Brown midrib2 (Bmr2) encodes the major 4-coumarate:coenzyme A ligase involved in lignin biosynthesis in sorghum (Sorghum bicolor (L.) Moench)

Successful modification of plant cell-wall composition without compromising plant integrity is dependent on being able to modify the expression of specific genes, but this can be very challenging when the target genes are members of multigene families. 4-coumarate:CoA ligase (4CL) catalyzes the formation of 4-coumaroyl CoA, a precursor of both flavonoids and monolignols, and is an attractive target for transgenic down-regulation aimed at improving agro-industrial properties. Inconsistent phenotypes of transgenic plants have been attributed to variable levels of down-regulation of multiple 4CL genes. Phylogenetic analysis of the sorghum genome revealed 24 4CL(-like) proteins, five of which cluster with bona fide 4CLs from other species. Using a map-based cloning approach and analysis of two independent mutant alleles, the sorghum brown midrib2 (bmr2) locus was shown to encode 4CL. In vitro enzyme assays indicated that its preferred substrate is 4-coumarate. Missense mutations in the two bmr2 alleles result in loss of 4CL activity, probably as a result of improper folding as indicated by molecular modeling. Bmr2 is the most highly expressed 4CL in sorghum stems, leaves and roots, both at the seedling stage and in pre-flowering plants, but the products of several paralogs also display 4CL activity and compensate for some of the lost activity. The contribution of the paralogs varies between developmental stages and tissues. Gene expression assays indicated that Bmr2 is under auto-regulatory control, as reduced 4CL activity results in over-expression of the defective gene. Several 4CL paralogs are also up-regulated in response to the mutation.     

1H, 13C, 15N and 31P chemical shift assignments of a human Xist RNA A-repeat tetraloop hairpin essential for X-chromosome inactivation

Initiation of X-chromosome inactivation in female mammals depends on the non-coding RNA Xist. We have solved the NMR structure of a 14-nucleotide hairpin with a novel AUCG tetraloop fold from a Xist A-repeat that is essential for silencing. The ¹H, ¹³C, ¹⁵N and ³¹P chemical shift assignments are reported.     

Can electromagnetic fields influence the structure and enzymatic digest of proteins? A critical evaluation of microwave-assisted proteomics protocols

This study reevaluates the putative advantages of microwave-assisted tryptic digests compared to conventionally heated protocols performed at the same temperature. An initial investigation of enzyme stability in a temperature range of 37-80°C demonstrated that trypsin activity declines sharply at temperatures above 60°C, regardless if microwave dielectric heating or conventional heating is employed. Tryptic digests of three proteins of different size (bovine serum albumin, cytochrome c and β-casein) were thus performed at 37°C and 50°C using both microwave and conventional heating applying accurate internal fiber-optic probe reaction temperature measurements. The impact of the heating method on protein degradation and peptide fragment generation was analyzed by SDS-PAGE and MALDI-TOF-MS. Time-dependent tryptic digestion of the three proteins and subsequent analysis of the corresponding cleavage products by MALDI-TOF provided virtually identical results for both microwave and conventional heating. In addition, the impact of electromagnetic field strength on the tertiary structure of trypsin and BSA was evaluated by molecular mechanics calculations. These simulations revealed that the applied field in a typical laboratory microwave reactor is 3-4 orders of magnitude too low to induce conformational changes in proteins or enzymes.     

Temporal Variation in Seed Predation by Insects in a Population of Syagrus romanzoffiana (Arecaceae) in Santa Catarina Island, SC, Brazil

Insect seed predation may vary depending on seed production. The present study considers the hypothesis that the rates of seed predation tend to be smaller in years of higher fruit production. Thus, we monitored the production of fruits and predation of seeds of the palm Syagrus romanzoffiana over 2?years in the Atlantic Forest (Parque Municipal da Lagoa do Peri, Florianópolis, SC, Brazil), between July 2006 and June 2008. Plots of 0.25?m² were fitted under 20 mother plants and fruits were monthly collected for assessment of abundance and seed predation. There was variation in fruit production between the 2?years and among reproductive plants. Predation rates were high and occurred in the predispersal phase by the Curculionidae Revena rubiginosa Boheman, Anchylorhynchus aegrotus Fahraeus, and Anchylorhynchus variabilis Gyllenhal. Seed predation by these species of Anchylorhynchus is first registered in the present study. In average, about 60% of the seeds monthly produced in the population tend to escape insect predation in year of high or low production, becoming available for recruitment. The predation rate was not related to the amount of fruits produced per reproductive plant. Also, different than expected, there was a positive relation between the rates of seed predation and the total of fruits produced monthly on the plots. Thus, no evidence for the satiation of insect seed predators was found in this study with S. romanzoffiana.     

