Homocysteine inhibits the proliferation and invasive potential of HT-1080 human fibrosarcoma cells

The impairment of homocysteine metabolism has been related to several disorders and diseases. Recently, homocysteine has been shown to inhibit key steps of angiogenesis, including endothelial cell proliferation, invasion, and remodeling of the extracellular matrix. Since these are also key steps in tumor invasion and metastasis, it can be hypothesized that homocysteine can also interfere in these processes. Therefore, we studied the effects of homocysteine on tumor proliferation and invasion, as well as on urokinase, a key extracellular matrix-degrading protease, using a model human tumor cell line. This study demonstrates that, in fact, homocysteine inhibits HT-1080 proliferation and invasion, and is a potent inhibitor of tumor cell urokinase expression.     

The two paralogue phoN (phosphinothricin acetyl transferase) genes of Pseudomonas putida encode functionally different proteins

Phosphinothricin (PPT) is a non‐specific inhibitor of glutamine synthetase that has been employed as herbicide for selection of transgenic plants expressing cognate resistance genes. While the soil bacterium Pseudomonas putida KT2440 has been generally considered PPT‐sensitive, inspection of its genome sequence reveals the presence of two highly similar open reading frames (PP_1924 and PP_4846) encoding acetylases with a potential to cause tolerance to the herbicide. To explore this possibility, each of these genes (named phoN1 and phoN2) was separately cloned and their activities examined in vivo and in vitro. Genetic and biochemical evidence indicated that phoN1 encodes a bona fide PPT‐acetyl transferase, the expression of which suffices to make P. putida tolerant to high concentrations of the herbicide. In contrast, PhoN2 does not act on PPT but displays instead activity against methionine sulfoximine (MetSox), another glutamine synthetase inhibitor. When the geometry of the substrate‐binding site of PhoN1 was grafted with the equivalent residues of the predicted PhoN2 structure, the resulting protein increased significantly MetSox resistance of the expression host concomitantly with the loss of activity on PPT. These observations uncover intricate biochemical and genetic interactions among soil microorganisms and how they can be perturbed by exposure to generic herbicides in soil.     

Histamine prevents polyamine accumulation in mouse C57.1 mast cell cultures.

The effects of histamine on polyamine uptake and metabolism was studied in a mouse mast cell line (C57.1), as a cell model in which both biogenic amines are important for maintaining cell function and viability. Results obtained after incubations with exogenous histamine indicated that histamine prevents polyamine accumulation by affecting polyamine uptake. A plasma membrane transport system for polyamines has been also studied in mast cells. It seems to be a Na(+)-dependent uptake with high affinity for both spermine and spermidine and lower affinity for putrescine and agmatine. Polyamine uptake was reduced in both cells treated with exogenous histamine and histamine-preloaded cells. However, ornithine decarboxylase activity and cell proliferation were not affected by histamine. Incubation with histamine enhanced the spermidine/spermine acetyl transferase induction caused by N(1)-ethyl-N(11)-[(cyclopropyl)methyl]-4,8-diazaundecane, suggesting that polyamine acetylation could be another mechanism by which histamine prevents polyamine accumulation in C57.1 mast cells.     

Comparison of midgut bacterial diversity in tropical caterpillars (Lepidoptera: Saturniidae) fed on different diets

As primary consumers of foliage, caterpillars play essential roles in shaping the trophic structure of tropical forests. The caterpillar midgut is specialized in plant tissue processing; its pH is exceptionally alkaline and contains high concentrations of toxic compounds derived from the ingested plant material (secondary compounds or allelochemicals) and from the insect itself. The midgut, therefore, represents an extreme environment for microbial life. Isolates from different bacterial taxa have been recovered from caterpillar midguts, but little is known about the impact of these microorganisms on caterpillar biology. Our long-term goals are to identify midgut symbionts and to investigate their functions. As a first step, different diet formulations were evaluated for rearing two species of tropical saturniid caterpillars. Using the polymerase chain reaction (PCR) with primers hybridizing broadly to sequences from the bacterial domain, 16S rRNA gene libraries were constructed with midgut DNA extracted from caterpillars reared on different diets. Amplified rDNA restriction analysis indicated that bacterial sequences recovered from the midguts of caterpillars fed on foliage were more diverse than those from caterpillars fed on artificial diet. Sequences related to Methylobacterium sp., Bradyrhizobium sp., and Propionibacterium sp. were detected in all caterpillar libraries regardless of diet, but were not detected in a library constructed from the diet itself. Furthermore, libraries constructed with DNA recovered from surface-sterilized eggs indicated potential for vertical transmission of midgut symbionts. Taken together, these results suggest that microorganisms associated with the tropical caterpillar midgut may engage in symbiotic interactions with these ecologically important insects.     

