Multi-site-specific 16S rRNA methyltransferase RsmF from Thermus thermophilus

Cells devote a significant effort toward the production of multiple modified nucleotides in rRNAs, which fine tune the ribosome function. Here, we report that two methyltransferases, RsmB and RsmF, are responsible for all four 5-methylcytidine (m⁵C) modifications in 16S rRNA of Thermus thermophilus. Like Escherichia coli RsmB, T. thermophilus RsmB produces m⁵C967. In contrast to E. coli RsmF, which introduces a single m⁵C1407 modification, T. thermophilus RsmF modifies three positions, generating m⁵C1400 and m⁵C1404 in addition to m⁵C1407. These three residues are clustered near the decoding site of the ribosome, but are situated in distinct structural contexts, suggesting a requirement for flexibility in the RsmF active site that is absent from the E. coli enzyme. Two of these residues, C1400 and C1404, are sufficiently buried in the mature ribosome structure so as to require extensive unfolding of the rRNA to be accessible to RsmF. In vitro, T. thermophilus RsmF methylates C1400, C1404, and C1407 in a 30S subunit substrate, but only C1400 and C1404 when naked 16S rRNA is the substrate. The multispecificity of T. thermophilus RsmF is potentially explained by three crystal structures of the enzyme in a complex with cofactor S-adenosyl-methionine at up to 1.3 Å resolution. In addition to confirming the overall structural similarity to E. coli RsmF, these structures also reveal that key segments in the active site are likely to be dynamic in solution, thereby expanding substrate recognition by T. thermophilus RsmF.     

Changes in biodiversity of phytoplankton,zooplankton, fishes and macrobenthos in the Southern Caspian Sea after the invasion of the ctenophore <Emphasis Type="Italic">Mnemiopsis Leidyi</Emphasis>

Monitoring for 6 years (2001–2006) showed that the population explosion of the alien ctenophore Mnemiopsis leidyi in the southern Caspian Sea coincided with a decline in the abundance and species number of mesozooplankton. While this decline appeared to have reduced the nourishment of sprat (also known as kilka), it seemed to have affected phytoplankton favorably mainly due to the decrease in grazing pressure. During 2001–2002, when M. leidyi abundance and biomass were at their highest levels, abundance of dinoflagellates and cyanophytes exceeded that of diatoms. Before the invasion (1996) and in some years after the invasion (2003, 2004 and 2006) diatom abundance was higher than the abundance of other groups. In September 2005, an unprecedented bloom of the toxic cyanophyte Nodularia sp. was observed in the southern Caspian Sea. Disappearance of edible zooplankton such as Eurytemora spp. was among the first changes observed after the expansion of M. leidyi in the area. Some changes in the macrobenthic fauna were also conspicuous after the increase of this ctenophore. While the biomass of some deposit feeders, such as the polychaete Nereis diversicolor and oligochaete species increased, benthic crustaceans decreased sharply in abundance during 2001–2003 and completely disappeared during 2004–2006. Iranian catches of kilka, the most abundant and widespread zooplanktivorous fish, decreased significantly in the southern Caspian Sea after 1999. Iranian landings of kilka dropped ~70% from 69,070 ± 20,270 t during 1995–2000 to 23,430 ± 12,240 t during 2001–2006, resulting in a loss of at least 125 million US dollars to the economy. There were also changes in the total catches of large predators such as the kutum and mullet, which mainly feed on kilka, between 1991 and 2006.     

Cutting edge: Thymic NK cells develop independently from T cell precursors

Although NK cells in the mouse are thought to develop in the bone marrow, a small population of NK cells in the thymus has been shown to derive from a GATA3-dependent pathway. Characteristically, thymic NK cells express CD127 and few Ly49 molecules and lack CD11b. Because these NK cells develop in the thymus, the question of their relationship to the T cell lineage has been raised. Using several different mouse models, we find that unlike T cells, thymic NK cells are not the progeny of Rorc-expressing progenitors and do not express Rag2 or rearrange the TCRγ locus. We further demonstrate that thymic NK cells develop independently of the Notch signaling pathway, supporting the idea that thymic NK cells represent bona fide NK cells that can develop independently of all T cell precursors.     

Clustering of protein families into functional subtypes using Relative Complexity Measure with reduced amino acid alphabets

Background  

Phylogenetic analysis can be used to divide a protein family into subfamilies in the absence of experimental information. Most phylogenetic analysis methods utilize multiple alignment of sequences and are based on an evolutionary model. However, multiple alignment is not an automated procedure and requires human intervention to maintain alignment integrity and to produce phylogenies consistent with the functional splits in underlying sequences. To address this problem, we propose to use the alignment-free Relative Complexity Measure (RCM) combined with reduced amino acid alphabets to cluster protein families into functional subtypes purely on sequence criteria. Comparison with an alignment-based approach was also carried out to test the quality of the clustering.     