Occurrence and assemblage composition of millipedes (Myriapoda, Diplopoda) and terrestrial isopods (Crustacea, Isopoda, Oniscidea) in urban areas of Switzerland

Terrestrial isopods and millipedes, members of the invertebrate macro-decomposer guild, were collected through pitfall traps in three Swiss cities (Zurich, Lucerne, Lugano). A total of 7,198 individuals of 17 isopod species (7093 ind.), and 10 millipede species (105 ind.) were captured. Besides the Alpine endemic isopod (Trichoniscus alemannicus) and millipede (Cylindroiulus verhoeffi), urban assemblages were mainly composed of widespread, native European and even cosmopolitan species, which are frequent in anthropogenic areas. Overall species richness (isopods and millipedes combined) was similar in Zurich (17 species) and Lucerne (16), while only 13 species were sampled in Lugano. According to the Sørensen index of similarity, species composition of Zurich and Lucerne were more alike, while the one of Lugano was more distinct from the other two cities. This result can be explained by the spatial proximity of Zurich and Lucerne in the north of the Alps compared to Lugano, which is located more distantly and in the south of the Alps. Dominant isopods and millipedes in Zurich and Lucerne were found to be widespread synanthropic species in temperate Europe(Porcellio scaber, Trachelipus rathkii and Ophyiulus pilosus) while the dominant isopod in Lugano (Trachelipus razzautii) is a species with a north-eastern Mediterranean distribution. Our study reveals that the urban millipede and isopod fauna in Swiss cities mainly consists of widespread species, but species of narrower distribution (e.g. Trichoniscus alemannicus, Cylindroiulus verhoeffi) may also find suitable habitats in cities. Despite some signs of biotic homogenization, our study also found compositional differences of millipede and isopod assemblages between northern and southern cities that suggest geographical effects of the regional species pool.     

Phylogenetic and Functional Heterogeneity of Sediment Biofilms along Environmental Gradients in a Glacial Stream

We used in situ hybridization with fluorescently labeled rRNA-targeted oligonucleotide probes concurrently with measurements of bacterial carbon production, biomass, and extracellular polymeric substances (EPS) to describe the bacterial community in sediments along a glacial stream. The abundance of sediment-associated Archaea, as detected with the ARCH915 probe, decreased downstream of the glacier snout, and a major storm increased their relative abundance by a factor of 5.5 to 7.9. Bacteria of the Cytophaga-Flavobacterium group were also sixfold to eightfold more abundant in the storm aftermath. Furthermore, elevated numbers of Archaea and members of the Cytophaga-Flavobacterium group characterized the phylogenetic composition of the supraglacial ice community. We postulate that glacial meltwaters constitute a possible source of allochthonous bacteria to the stream biofilms. Although stream water temperature increased dramatically from the glacier snout along the stream (3.5 km), sediment chlorophyll a was the best predictor for bacterial carbon production and specific growth rates along the stream. Concomitant with an increase in sediment chlorophyll a, the EPS carbohydrate-to-bacterial-cell ratio declined 11- to 15-fold along the stream prior to the storm, which is indicative of a larger biofilm matrix in upstream reaches. We assume that a larger biofilm matrix is required to assure prolonged transient storage and enzymatic processing of allochthonous macromolecules, which are likely the major substrate for microbial heterotrophs. Bacteria of the Cytophaga-Flavobacterium cluster, which are well known to degrade complex macromolecules, were most abundant in these stream reaches. Downstream, higher algal biomass continuously supplies heterotrophs with easily available exudates, therefore making a larger matrix unnecessary. As a result, bacterial carbon production and specific growth rates were higher in downstream reaches.     

Hypochlorite-modified (lipo)proteins are present in rabbit lesions in response to dietary cholesterol.

Myeloperoxidase (MPO), a heme enzyme secreted by activated phagocytes, generates an array of oxidants proposed to play critical roles in host defense, tissues damage, and foam cell formation. Although neutrophils are the major source for MPO, the enzyme could be identified abundantly in circulating monocytes and monocytes/macrophages in rabbit lesions. MPO is the only enzyme known to generate hypochlorous acid (HOCl) and HOCl-modified lipoproteins have pronounced atherogenic and/or proinflammatory features in vivo and in vitro. Using specific monoclonal antibodies, HOCl-modified (lipo)proteins were detected in atherosclerotic plaques of heterozygous Watanabe heritable hyperlipidemic rabbits and to a lesser extent in a specific strain of New Zealand White rabbits with a high atherosclerotic response to hypercholesterolemia. Colocalization of immunoreactive MPO and HOCl-modified-epitopes in serial sections of rabbit lesions provides convincing evidence for MPO-H2O2-chloride system-mediated oxidation of (lipo)proteins under in vivo conditions. We propose that monocyte-derived MPO could connect chronic inflammatory conditions with arterial lipid/lipoprotein deposition during diet-induced atherogenesis in rabbits.