Hardy-Weinberg Equilibrium in different mitochondrial haplogroups of four genes associated with neuroprotection and neurodegeneration.

BACKGROUND: Malfunctioning or damaged mitochondria result in altered energy metabolism, redox equilibrium, and cellular dynamics and is a central point in the pathogenesis of neurological disorders such as Alzheimer's disease, Parkinson's disease, Huntington's disease and Amyotrophic Lateral Sclerosis. Therefore, it is of utmost importance to identify mitochondrial genetic susceptibility markers for neurodegenerative diseases. Potential markers include the respiratory chain enzymes Riboflavin kinase (RFK), Flavin adenine dinucleotide synthetase (FAD), Succinate dehydrogenase B subunit (SDHB), and Cytochrome C1 (CYC1). These enzymes are associated with neuroprotection and neurodegeneration. OBJECTIVE: To test if variants in genes RFK, FAD, SDHB and CYC1 deviate from Hardy-Weinberg Equilibrium (HWE) in different human mitochondrial haplogroups. METHODS: Sequence variants in genes RFK, FAD, SDHB and CYC1 of 2,504 non-affected individuals of the 1,000 genomes project were used for mitochondrial haplogroup assessment and HWE calculations in different mitochondrial haplogroups. RESULTS: We show that RFK variants deviate from HWE in haplogroups G, H, L, V and W, variants of FAD in haplogroups B, J, L, U, and C, variants of SDHB in relation to the C, W, and A and CYC1 variants in B, L, U, D, and T. HWE deviation indicates action of selective pressures and genetic drift. CONCLUSIONS: HWE deviation of particular variants in relation to global populational HWE, could be, at least in part, associated with the differential susceptibility of specific populations and ethnicities to neurodegenerative diseases. Our data might contribute to the epidemiology and diagnostic/prognostic methods for neurodegenerative diseases.

Hardy-Weinberg Equilibrium in different mitochondrial haplogroups of four genes associated with neuroprotection and neurodegeneration / Equilíbrio de Hardy-Weinberg em diferentes haplogrupos mitocondriais de quatro genes associados à neuroproteção e neurodegeneração

ABSTRACT Background: Malfunctioning or damaged mitochondria result in altered energy metabolism, redox equilibrium, and cellular dynamics and is a central point in the pathogenesis of neurological disorders such as Alzheimer's disease, Parkinson's disease, Huntington's disease and Amyotrophic Lateral Sclerosis. Therefore, it is of utmost importance to identify mitochondrial genetic susceptibility markers for neurodegenerative diseases. Potential markers include the respiratory chain enzymes Riboflavin kinase (RFK), Flavin adenine dinucleotide synthetase (FAD), Succinate dehydrogenase B subunit (SDHB), and Cytochrome C1 (CYC1). These enzymes are associated with neuroprotection and neurodegeneration. Objective: To test if variants in genes RFK, FAD, SDHB and CYC1 deviate from Hardy-Weinberg Equilibrium (HWE) in different human mitochondrial haplogroups. Methods: Sequence variants in genes RFK, FAD, SDHB and CYC1 of 2,504 non-affected individuals of the 1,000 genomes project were used for mitochondrial haplogroup assessment and HWE calculations in different mitochondrial haplogroups. Results: We show that RFK variants deviate from HWE in haplogroups G, H, L, V and W, variants of FAD in haplogroups B, J, L, U, and C, variants of SDHB in relation to the C, W, and A and CYC1 variants in B, L, U, D, and T. HWE deviation indicates action of selective pressures and genetic drift. Conclusions: HWE deviation of particular variants in relation to global populational HWE, could be, at least in part, associated with the differential susceptibility of specific populations and ethnicities to neurodegenerative diseases. Our data might contribute to the epidemiology and diagnostic/prognostic methods for neurodegenerative diseases.

RESUMO Introdução: Mitocôndrias defeituosas ou danificadas resultam em alterações do metabolismo energético, equilíbrio redox e dinâmica celular e são, portanto, identificadas como o ponto central da patogênese em muitos distúrbios neurológicos, como a doença de Alzheimer, a doença de Parkinson, a doença de Huntington e a Esclerose Lateral Amiotrófica. Portanto, é de fundamental importância identificar marcadores de susceptibilidade genética mitocondrial para doenças neurodegenerativas. Entre os potenciais marcadores relevantes estão as enzimas da cadeia respiratória riboflavina quinase (RFK), flavina adenina dinucleotídeo sintetase (FAD), succinato desidrogenase subunidade B (SDHB) e citocromo C1 (CYC1). Estas enzimas estão associadas à neuroproteção e à neurodegeneração. Objetivo: Testar se variantes nas sequências dos genes RFK, FAD, SDHB e CYC1 desviam do Equilíbrio de Hardy-Weinberg (HWE) em diferentes haplogrupos mitocondriais humanos. Métodos: Neste trabalho utilizamos os variantes nos genes RFK, FAD, SDHB e CYC1 de sequências de 2.504 indivíduos não afetados do projeto de 1.000 genomas para o cálculo dos valores de HWE em diferentes haplogrupos mitocondriais. Resultados: As variantes de RFK desviam de HWE nos haplogrupos G, H, L, V e W, variantes de FAD nos haplogrupos B, J, L, U e C, variantes de SDHB em relação às variantes C, W e A e CYC1 em B, L, U, D e T. O desvio de HWE indica a ação de pressões seletivas e desvio genético. Conclusões: O desvio do HWE de variantes particulares em relação ao HWE populacional global poderia estar, pelo menos em parte, associado à suscetibilidade diferencial de populações e etnias específicas a doenças neurodegenerativas. Nossos dados podem contribuir para a epidemiologia e métodos diagnósticos/prognósticos para doenças neurodegenerativas.