Salt tolerance in <Emphasis Type="Italic">Solanum pennellii</Emphasis>: antioxidant response and related QTL

Background  

Excessive soil salinity is an important problem for agriculture, however, salt tolerance is a complex trait that is not easily bred into plants. Exposure of cultivated tomato to salt stress has been reported to result in increased antioxidant content and activity. Salt tolerance of the related wild species, Solanum pennellii, has also been associated with similar changes in antioxidants. In this work, S. lycopersicum M82, S. pennellii LA716 and a S. pennellii introgression line (IL) population were evaluated for growth and their levels of antioxidant activity (total water-soluble antioxidant activity), major antioxidant compounds (phenolic and flavonoid contents) and antioxidant enzyme activities (superoxide dismutase, catalase, ascorbate peroxidase and peroxidase) under both control and salt stress (150 mM NaCl) conditions. These data were then used to identify quantitative trait loci (QTL) responsible for controlling the antioxidant parameters under both stress and nonstress conditions.     

Comparative study of effects of polyols, salts, and alcohols on trichloroacetic acid-induced state of cytochrome c

A systematic investigation of the effect of polyethylene glycols, salts, and alcohols on the trichloroacetic acid (TCA)-induced state of ferricytochrome c was made using various spectroscopic techniques. Native cytochrome c (Cyt c) has a fluorescence maximum at 335 nm, whereas the TCA-induced state of Cyt c has a red shift of 7 nm with enhanced fluorescence intensity. The near- and far-UV CD spectra showed a significant loss of tertiary and secondary structure, although the protein is relatively less unfolded as compared with a conformation at pH 2.0. Addition of 70% (v/v) polyols to TCA (3.3 mM)-induced state of Cyt c resulted in increased 1-anilino-8-naphthalene sulfonate binding and increased mean residue ellipticity at 222 nm, indicating increase in compactness with enhanced exposure of hydrophobic surface area. Also, the stabilizing effect of salts and alcohols on the TCA-induced state was studied and compared with their effect on trifluoroacetic acid-unfolded state of Cyt c. Among all the polyols, salts, and alcohols studied, PEG-400, K3[Fe(CN)6], and butanol were the most efficient in inducing secondary structure in TCA-induced state as examined by the above-mentioned spectroscopic techniques. For salts, the efficiency in inducing the secondary structure followed the order K3[Fe(CN)6] > KClO4 > K2SO4 > KCl. For alcohols, this order was found to be as follows: butanol > propanol > ethanol > methanol.     

Contents of metals in some wild mushrooms

The concentrations of 7 metals (lead, cadmium, manganese, copper, nickel, silver, and chromium) were determined in 32 different species of wild mushrooms. The mushroom samples, which have been using for food and some medical purposes, were collected from Konya, an Inner Anatolian region of Turkey. The highest metal concentrations were determined as 39 mg/kg Pb and 3.72 mg/kg Cd in Trichaptum abietinum, 467 mg/kg Mn in Panaeolus sphinctrinus, 326 mg/kg Cu in Trametes versicolor, 69.4 mg/kg Ni in Helvella spadicea, 6.97 mg/kg Ag in Agaricus campestris, and 84.5 mg/kg Cr in Phellinus igniarius. The maximum contents are 1.52, 2.22, and 60.2 mg/kg in Pleurotus eryngii (for Pb), Amanita vaginata (for Cd), and Helvella leucomelana (for Cu), respectively. These results were compared according to the WHO/FAO standard.     

Soft-electron beam and gamma-radiation sensitivity and DNA damage in phosphine-resistant and -susceptible strains of Rhyzopertha dominica

The soft-electron beam (low-energy electrons) and gamma-radiation sensitivities of phosphine-resistant (PHR) and -susceptible (PHS) strains of adults lesser grain borer Rhyzopertha dominica (F.) were studied, with particular reference to DNA damage assessed using single-cell electrophoresis (comet assay). Results showed that mortality in adult R. dominica varied significantly between both PHR and PHS strains. Adults of the PHR strain were found to be more tolerant toward soft-electron and gamma radiation than adults of the PHS strain. Studies on the longevity of strains showed that mean survival time and dose rate were highly correlated with both strains and treatments. Results also showed that adults of the PHR strain lived longer than adults of PHS strain for both treatments. Radiation sensitivity indices, however, decreased as radiation dose increased in both strains. Analysis of DNA damage, after 40- and 160-Gy gamma radiation, was carried out using cells obtained from both strains. Gamma-irradiated adults of both strains showed typical DNA fragmentation, compared with cells from nonirradiated adults, which showed more intact DNA. Investigations using the comet assay showed that tail length, moment, olive-tail moment, percentage of tail DNA, and percentage of DNA damage were all greater in the PHS strain compared with the PHR strain and the control insects. Results also showed that DNA damage remained at a constant level for up to 24 h after irradiation. The results have been discussed in relation to the observed strain differences in radiation sensitivity and resistance to phosphine.     

Cloning and functional characterization of the ambler class C beta-lactamase of Yersinia ruckeri

Yersinia ruckeri is a gram-negative pathogen causing enteric redmouth disease in salmonids. Previous studies have reported that Y. ruckeri harbors an ampC gene that is expressed at low level. In this present work, the entire ampC gene of Y. ruckeri was cloned and expressed in Escherichia coli. The AmpC enzyme confers resistance to aminopenicillins and narrow-spectrum cephalosporins, which fit well with the kinetic properties of the purified enzyme. Phylogenetic analysis showed that YRC-1 did not share significant sequence identity with known plasmid-mediated or chromosomal AmpC enzymes. This work provides further evidence that fish-pathogenic gram-negative rod species may constitute a reservoir of antibiotic resistance genes.