PS II model based analysis of transient fluorescence yield measured on whole leaves of Arabidopsis thaliana after excitation with light flashes of different energies

Our recently presented PS II model (Belyaeva et al., 2008) was improved in order to permit a consistent simulation of Single Flash Induced Transient Fluorescence Yield (SFITFY) traces that were earlier measured by Steffen et al. (2005) on whole leaves of Arabidopsis (A.) thaliana at four different energies of the actinic flash. As the essential modification, the shape of the actinic flash was explicitly taken into account assuming that an exponentially decaying rate simulates the time dependent excitation of PS II by the 10 ns actinic flash. The maximum amplitude of this excitation exceeds that of the measuring light by 9 orders of magnitude. A very good fit of the SFITFY data was achieved in the time domain from 100 ns to 10 s for all actinic flash energies (the maximum energy of 7.5 × 10¹⁶ photons/(cm² flash) is set to 100%, the relative energies of weaker actinic flashes were of ∼8%, 4%, ∼1%). Our model allows the calculation and visualization of the transient PS II redox state populations ranging from the dark adapted state, via excitation energy and electron transfer steps induced by pulse excitation, followed by final relaxation into the stationary state eventually attained under the measuring light. It turned out that the rate constants of electron transfer steps are invariant to intensity of the actinic laser flash. In marked contrast, an increase of the actinic flash energy by more than two orders of magnitude from 5.4 × 10¹⁴ photons/(cm² flash) to 7.5 × 10¹⁶ photons/(cm² flash), leads to an increase of the extent of fluorescence quenching due to carotenoid triplet (³Car) formation by a factor of 14 and of the recombination reaction between reduced primary pheophytin (Phe⁻) and P680⁺ by a factor of 3 while the heat dissipation in the antenna complex remains virtually constant.The modified PS II model offers new opportunities to compare electron transfer and dissipative parameters for different species (e.g. for the green algae and the higher plant) under varying illumination conditions.     

Cysteine S-glycosylation, a new post-translational modification found in glycopeptide bacteriocins

O-glycosylation is a ubiquitous eukaryotic post-translational modification, whereas early reports of S-linked glycopeptides have never been verified. Prokaryotes also glycosylate proteins, but there are no confirmed examples of sidechain glycosylation in ribosomal antimicrobial polypeptides collectively known as bacteriocins. Here we show that glycocin F, a bacteriocin secreted by Lactobacillus plantarum KW30, is modified by an N-acetylglucosamine β-O-linked to Ser18, and an N-acetylhexosamine S-linked to C-terminal Cys43. The O-linked N-acetylglucosamine is essential for bacteriostatic activity, and the C-terminus is required for full potency (IC₅₀ 2 nM). Genomic context analysis identified diverse putative glycopeptide bacteriocins in Firmicutes. One of these, the reputed lantibiotic sublancin, was shown to contain a hexose S-linked to Cys22.     

Insights into metalloregulation by M-box riboswitch RNAs via structural analysis of manganese-bound complexes

The M-box riboswitch couples intracellular magnesium levels to expression of bacterial metal transport genes. Structural analyses on other riboswitch RNA classes, which typically respond to a small organic metabolite, have revealed that ligand recognition occurs through a combination of base-stacking, electrostatic, and hydrogen-bonding interactions. In contrast, the M-box RNA triggers a change in gene expression upon association with an undefined population of metals, rather than responding to only a single ligand. Prior biophysical experimentation suggested that divalent ions associate with the M-box RNA to promote a compacted tertiary conformation, resulting in sequestration of a short sequence tract otherwise required for downstream gene expression. Electrostatic shielding from loosely associated metals is undoubtedly an important influence during this metal-mediated compaction pathway. However, it is also likely that a subset of divalent ions specifically occupies cation binding sites and promotes proper positioning of functional groups for tertiary structure stabilization. To better elucidate the role of these metal binding sites, we resolved a manganese-chelated M-box RNA complex to 1.86 Å by X-ray crystallography. These data support the presence of at least eight well-ordered cation binding pockets, including several sites that had been predicted by biochemical studies but were not observed in prior structural analysis. Overall, these data support the presence of three metal-binding cores within the M-box RNA that facilitate a network of long-range interactions within the metal-bound, compacted conformation.     

Plasmodium sporozoite disintegration during skin passage limits malaria parasite transmission

During transmission of malaria‐causing parasites from mosquitoes to mammals, Plasmodium sporozoites migrate rapidly in the skin to search for a blood vessel. The high migratory speed and narrow passages taken by the parasites suggest considerable strain on the sporozoites to maintain their shape. Here, we show that the membrane‐associated protein, concavin, is important for the maintenance of the Plasmodium sporozoite shape inside salivary glands of mosquitoes and during migration in the skin. Concavin‐GFP localizes at the cytoplasmic periphery and concavin(−) sporozoites progressively round up upon entry of salivary glands. Rounded concavin(−) sporozoites fail to pass through the narrow salivary ducts and are rarely ejected by mosquitoes, while normally shaped concavin(−) sporozoites are transmitted. Strikingly, motile concavin(−) sporozoites disintegrate while migrating through the skin leading to parasite arrest or death and decreased transmission efficiency. Collectively, we suggest that concavin contributes to cell shape maintenance by riveting the plasma membrane to the subtending inner membrane complex. Interfering with cell shape maintenance pathways might hence provide a new strategy to prevent a malaria infection.     

