New types of metacaspases in phytoplankton reveal diverse origins of cell death proteases

Metacaspases are evolutionarily distant homologs of caspases that are found outside the metazoan and are known to have key roles in programmed cell death (PCD). Two types of metacaspases (types I and II) have been defined in plants based on their domain structures; these have similarities to metazoan ‘initiator'' and ‘executioner'' caspases. However, we know little about metacaspases in unicellular organisms and even less about their roles in cell death. We identified a novel group of metacaspases in sequenced phytoplanktonic protists that show domain architectures distinct from either type I or II enzymes; we designate them as type III. Type III metacaspases exhibit a rearrangement of domain structures between N- and C-terminus. In addition, we found a group of metacaspase-like proteases in phytoplankton that show sequence homology with other metacaspases, but defy classification in conventional schemes. These metacaspase-like proteases exist in bacteria alongside a variant of type I metacaspases and we propose these bacterial metacaspases are the origins of eukaryotic metacaspases. Type II and III metacaspases were not detected in bacteria and they might be variants of bacterial type I metacaspases that evolved in plants and phytoplanktonic protists, respectively, during the establishment of plastids through the primary and secondary endosymbiotic events. A complete absence of metacaspases in protists that lost plastids, such as oömycetes and ciliates indicates the gene loss during the plastid-to-nucleus gene transfer. Taken together, our findings suggest endosymbiotic gene transfer (EGT) is a key mechanism resulting in the evolutionary diversity of cell death proteases.     

Environmental levels of oestrogenic and antiandrogenic compounds feminize digit ratios in male rats and their unexposed male progeny

Digit length ratios, especially the second-to-fourth digit ratio (2D : 4D), are associated with various pathological and behavioural conditions in many species including humans and are dependent upon prenatal androgen to oestrogen balance. It is unknown whether digit ratios are modified by environmental exposure to ubiquitous endocrine disruptors. We studied the effect on adult male Wistar rat digit ratios of a gestational exposure to the oestrogenic and antiandrogenic compounds bisphenol A (BPA), genistein and vinclozolin, in low doses, and in combination with investigating in parallel a possible sexual dimorphism of this trait. We also investigated the effects on the male progeny not exposed during gestation. X-rays were taken of the left and right forepaws, and 2D–5D proximal to distal phalanx distances were measured by a standardized procedure based on semi-automatic image analysis. We provide evidence that there is a sexual dimorphism of digit ratios in the Wistar rat, and we found that BPA alone or in combination with genistein and vinclozolin significantly feminized digit ratios in male rats. Intriguingly, significant feminization of digit ratios was also found in the unexposed male progeny of males that had been exposed to compound mixtures. In conclusion, prenatal environmental levels of endocrine-active substances permanently disrupt digit ratios. Digit ratio measurement in adults is thus a promising biomarker of prenatal exposure to low-dose endocrine disruptors in rodents, with potential implications for future studies in humans.     

Effects of silicate resupply to silicate‐deprived Thalassiosira weissflogii (Bacillariophyceae) in stationary or senescent phase: short‐term patterns of growth and cell death

The ability of nutrient‐deprived phytoplankton to recover in the short term when nutrients are resupplied has been studied for nitrogen and phosphorus, but the case for silicate (Si) is poorly understood. Si‐limited Thalassiosira weissflogii (Grunow) Fryxell et Hasle (grown in batch culture) was harvested in stationary phase (when cell numbers stopped increasing ~2 d after Si depletion) and senescence (when cell numbers declined ~4 d after Si depletion) and Si was resupplied at different concentrations (from 0 to 100 μM). Cell numbers, proportion of dead cells, variable fluorescence emissions (F_v/F_m), and activities of proteases were measured during Si depletion and for 24 h after Si resupply. As Si was depleted, the specific growth rate declined, dead cells increased from ~2% in log phase, to ~25% in stationary phase to over 35% in senescence, and activities of proteases associated with cell death increased several‐fold. Concentration‐dependent recovery of growth rate was seen after 24 h for cultures resupplied with Si in stationary phase but not in senescence. However, resupply of Si at 100 μM to stationary phase cultures alone increased protease activity to nearly the levels seen in senescence. Differences in the responses to Si resupply suggest that the ability and time to recover from Si depletion depend not only on the growth phase but also on the concentration resupplied.     

Increased expression and altered subunit composition of proteasomes induced by continuous proteasome inhibition establish apoptosis resistance and hyperproliferation of Burkitt lymphoma cells

