment bias rendering them unusable for population-wide analyses. After testing multiple next generation DNA sequencing methods, I helped develop a sequencing library preparation method based on multiplexing and hybridization enrichment of mtDNAs for sequencing by synthesis that has since become widely used in labs across the globe. Comparing the same samples sequenced by both the traditional and new methods for five ethnolinguistic populations showed that these new methods were robust and could lead to different inferences about population histories while avoiding a sampling bias. Based on the results of this thesis it is now recommended for researchers to sequence complete mtDNA genomes for all relevant samples within a collection. By applying these methods to additional Siberian populations it was possible to better describe maternal population contact and identify demographic changes over time. This additional information allowed for the identification of putative drops in the maternal effective population sizes in the Siberian populations examined here. When examining the potential migrations and population contact between Turkic-speaking Yakuts and the Tungusic-speaking Even and Evenks, there exists a differential sharing of haplotypes suggesting that the Tungusic speaking populations herein were already in the northern region and split prior to the expansion of the Yakuts into their territory. The putative origin of the Yakuts as being around Lake Baikal was given additional support from the analyses included in this study and the origins of the Dolgans were shown to predominately include the admixture of Yakuts and Evenks.

Trichomonas vaginalis is a human pathogen that affects annually approximately 258 million people worldwide. This parasite possesses organelles of mitochondrial origin called hydrogenosomes, which generate ATP under anaerobic conditions. The identification of the protein content at the subcellular level may provide new targets for antiparasitic drugs developments as well as it contributes for our understanding of the organelles function and evolution. The availability of protocols for organelles purification and the complete genome sequence allow the study of the organellar proteomes using mass spectrometry and bioinformatics, providing a powerful strategy that combine cell biology and proteomics. In our research, we used several approaches to identify the protein composition in hydrogenosomes and mitosomes. We performed transcriptomic and proteomic analysis to investigate the molecular responses of Trichomonas vaginalis upon iron availability. Furthermore, the changes in the proteome during the development of metronidazole resistance were also studied. The organelles separated by differential and Optiprep-sucrose gradient centrifugation were analyzed with nano- RP-HPLC/MALDI-TOF/TOF. We also used Triton X-114 phase partitioning to separate membrane proteins and iTRAQ technique to label the peptides...