A likelihood-based approach to estimating and testing for isolation by distance

Simple regression of genetic similarities between pairs of populations on their corresponding geographic distances is frequently used to detect the presence of isolation by distance (IBD). However, these pairwise values are obviously not independent and there is no parametric procedure for estimating and testing for the IBD intercepts and slopes based on standard regression theory. Nonparametric tests, such as the Mantel test, and resampling techniques, such as bootstrapping, have been exploited with limited success. Here, I describe a likelihood-based analysis to allow for simultaneously detecting patterns of correlated residuals and estimating and testing for the presence of IBD. It is shown, through the analysis of two molecular datasets in pine species, that different covariance structures of the residuals exist. More over, the likelihood ratio tests under these covariance structures are less sensitive to the presence of IBD than the Mantel test and the simple regression analysis but more sensitive than the bootstrap and jackknife samples over independent populations or population pairs. Because the likelihood analysis directly models and accounts for nonindependence of residuals, it should legitimately detect the presence of IBD, thereby allowing for accurate inferences about evolutionary and demographic processes influencing the extent and patterns of IBD.     

Population biology of the little gulper shark Centrophorus uyato in Lebanese waters

A total of 38 individuals of the heavily exploited little gulper shark Centrophorus uyato were collected from Lebanese coastal waters using bottom longlines and trammel nets of different meshes at depths ranging from 115 to 600 m between May 2013 and February 2014. Their total lengths were between 45 and 94 cm and their total mass was from 870 to 6700 g. The sex ratio was not significantly different from 1:1, with 20 males and 18 females, but bathymetric sexual segregation occurred. Catch per net setting (CNS) was used as a relative abundance index to detect spatial distribution; trammel nets showed largest CNS ranging from 4·9 to 5·45 kg per unit effort in the north and south, at depths from 120 to 200 m, during spring and summer. The mass–length relationships demonstrated negative allometric growth (b < 3) (males: M_T = 0.3585L_T^2·071, r² = 0·94; females: M_T = 0.0239L_T^2·735, r² = 0·64). The condition factor as well as the gonado‐somatic and hepato‐somatic indices of C. uyato in the study area were also calculated. The results are discussed in relation to the distribution, growth and reproduction as well as the management of C. uyato.     

The combined use of raw and phylogenetically independent methods of outlier detection uncovers genome‐wide dynamics of local adaptation in a lizard

Local adaptation is a dynamic process by which different allele combinations are selected in different populations at different times, and whose genetic signature can be inferred by genome‐wide outlier analyses. We combined gene flow estimates with two methods of outlier detection, one of them independent of population coancestry (CIOA) and the other one not (ROA), to identify genetic variants favored when ecology promotes phenotypic convergence. We analyzed genotyping‐by‐sequencing data from five populations of a lizard distributed over an environmentally heterogeneous range that has been changing since the split of eastern and western lineages ca. 3 mya. Overall, western lizards inhabit forest habitat and are unstriped, whereas eastern ones inhabit shrublands and are striped. However, one population (Lerma) has unstriped phenotype despite its eastern ancestry. The analysis of 73,291 SNPs confirmed the east–west division and identified nonoverlapping sets of outliers (12 identified by ROA and 9 by CIOA). ROA revealed ancestral adaptive variation in the uncovered outliers that were subject to divergent selection and differently fixed for eastern and western populations at the extremes of the environmental gradient. Interestingly, such variation was maintained in Lerma, where we found high levels of heterozygosity for ROA outliers, whereas CIOA uncovered innovative variants that were selected only there. Overall, it seems that both the maintenance of ancestral variation and asymmetric migration have counterbalanced adaptive lineage splitting in our model species. This scenario, which is likely promoted by a changing and heterogeneous environment, could hamper ecological speciation of locally adapted populations despite strong genetic structure between lineages.     

Direct Detection of Membrane-Inserting Fragments Defines the Translocation Pores of a Family of Pathogenic Toxins

Large clostridial toxins (LCTs) are a family of homologous proteins toxins that are directly responsible for the symptoms associated with a number of clostridial infections that cause disease in humans and in other animals. LCTs damage tissues by delivering a glucosyltransferase domain, which inactivates small GTPases, across the endosomal membrane and into the cytosol of target cells. Elucidating the mechanism of translocation for LCTs has been hampered by difficulties associated with identifying marginally hydrophobic segments that insert into the bounding membrane to form the translocation pore. Here, we directly measured the membrane-insertion partitioning propensity for segments spanning the putative pore-forming region using a translocon-mediated insertion assay and synthetic peptides. We identified membrane-inserting segments, as well as a conserved and functionally important negatively charged residue that requires protonation for efficient membrane insertion. We provide a model of the LCT pore, which provides insights into translocation for this enigmatic family of α-helical translocases.