Genetic diversity of medically important and emerging Candida species causing invasive infection.

BACKGROUND: Genetic variation in the ribosomal DNA (rDNA) internal transcribed spacer (ITS) region has been studied among fungi. However, the numbers of ITS sequence polymorphisms in the various Candida species and their associations with sources of invasive fungal infections remain poorly investigated. Here, we characterized the intraspecific and interspecific ITS diversity of Candida spp. strains collected from patients with bloodstream or oroesophageal candidiasis. METHODS: We selected cultures of representative medically important species of Candida as well as some rare and emerging pathogens. Identification was performed by micromorphology and by biochemical testing using an ID32C system, as well as by the sequencing of rDNA ITS. The presence of intraspecific ITS polymorphisms was characterized based on haplotype networks, and interspecific diversity was characterized based on Bayesian phylogenetic analysis. RESULTS: Among 300 Candida strains, we identified 76 C. albicans, 14 C. dubliniensis, 40 C. tropicalis, 47 C. glabrata, 34 C. parapsilosis (sensu stricto), 31 C. orthopsilosis, 3 C. metapsilosis, 21 Meyerozyma guilliermondii (C. guilliermondii), 12 Pichia kudriavzevii (C. krusei), 6 Clavispora lusitaniae (C. lusitaniae), 3 C. intermedia, 6 Wickerhamomyces anomalus (C. pelliculosa), and 2 C. haemulonii strains, and 1 C. duobushaemulonii, 1 Kluyveromyces marxianus (C. kefyr), 1 Meyerozyma caribbica (C. fermentati), 1 Pichia norvegensis (C. norvegensis), and 1 Lodderomyces elongisporus strain. Out of a total of seven isolates with inconsistent ID32C profiles, ITS sequencing identified one C. lusitaniae strain, three C. intermedia strains, two C. haemulonii strains and one C. duobushaemulonii strain. Analysis of ITS variability revealed a greater number of haplotypes among C. albicans, C. tropicalis, C. glabrata and C. lusitaniae, which are predominantly related to endogenous sources of acquisition. Bayesian analysis confirmed the major phylogenetic relationships among the isolates and the molecular identification of the different Candida spp. CONCLUSIONS: Molecular studies based on ITS sequencing are necessary to identify closely related and emerging species. Polymorphism analysis of the ITS rDNA region demonstrated its utility as a genetic marker for species identification and phylogenetic relationships as well as for drawing inferences concerning the natural history of hematogenous infections caused by medically important and emerging Candida species.

Trypanosoma cruzi: Genome characterization of phosphatidylinositol kinase gene family (PIK and PIK-related) and identification of a novel PIK gene.

Chagas disease is caused by the protozoan Trypanosoma cruzi which affects 10 million people worldwide. Very few kinases have been characterized in this parasite, including the phosphatidylinositol kinases (PIKs) that are at the heart of one of the major pathways of intracellular signal transduction. Recently, we have classified the PIK family in T. cruzi using five different models based on the presence of PIK conserved domains. In this study, we have mapped PIK genes to the chromosomes of two different T. cruzi lineages (G and CL Brener) and determined the cellular localization of two PIK members. The kinases have crucial roles in metabolism and are assumed to be conserved throughout evolution. For this reason, they should display a conserved localization within the same eukaryotic species. In spite of this, there is an extensive polymorphism regarding PIK localization at both genomic and cellular levels, among different T. cruzi isolates and between T. cruzi and Trypanosomabrucei, respectively. We showed in this study that the cellular localization of two PIK-related proteins (TOR1 and 2) in the T. cruzi lineage is distinct from that previously observed in T. brucei. In addition, we identified a new PIK gene with peculiar feature, that is, it codes for a FYVE domain at N-terminal position. FYVE-PIK genes are phylogenetically distant from the groups containing exclusively the FYVE or PIK domain. The FYVE-PIK architecture is only present in trypanosomatids and in virus such as Acanthamoeba mimivirus, suggesting a horizontal acquisition. Our Bayesian phylogenetic inference supports this hypothesis. The exact functions of this FYVE-PIK gene are unknown, but the presence of FYVE domain suggests a role in membranous compartments, such as endosome. Taken together, the data presented here strengthen the possibility that trypanosomatids are characterized by extensive genomic plasticity that may be considered in designing drugs and vaccines for prevention of Chagas disease.

