Recombinant <Emphasis Type="Italic">Escherichia coli</Emphasis> produces tailor-made biopolyester granules for applications in fluorescence activated cell sorting: functional display of the mouse interleukin-2 and myelin oligodendrocyte glycoprotein

Background  

Fluorescence activated cell sorting (FACS) is a powerful technique for the qualitative and quantitative detection of biomolecules used widely in both basic research and clinical diagnostic applications. Beads displaying a specific antigen are used to bind antibodies which are then fluorescently labelled using secondary antibodies. As the individual suspension bead passes through the sensing region of the FACS machine, fluorescent signals are acquired and analysed. Currently, antigens are tediously purified and chemically cross-linked to preformed beads. Purification and coupling of proteins often renders them inactive and they will not be displayed in its native configuration. As an alternative, we genetically engineered Escherichia coli to produce biopolyester (polyhdroxyalkanoate=PHA) granules displaying diagnostically relevant antigens in their native conformation and suitable for FACS analysis.     

MicroRNAs show a wide diversity of expression profiles in the developing and mature central nervous system

Background  

MicroRNA (miRNA) encoding genes are abundant in vertebrate genomes but very few have been studied in any detail. Bioinformatic tools allow prediction of miRNA targets and this information coupled with knowledge of miRNA expression profiles facilitates formulation of hypotheses of miRNA function. Although the central nervous system (CNS) is a prominent site of miRNA expression, virtually nothing is known about the spatial and temporal expression profiles of miRNAs in the brain. To provide an overview of the breadth of miRNA expression in the CNS, we performed a comprehensive analysis of the neuroanatomical expression profiles of 38 abundant conserved miRNAs in developing and adult zebrafish brain.     

High molecular weight dissolved organic matter enrichment selects for methylotrophs in dilution to extinction cultures

The role of bacterioplankton in the cycling of marine dissolved organic matter (DOM) is central to the carbon and energy balance in the ocean, yet there are few model organisms available to investigate the genes, metabolic pathways, and biochemical mechanisms involved in the degradation of this globally important carbon pool. To obtain microbial isolates capable of degrading semi-labile DOM for growth, we conducted dilution to extinction cultivation experiments using seawater enriched with high molecular weight (HMW) DOM. In total, 93 isolates were obtained. Amendments using HMW DOM to increase the dissolved organic carbon concentration 4x (280 μM) or 10x (700 μM) the ocean surface water concentrations yielded positive growth in 4–6% of replicate dilutions, whereas <1% scored positive for growth in non-DOM-amended controls. The majority (71%) of isolates displayed a distinct increase in cell yields when grown in increasing concentrations of HMW DOM. Whole-genome sequencing was used to screen the culture collection for purity and to determine the phylogenetic identity of the isolates. Eleven percent of the isolates belonged to the gammaproteobacteria including Alteromonadales (the SAR92 clade) and Vibrio. Surprisingly, 85% of isolates belonged to the methylotrophic OM43 clade of betaproteobacteria, bacteria thought to metabolically specialize in degrading C1 compounds. Growth of these isolates on methanol confirmed their methylotrophic phenotype. Our results indicate that dilution to extinction cultivation enriched with natural sources of organic substrates has a potential to reveal the previously unsuspected relationships between naturally occurring organic nutrients and the microorganisms that consume them.     

Rac signaling in breast cancer: a tale of GEFs and GAPs

Rac GTPases, small G-proteins widely implicated in tumorigenesis and metastasis, transduce signals from tyrosine-kinase, G-protein-coupled receptors (GPCRs), and integrins, and control a number of essential cellular functions including motility, adhesion, and proliferation. Deregulation of Rac signaling in cancer is generally a consequence of enhanced upstream inputs from tyrosine-kinase receptors, PI3K or Guanine nucleotide Exchange Factors (GEFs), or reduced Rac inactivation by GTPase Activating Proteins (GAPs). In breast cancer cells Rac1 is a downstream effector of ErbB receptors and mediates migratory responses by ErbB1/EGFR ligands such as EGF or TGFα and ErbB3 ligands such as heregulins. Recent advances in the field led to the identification of the Rac-GEF P-Rex1 as an essential mediator of Rac1 responses in breast cancer cells. P-Rex1 is activated by the PI3K product PIP3 and Gβγ subunits, and integrates signals from ErbB receptors and GPCRs. Most notably, P-Rex1 is highly overexpressed in human luminal breast tumors, particularly those expressing ErbB2 and estrogen receptor (ER). The P-Rex1/Rac signaling pathway may represent an attractive target for breast cancer therapy.     