The proteasome is the main protease for extralysosomal protein degradation in eukaryotic cells, and constitutes a sophisticated high molecular mass proteinase complex underlying a tightly coordinated expression and assembly of multiple subunits and subcomplexes. Here we show that continuous inhibition of proteasomal chymotrypsin-like peptidase activity by the proteasome inhibitor bortezomib induces in human Namalwa Burkitt lymphoma cells increased de novo biogenesis of proteasomes accompanied by increased expression of the proteasome maturation protein POMP, increased expression of 19S-20S-19S proteasomes, and abrogation of expression of beta 1i, beta 2i and beta 5i immunosubunits and PA28 in favor of increased expression of constitutive proteolytic beta1, beta2 and beta 5 subunits and 19S regulatory complexes. These alterations of proteasome expression and subunit composition are accompanied by an increase in proteasomal caspase-like, trypsin-like and chymotrypsin-like peptidase activities, not inhibitable by high doses of bortezomib. Cells harboring these proteasomal alterations display rapid proliferation and cell cycle progression, and acquire resistance to apoptosis induced by proteasome inhibitors, gamma-irradiation and staurosporine. This acquired apoptosis resistance is accompanied by de novo expression of anti-apoptotic Hsp27 protein and the loss of ability to accumulate and stabilize pro-apoptotic p53 protein. Thus, increased expression, altered subunit composition and increased activity of proteasomes constitute a hitherto unknown adaptive and autoregulatory feedback mechanism to allow cells to survive the lethal challenge of proteasome inhibition and to establish a hyperproliferative and apoptosis-resistant phenotype.     

Virus morphological diversity and relationship to bacteria and chlorophyll across a freshwater trophic gradient in the Lake Michigan watershed

The effect of starvation on sexual reproduction in cyclic parthenogenetic rotifers has been studied using life history experiment. Short-time starvation of rotifers that experienced starvation immediately after hatching from resting eggs can cause high induction of sexual reproduction up to the 10th generation. However, it is not clear whether the induction of sexual reproduction can occur beyond the 10th generation. To investigate this phenomenon, we conducted a sex induction study using the monogonont rotifer Brachionus manjavacas. Newborn stem females were starved for 12 h, while controls were supplied with 7.0 × 10⁶ cells ml^?1 of Nannochloropsis oculata. In a life history experiment, the rotifers were individually cultured in 96-well microplates containing 0.2 ml of seawater (22 ppt) in each well at 25 °C with daily feeding thereafter. Mixis induction in offspring from starved stem females was significantly higher than in those from non-starved stem females up to the 40th generation. The effect of accumulative generations increased mixis induction up to the 20th generation. Effect on future generations of the rise in mixis ratio by the starvation to stem females may facilitate colonization by favoring population growth via female parthenogenesis and by decreasing food requirements for survival and reproduction.     

Alkaline cooking and tortilla quality in maize grains from the humid,tropical lands of Mexico

Maize (Zea mays L.) tortilla is the major staple food for the Mexican population. Nine tropical maize genotypes were evaluated. All samples had white grains, a common characteristic in tropical maize, and therefore they were appropriate for nixtamalized flour industry. Grain, flour, masa and tortilla characteristics of each maize genotype were evaluated. Length, width, thickness, weight of 1000 grains and hardness of grain were determined. Moisture content, proteins, fat, ash, mean particle size, water absorption index, enthalpy, and flour temperature were also evaluated. Adhesiveness and cohesiveness were evaluated in masa. Moisture content, protein, capacity to puff up, roll making, tension and cutting strength were determined in tortillas. There were significant differences (p≤0.05) in most of the evaluated characteristics. Grain length values varied between 9.26 and 11.02 mm for populations 23 and 22, respectively. Grain hardness oscillated between 11.17 (population 32) and 14.75 (landrace Mejen). According to the weight of 1000 grains most genotypes had small grains. The minimum and maximum moisture values of flour and tortillas were 8.33-9.99% and 46.20-50.36%, respectively. The texture of tortillas elaborated from population 32 and landrace Mejen had the lowest tension and cutting strength, resulting the best genotypes for making tortilla.     

Physiological responses of phytoplankton communities in the Irish Sea to simulated upwelling

Understanding the dynamics of upwelling systems, especially the interactions between nutrients and light, has benefited from the application of models of varying complexity. Validation of such models using unialgal cultures or field observations has often proven difficult, but short-term incubations of contained natural assemblages and use of instantaneous physiological indicators offer an alternative approach. In May and June 1996, phytoplankton communities deep in the euphotic zone were sampled from nearly identical physical environments. Replicate samples (20 l volume) were incubated on deck at 50% surface irradiance with either no nutrient additions (Controls) or additions of 20 μM nitrate (Enrichments). Over 24 h, variable fluorescence (F _v:F _m), nitrate reductase activity (NR), nutrients, chlorophyll a and particulate C and N were monitored. Initial chlorophyll a (～3 μg l^?1), phosphate (～0.2 μM), nitrate (～1.5 μM) and silicate (～3 μM) were similar in both months. Changes in NR and F _v:F _m indicated clear physiological responses to changes in irradiance and added nitrate that differed between months. In May, Controls and Enrichments responded in the same way. F _v:F _m stayed constant (0.5), chlorophyll a increased slightly, and NR activity increased markedly in all samples. In contrast, in June, treatments responded quite differently. F _v:F _m was near the theoretical maximum (0.7–0.8) initially and remained constant in Enrichments, but fell sharply in Controls. Declines in controls were also seen for chlorophyll a, and NR activity. Thus, the addition of 20 μM nitrate had a significant effect even though ambient levels of nitrate (>1 μM) should not have been limiting. Small (<20 μm) flagellates predominated in the May samples, but in June large and chain-forming centric diatoms constituted a significant proportion of the phytoplankton community. We conclude that the response of a phytoplankton community to environmental changes can depend on factors that are poorly represented by bulk measurements of chlorophyll, nutrients and particulate elements.