The genome sequence of Leishmania (Leishmania) amazonensis: functional annotation and extended analysis of gene models.

We present the sequencing and annotation of the Leishmania (Leishmania) amazonensis genome, an etiological agent of human cutaneous leishmaniasis in the Amazon region of Brazil. L. (L.) amazonensis shares features with Leishmania (L.) mexicana but also exhibits unique characteristics regarding geographical distribution and clinical manifestations of cutaneous lesions (e.g. borderline disseminated cutaneous leishmaniasis). Predicted genes were scored for orthologous gene families and conserved domains in comparison with other human pathogenic Leishmania spp. Carboxypeptidase, aminotransferase, and 3'-nucleotidase genes and ATPase, thioredoxin, and chaperone-related domains were represented more abundantly in L. (L.) amazonensis and L. (L.) mexicana species. Phylogenetic analysis revealed that these two species share groups of amastin surface proteins unique to the genus that could be related to specific features of disease outcomes and host cell interactions. Additionally, we describe a hypothetical hybrid interactome of potentially secreted L. (L.) amazonensis proteins and host proteins under the assumption that parasite factors mimic their mammalian counterparts. The model predicts an interaction between an L. (L.) amazonensis heat-shock protein and mammalian Toll-like receptor 9, which is implicated in important immune responses such as cytokine and nitric oxide production. The analysis presented here represents valuable information for future studies of leishmaniasis pathogenicity and treatment.

Identification of genes differentially expressed in hyphae of Candida albicans

A transição morfológica de levedura para hifa é essencial para a virulência de Candida albicans. Esta transição envolve a expressão de genes hifa-específicos que estão sob o controle de fatores transcricionais. Para descobrir genes hifa-específicos utilizamos um método de triagem diferencial, onde clones de biblioteca de DNA genômico foram hibridizados com sondas de cDNA de levedura e hifa. Dois clones com aumento de expressão em hifa foram selecionados. O sequenciamento dos insertos destes clones permitiu a identificação de dois genes metabólicos importantes: CaHXT7 (high-affinity hexose transporter) e CaYLL34 (da família AAA ATPase). As ORFs completas destes genes foram caracterizadas e a análise das proteínas hipotéticas revelou que: (1) CaHxt7p tem um domínio de transportador de hexose e (2) CaYll34 tem dois domínios AAA ATPase. Este é o primeiro estudo que demonstra aumento de expressão de genes metabólicos em hifas de C. albicans. Ainda, a associação dos produtos de CaHXT7 e CaYLL34 com a formação de hifas torna estas proteínas potenciais novos alvos para drogas antifúngicas.

Identification of genes differentially expressed in hyphae of Candida albicans

The ability to switch from yeast to hyphal forms is essential for Candida albicans virulence. This morphological switch involves the expression of hyphal-specific genes under the control of transcriptional factors. To contribute to the discovery of hyphal-specific genes, we used a differential screening method where clones of a genomic DNA library were hybridized with yeast and hyphal cDNA probes. Two clones with increased expression in hyphae were selected for study. Sequencing these clones, we found that they encoded two important metabolic genes, CaHXT7 (high-affinity hexose transporter) and CaYLL34 (member of the AAA ATPase family). CaHXT7 and CaYLL34 ORFs were completely determined. Analyses of the putative proteins show that: (1) CaHxt7p has one hexose transporter domain and (2) CaYll34p has two AAA ATPase domains. These results show, for the first time, increased expression of metabolic genes in C. albicans hyphae. Also, because the proteins encoded by CaHXT7 and CaYLL34 may be necessary for the switching to hyphae, they could be new targets for antifungal drugs.

A transição morfológica de levedura para hifa é essencial para a virulência de Candida albicans. Esta transição envolve a expressão de genes hifa-específicos que estão sob o controle de fatores transcricionais. Para descobrir genes hifa-específicos utilizamos um método de triagem diferencial, onde clones de biblioteca de DNA genômico foram hibridizados com sondas de cDNA de levedura e hifa. Dois clones com aumento de expressão em hifa foram selecionados. O sequenciamento dos insertos destes clones permitiu a identificação de dois genes metabólicos importantes: CaHXT7 (high-affinity hexose transporter) e CaYLL34 (da família AAA ATPase). As ORFs completas destes genes foram caracterizadas e a análise das proteínas hipotéticas revelou que: (1) CaHxt7p tem um domínio de transportador de hexose e (2) CaYll34 tem dois domínios AAA ATPase. Este é o primeiro estudo que demonstra aumento de expressão de genes metabólicos em hifas de C. albicans. Ainda, a associação dos produtos de CaHXT7 e CaYLL34 com a formação de hifas torna estas proteínas potenciais novos alvos para drogas antifúngicas.