Comparative genomic evidence for a close relationship between the dimorphic prosthecate bacteria Hyphomonas neptunium and Caulobacter crescentus

The dimorphic prosthecate bacteria (DPB) are alpha-proteobacteria that reproduce in an asymmetric manner rather than by binary fission and are of interest as simple models of development. Prior to this work, the only member of this group for which genome sequence was available was the model freshwater organism Caulobacter crescentus. Here we describe the genome sequence of Hyphomonas neptunium, a marine member of the DPB that differs from C. crescentus in that H. neptunium uses its stalk as a reproductive structure. Genome analysis indicates that this organism shares more genes with C. crescentus than it does with Silicibacter pomeroyi (a closer relative according to 16S rRNA phylogeny), that it relies upon a heterotrophic strategy utilizing a wide range of substrates, that its cell cycle is likely to be regulated in a similar manner to that of C. crescentus, and that the outer membrane complements of H. neptunium and C. crescentus are remarkably similar. H. neptunium swarmer cells are highly motile via a single polar flagellum. With the exception of cheY and cheR, genes required for chemotaxis were absent in the H. neptunium genome. Consistent with this observation, H. neptunium swarmer cells did not respond to any chemotactic stimuli that were tested, which suggests that H. neptunium motility is a random dispersal mechanism for swarmer cells rather than a stimulus-controlled navigation system for locating specific environments. In addition to providing insights into bacterial development, the H. neptunium genome will provide an important resource for the study of other interesting biological processes including chromosome segregation, polar growth, and cell aging.     

Effects of melatonin and undernutrition on the viability of ovine embryos during anestrus and the breeding season

This study examined the effects of melatonin and level of nutrition on embryo yield during anestrous and breeding season. Adult Rasa Aragonesa ewes were assigned randomly to one of the four treatment groups in two experiments using a 2x2x2 factorial design. Individuals were treated (+MEL) or not treated (-MEL) with a subcutaneous implant of melatonin for 42d (Melovine, CEVA) and fed 1.5 (control, C) or 0.5 (low, L) times the daily maintenance requirements for 20d. Ewes were mated at oestrus (Day=0) and embryos were recovered on Day 5. Level of nutrition and melatonin supplements did not have a significant effect on ovulation rate or the number of recovered ova per ewe in the Reproductive Season (RS) and the Anestrous Season (AS). During the RS, undernutrition reduced the number of viable embryos per ewe (C: 1.1+/-0.2; L: 0.6+/-0.2; P<0.05); however, the number of viable embryos per ewe in the L+MEL group (0.2+/-0.15) was significantly lower than it was in the L, C+MEL and C groups (0.9+/-0.3, 1.2+/-0.3, 1.0+/-0.4, respectively; P<0.05). In the AS, nutrition did not have a significant effect on the number of viable embryos per ewe, although melatonin supplements might have improved rates slightly. Embryo viability rate (% viable embryos/embryos recovered) was unaffected by melatonin supplements or level of nutrition in the RS and the AS. Season had a strong effect on the number of viable embryos per functional corpus luteum among ewes in the L+MEL group, only (RS: 0.2+/-0.1; AS: 0.6+/-0.2; P<0.05). In conclusion, undernutrition impaired the viability of sheep embryos in the RS, particularly among ewes that were given melatonin supplements subcutaneously, but melatonin appeared to improve embryo quality in the AS, which suggests that the mechanisms involved in the interactive effects of melatonin and nutrition on embryo development are influenced by season.     

Specialization clines in the pollination systems of agaves (Agavaceae) and columnar cacti (Cactaceae): A phylogenetically controlled meta-analysis

The biogeography of plant-animal interactions is a novel topic on which many disciplines converge (e.g., reproductive biology, biogeography, and evolutionary biology). Narrative reviews have indicated that tropical columnar cacti and agaves have highly specialized pollination systems, while extratropical species have generalized systems. However, this dichotomy has never been quantitatively tested. We tested this hypothesis using traditional and phylogenetically informed meta-analysis. Three effect sizes were estimated from the literature: diurnal, nocturnal, and hand cross-pollination (an indicator of pollen limitation). Columnar cactus pollination systems ranged from purely bat-pollinated in the tropics to generalized pollination, with diurnal visitors as effective as nocturnal visitors in extratropical regions; even when phylogenetic relatedness among species is taken into account. Metaregressions identified a latitudinal increase in pollen limitation in columnar cacti, but this increase was not significant after correcting for phylogeny. The currently available data for agaves do not support any latitudinal trend. Nectar production of columnar cacti varied with latitude. Although this variation is positively correlated with pollination by diurnal visitors, it is influenced by phylogeny. The degree of specificity in the pollination systems of columnar cacti is heavily influenced by ecological factors and has a predictable geographic pattern.     

