Human mitochondrial disease-like symptoms caused by a reduced tRNA aminoacylation activity in flies

The translation of genes encoded in the mitochondrial genome requires specific machinery that functions in the organelle. Among the many mutations linked to human disease that affect mitochondrial translation, several are localized to nuclear genes coding for mitochondrial aminoacyl-transfer RNA synthetases. The molecular significance of these mutations is poorly understood, but it is expected to be similar to that of the mutations affecting mitochondrial transfer RNAs. To better understand the molecular features of diseases caused by these mutations, and to improve their diagnosis and therapeutics, we have constructed a Drosophila melanogaster model disrupting the mitochondrial seryl-tRNA synthetase by RNA interference. At the molecular level, the knockdown generates a reduction in transfer RNA serylation, which correlates with the severity of the phenotype observed. The silencing compromises viability, longevity, motility and tissue development. At the cellular level, the knockdown alters mitochondrial morphology, biogenesis and function, and induces lactic acidosis and reactive oxygen species accumulation. We report that administration of antioxidant compounds has a palliative effect of some of these phenotypes. In conclusion, the fly model generated in this work reproduces typical characteristics of pathologies caused by mutations in the mitochondrial aminoacylation system, and can be useful to assess therapeutic approaches.     

Genetic variation in the Cytb gene of human cerebral Taenia solium cysticerci recovered from clinically and radiologically heterogeneous patients with neurocysticercosis

Neurocysticercosis (NC) is a clinically and radiologically heterogeneous parasitic disease caused by the establishmentof larval Taenia solium in the human central nervous system. Host and/or parasite variations may be related to this observed heterogeneity. Genetic differences between pig and human-derived T. solium cysticerci have been reported previously. In this study, 28 cysticerci were surgically removed from 12 human NC patients, the mitochondrial gene that encodes cytochrome b was amplified from the cysticerci and genetic variations that may be related to NC heterogeneity were characterised. Nine different haplotypes (Ht), which were clustered in four haplogroups (Hg), were identified. Hg 3 and 4 exhibited a tendency to associate with age and gender, respectively. However, no significant associations were found between NC heterogeneity and the different T. solium cysticerci Ht or Hg. Parasite variants obtained from patients with similar NC clinical or radiological features were genetically closer than those found in groups of patients with a different NC profile when using the Mantel test. Overall, this study establishes the presence of genetic differences in the Cytb gene of T. solium isolated from human cysticerci and suggests that parasite variation could contribute to NC heterogeneity.     

Doxorubicin treatment activates a Z-VAD-sensitive caspase, which causes deltapsim loss, caspase-9 activity, and apoptosis in Jurkat cells

Doxorubicin induces caspase-3 activation and apoptosis in Jurkat cells but inhibition of this enzyme did not prevent cell death, suggesting that another caspase(s) is critically implicated. Western blot analysis of cell extracts indicated that caspases 2, 3, 4, 6, 7, 8, 9, and 10 were activated by doxorubicin. Cotreatment of cells with the caspase inhibitors Ac-DEVD-CHO, Z-VDVAD-fmk, Z-IETD-fmk, and Z-LEHD-fmk alone or in combination, or overexpression of CrmA, prevented many morphological features of apoptosis but not loss of mitochondrial membrane potential (delta(psi)m), phospatidilserine exposure, and cell death. Western blot analysis of cells treated with doxorubicin in the presence of inhibitors allowed elucidation of the sequential order of caspase activation. Z-IETD-fmk or Z-LEHD-fmk, which inhibit caspase-9 activity, blocked the activation of all caspases studied, lamin B degradation, and the development of apoptotic morphology, but not cell death. All morphological and biochemical features of apoptosis, as well as cell death, were prevented by cotreatment of cells with the general caspase inhibitor Z-VAD-fmk or by overexpression of Bcl-2. Doxorubicin cytotoxicity was also blocked by the protein synthesis inhibitor cycloheximide. Delayed addition of Z-VAD-fmk after doxorubicin treatment, but prior to the appearance of cells displaying a low delta(psi)m, prevented cell death. These results, taken together, suggest that the key mediator of doxorubicin-induced apoptosis in Jurkat cells may be an inducible, Z-VAD-sensitive caspase (caspase-X), which would cause delta(psi)m loss, release of apoptogenic factors from mitochondria, and cell death.     