Direct Proteolytic Cleavage of NLRP1B Is Necessary and Sufficient for Inflammasome Activation by Anthrax Lethal Factor

Inflammasomes are multimeric protein complexes that respond to infection by recruitment and activation of the Caspase-1 (CASP1) protease. Activated CASP1 initiates immune defense by processing inflammatory cytokines and by causing a rapid and lytic cell death called pyroptosis. Inflammasome formation is orchestrated by members of the nucleotide-binding domain and leucine-rich repeat (NLR) or AIM2-like receptor (ALR) protein families. Certain NLRs and ALRs have been shown to function as direct receptors for specific microbial ligands, such as flagellin or DNA, but the molecular mechanism responsible for activation of most NLRs is still poorly understood. Here we determine the mechanism of activation of the NLRP1B inflammasome in mice. NLRP1B, and its ortholog in rats, is activated by the lethal factor (LF) protease that is a key virulence factor secreted by Bacillus anthracis, the causative agent of anthrax. LF was recently shown to cleave mouse and rat NLRP1 directly. However, it is unclear if cleavage is sufficient for NLRP1 activation. Indeed, other LF-induced cellular events have been suggested to play a role in NLRP1B activation. Surprisingly, we show that direct cleavage of NLRP1B is sufficient to induce inflammasome activation in the absence of LF. Our results therefore rule out the need for other LF-dependent cellular effects in activation of NLRP1B. We therefore propose that NLRP1 functions primarily as a sensor of protease activity and thus could conceivably detect a broader spectrum of pathogens than just B. anthracis. By adding proteolytic cleavage to the previously established ligand-receptor mechanism of NLR activation, our results illustrate the remarkable flexibility with which the NLR architecture can be deployed for the purpose of pathogen-detection and host defense.     

Repeated mass strandings of Miocene marine mammals from Atacama Region of Chile point to sudden death at sea

Marine mammal mass strandings have occurred for millions of years, but their origins defy singular explanations. Beyond human causes, mass strandings have been attributed to herding behaviour, large-scale oceanographic fronts and harmful algal blooms (HABs). Because algal toxins cause organ failure in marine mammals, HABs are the most common mass stranding agent with broad geographical and widespread taxonomic impact. Toxin-mediated mortalities in marine food webs have the potential to occur over geological timescales, but direct evidence for their antiquity has been lacking. Here, we describe an unusually dense accumulation of fossil marine vertebrates from Cerro Ballena, a Late Miocene locality in Atacama Region of Chile, preserving over 40 skeletons of rorqual whales, sperm whales, seals, aquatic sloths, walrus-whales and predatory bony fish. Marine mammal skeletons are distributed in four discrete horizons at the site, representing a recurring accumulation mechanism. Taphonomic analysis points to strong spatial focusing with a rapid death mechanism at sea, before being buried on a barrier-protected supratidal flat. In modern settings, HABs are the only known natural cause for such repeated, multispecies accumulations. This proposed agent suggests that upwelling zones elsewhere in the world should preserve fossil marine vertebrate accumulations in similar modes and densities.     

Kinetics of infective juvenile production of the entomopathogenic nematode Steinernema carpocapsae in submerged monoxenic culture

The effects of culture medium formulations on the kinetics of infective juvenile (IJ) production of the entomopathogenic nematode Steinernema carpocapsae in submerged monoxenic culture, were studied at the cylindrical-bottle scale using six culture media containing agave juice from Agave spp. among other ingredients. The IJ production kinetics was well modelled through a re-parameterised 3-parameter Gompertz model with kinetic parameters: IJ-lag phase lambda (IJ) (day), maximum IJ-stage production rate m (max )(day(-1)), and IJ-multiplication factor (C (IJ)/C (IJ,0))(max)(-). The variation of lambda (IJ) was not very important within fermentations (10.3-16.2 days); nonetheless, important effects were observed on m (max) (32.8-241.2 days(-1)) and (C (IJ)/C (IJ,0))(max) (66(-) to 611.4(-)). Particularly, maximum values of m (max) and (C (IJ)/C (IJ,0))(max) were obtained in medium A4 (0.276 l l(-1) agave juice, 17 g l(-1) yeast extract, 12 g l(-1) dried egg yolk, 0.025 l l(-1) corn oil). Also, the maximum IJ concentration (249,444 per ml) was achieved in A4-fermentations.     

Pristine but metal-rich Río Sucio (Dirty River) is dominated by Gallionella and other iron-sulfur oxidizing microbes

Whether the extreme conditions of acidity and heavy metal pollution of streams and rivers originating in pyritic formations are caused primarily by mining activities or by natural activities of metal-oxidizing microbes living within the geological formations is a subject of considerable controversy. Most microbiological studies of such waters have so far focused on acid mine drainage sites, which are heavily human-impacted environments, so it has been problematic to eliminate the human factor in the question of the origin of the key metal compounds. We have studied the physico-chemistry and microbiology of the Río Sucio in the Braulio Carrillo National Park of Costa Rica, 22 km from its volcanic rock origin. Neither the remote origin, nor the length of the river to the sampling site, have experienced human activity and are thus pristine. The river water had a characteristic brownish-yellow color due to high iron-dominated minerals, was slightly acidic, and rich in chemolithoautotrophic iron- and sulfur-oxidizing bacteria, dominated by Gallionella spp. Río Sucio is thus a natural acid-rock drainage system whose metal-containing components are derived primarily from microbial activities.