2009 Society for Medical Anthropology Conference: Reflections on the Future of Anthropology

In his plenary session entitled Five Questions on the Future, Harvard anthropologist Arthur Kleinman capitalized on the 2009 Society for Medical Anthropology Conference’s theme of Medical Anthropology at the Intersections to speculate on the future of the discipline.As he reflects on the field of anthropology, which had lacked theory, ethnography, and strong ties to public health and medicine, Harvard anthropologist Arthur Kleinman celebrates the accomplishments made by his contemporaries by saying, “My generation has made medical anthropology what it is today.” However, he is now looking to the future of the discipline, saying it must re-examine itself as a field.During the 2009 Society for Medical Anthropology Conference at Yale University, Kleinman capitalized on the theme of Medical Anthropology at the Intersections in his plenary session entitled Five Questions on the Future. Casting the conference itself as a kind of intersection, Kleinman not only lauded its size and diversity, but asserted that it marked a pivotal moment in which medical anthropology must re-evaluate its central questions.     

Availability of and access to critical habitats in regulated rivers: effects of low-head barriers on threatened lampreys

1.  Conservation of freshwater animal populations requires their access to, as well as sufficient availability of, critical habitats, such as those for reproduction. Abundant small-scale barriers may cause extensive fragmentation of freshwater habitat but, by comparison to larger structures their effects are rarely considered by catchment managers. The relationship between the distribution of, and access to, spawning habitat in a regulated river, characterized by abundant small barriers, was examined for river lamprey Lampetra fluviatilis , a threatened migratory fish.
2.  Telemetry of adult lamprey in the River Derwent, North East England was used to quantify upriver migration and access to spawning habitat, together with surveys of spawning habitat availability and spawning activity between 2002 and 2007.
3.  Access in to the Derwent appeared severely restricted by a tidal barrage, beyond which lamprey migrated rapidly in unobstructed reaches. Of all lamprey tagged in the lower 4 km of river, or ascending the barrage, 64% and 17% passed the first and second weirs respectively, with high flows crucial for this. Although over 98% of lamprey spawning habitat occurred more than 51 km upstream, on average just 1.8% of river lamprey spawners were recorded there.
4.  In order to protect or rehabilitate species or species assemblages, greater attention needs to be paid to the relative spatial distribution of low-head barriers and the resultant availability of key habitats within individual catchments. This is particularly important given the renewed emphasis internationally on low-head hydropower solutions as a source of renewable energy, and the rapid growth in numbers of low-head barriers in many catchments.     

Chemopreventive and renal protective effects for docosahexaenoic acid (DHA): implications of CRP and lipid peroxides

Background

The fish oil-derived ω-3 fatty acids, like docosahexanoic (DHA), claim a plethora of health benefits. We currently evaluated the antitumor effects of DHA, alone or in combination with cisplatin (CP) in the EAC solid tumor mice model, and monitored concomitant changes in serum levels of C-reactive protein (CRP), lipid peroxidation (measured as malondialdehyde; MDA) and leukocytic count (LC). Further, we verified the capacity of DHA to ameliorate the lethal, CP-induced nephrotoxicity in rats and the molecular mechanisms involved therein.

Results

EAC-bearing mice exhibited markedly elevated LC (2-fold), CRP (11-fold) and MDA levels (2.7-fold). DHA (125, 250 mg/kg) elicited significant, dose-dependent reductions in tumor size (38%, 79%; respectively), as well as in LC, CRP and MDA levels. These effects for CP were appreciably lower than those of DHA (250 mg/kg). Interestingly, DHA (125 mg/kg) markedly enhanced the chemopreventive effects of CP and boosted its ability to reduce serum CRP and MDA levels. Correlation studies revealed a high degree of positive association between tumor growth and each of CRP (r = 0.85) and leukocytosis (r = 0.89), thus attesting to a diagnostic/prognostic role for CRP. On the other hand, a single CP dose (10 mg/kg) induced nephrotoxicity in rats that was evidenced by proteinuria, deterioration of glomerular filtration rate (GFR, -4-fold), a rise in serum creatinine/urea levels (2–5-fold) after 4 days, and globally-induced animal fatalities after 7 days. Kidney-homogenates from CP-treated rats displayed significantly elevated MDA- and TNF-α-, but reduced GSH-, levels. Rats treated with DHA (250 mg/kg, but not 125 mg/kg) survived the lethal effects of CP, and showed a significant recovery of GFR; while their homogenates had markedly-reduced MDA- and TNF-α-, but -increased GSH-levels. Significant association was detected between creatinine level and those of MDA (r = 0.81), TNF-α ) r = 0.92) and GSH (r = -0.82); implying causal relationships.

Conclusion

DHA elicited prominent chemopreventive effects on its own, and appreciably augmented those of CP as well. The extent of tumor progression in various mouse groups was highly reflected by CRP levels (thus implying a diagnostic/prognostic role for CRP). Further, this study is the first to reveal that DHA can obliterate the lethal CP-induced nephrotoxicity and renal tissue injury. At the molecular level, DHA appears to act by reducing leukocytosis, systemic inflammation, and oxidative stress.