Long-chain fatty acids bound to alpha-fetoprotein and to serum albumin from fetal and adult pig

1. Pig alpha-fetoprotein (AFP) and albumin were isolated from fetal serum by DEAE-Sephadex ion exchange chromatography combined with Cibacron Blue-Sepharose and trypsin-Sepharose adsorptions. 2. AFP, fetal albumin and adult albumin carried 2.6, 2.4, and 1.9 moles of fatty acids per mole of protein, respectively. 3. Most of fatty acids bound to AFP were polyunsaturated: mainly arachidonic (20:4, n-6) and docosahexaenoic (22:6, n-3) acids, which accounted respectively for 21.7 and 18.8% of the total fatty acids. 4. By contrast, the fatty acids found in the albumins (fetal and adult) were preferentially saturated and monounsaturated. 5. Arachidonic acid was a minor component in both albumins, and no docosahexaenoic acid was detected.     

Betel Nut Chewing Is Strongly Associated With General and Central Obesity in Chinese Male Middle‐aged Adults

Betel nut chewing has been reported to increase the risk of cardiovascular disease and all‐cause mortality. The reason is unclear. In this study, we investigated the association between betel nut chewing and general obesity (BMI ≥25 kg/m²) and central obesity (waist circumference (WC) ≥90 cm). A total of 1,049 male subjects, aged ≥40 years, were recruited from Taichung city in Taiwan in 2004. The relationships between betel nut chewing and general and central obesity were studied by multiple linear and logistic regression analyses. The prevalence of current and former betel nut chewing was 7.0 and 10.5% in our male Taiwanese cohort. Current/former betel nut chewers had a higher prevalence of general and central obesity when compared with individuals who had never chewed betel nut. Adjusted for age, diabetes, hypertension, lipids, smoking, alcohol drinking, physical activity, income, and education level, the odds ratios (ORs; 95% confidence intervals) of general and central obesity among the lower consumption of betel nut chewers were 1.78 (1.07, 2.96) and 1.19 (0.70, 2.02), respectively, compared to 2.01 (1.18, 3.41) and 1.89 (1.10, 3.23), respectively, among higher consumption chewers compared to individuals who had never chewed betel nut. The increasing ORs of general and central obesity with higher betel nut consumption revealed dose–response effects. Using multiple linear regression analyses, after adjusting for potential confounders, betel nut consumption was statistically significantly associated with BMI and WC. In conclusion, betel nut chewing was independently associated with general and central obesity in Taiwanese men. Dose–response effects of the association between betel nut consumption and general obesity as well as central obesity were found.     

Refugia, differentiation and postglacial migration in arctic-alpine Eurasia, exemplified by the mountain avens (Dryas octopetala L.)

Many arctic-alpine organisms have vast present-day ranges across Eurasia, but their history of refugial isolation, differentiation and postglacial expansion is poorly understood. The mountain avens, Dryas octopetala sensu lato, is a long-lived, wind-dispersed, diploid shrub forming one of the most important components of Eurasian tundras and heaths in terms of biomass. We address differentiation and migration history of the species with emphasis on the western and northern Eurasian parts of its distribution area, also including some East Greenlandic and North American populations (partly referred to as the closely related D. integrifolia M. Vahl). We analysed 459 plants from 52 populations for 155 amplified fragment length polymorphisms (AFLP) markers. The Eurasian plants were separated into two main groups, probably reflecting isolation and expansion from two major glacial refugia, situated south and east of the North European ice sheets, respectively. Virtually all of northwestern Europe as well as East Greenland have been colonized by the Southern lineage, whereas northwest Russia, the Tatra Mountains and the arctic archipelago of Svalbard have been colonized by the Eastern lineage. The data indicate a contact zone between the two lineages in northern Scandinavia and possibly in the Tatra Mountains. The two single populations analysed from the Caucasus and Altai Mountains were most closely related to the Eastern lineage but were strongly divergent from the remaining eastern populations, suggesting survival in separate refugia at least during the last glaciation. The North American populations grouped with those from East Greenland, irrespective of their taxonomic affiliation, but this may be caused by independent hybridization with D. integrifolia and therefore not reflect the true relationship between populations from these areas.     

Effect of 24-Epibrassinolide on Chlorophyll Fluorescence and Photosynthetic CO<Subscript>2</Subscript> Assimilation in <Emphasis Type="Italic">Vicia faba</Emphasis> Plants Treated with the Photosynthesis-Inhibiting Herbicide Terbutryn

Simultaneous measurements of chlorophyll (Chl) fluorescence and CO₂ assimilation (A) in Vicia faba leaves were taken during the first weeks of growth to evaluate the protective effect of 24-epibrassinolide (EBR) against damage caused by the application of the herbicide terbutryn (Terb) at pre-emergence. V. faba seeds were incubated for 24 h in EBR solutions (2 × 10⁻⁶ or 2 × 10⁻⁵ mM) and immediately sown. Terb was applied at recommended doses (1.47 or 1.96 kg ha⁻¹) at pre-emergence. The highest dose of Terb strongly decreased CO₂ assimilation, the maximum quantum yield of PSII photochemistry in the dark-adapted state (F _V/F _M), the nonphotochemical quenching (NPQ), and the effective quantum yield (ΔF/F′_M) during the first 3–4 weeks after plant emergence. Moreover, Terb increased the basal quantum yield of nonphotochemical processes (F ₀/F _M)_, the degree of reaction center closure (1 − q _p), and the fraction of light absorbed in PSII antennae that was dissipated via thermal energy dissipation in the antennae (1 − F′_V/F′_M). The herbicide also significantly reduced plant growth at the end of the experiment as well as plant length, dry weight, and number of leaves. The application of EBR to V. faba seeds before sowing strongly diminished the effect of Terb on fluorescence parameters and CO₂ assimilation, which recovered 13 days after plant emergence and showed values similar to those of control plants. The protective effect of EBR on CO₂ assimilation was detected at a photosynthetic photon flux density (PFD) of 650 μmol m⁻² s⁻¹ and the effect on ΔF/F′_M and photosynthetic electron transport (J) was detected under actinic lightings up to 1750 μmol m⁻² s⁻¹. The highest dose of EBR also counteracted the decrease in plant growth caused by Terb, and plants registered the same growth values as controls.     

The mitochondrial DNA of <Emphasis Type="Italic">Dictyostelium discoideum</Emphasis>: complete sequence,gene content and genome organization

We present an overview of the gene content and organization of the mitochondrial genome of Dictyostelium discoideum. The mitochondria genome consists of 55,564?bp with an A + T content of 72.6%. The identified genes include those for two ribosomal RNAs (rnl and rns), 18 tRNAs, ten subunits of the NADH dehydrogenase complex (nad1, 2, 3, 4, 4L, 5, 6, 7, 9 and 11), apocytochrome b (cytb), three subunits of the cytochrome oxidase (cox1/2 and 3), four subunits of the ATP synthase complex (atp1, 6, 8 and 9), 15 ribosomal proteins, and five other ORFs, excluding intronic ORFs. Notable features of D. discoideum mtDNA include the following. (1) All genes are encoded on the same strand of the DNA and a universal genetic code is used. (2) The cox1 gene has no termination codon and is fused to the downstream cox2 gene. The 13 genes for ribosomal proteins and four ORF genes form a cluster 15.4?kb long with several gene overlaps. (3) The number of tRNAs encoded in the genome is not sufficient to support the synthesis of mitochondrial protein. (4) In total, five group I introns reside in rnl and cox1/2, and three of those in cox1/2 contain four free-standing ORFs. We compare the genome to other sequenced mitochondrial genomes, particularly that of Acanthamoeba castellanii.