Massively parallel tag sequencing reveals the complexity of anaerobic marine protistan communities

Background  

Recent advances in sequencing strategies make possible unprecedented depth and scale of sampling for molecular detection of microbial diversity. Two major paradigm-shifting discoveries include the detection of bacterial diversity that is one to two orders of magnitude greater than previous estimates, and the discovery of an exciting 'rare biosphere' of molecular signatures ('species') of poorly understood ecological significance. We applied a high-throughput parallel tag sequencing (454 sequencing) protocol adopted for eukaryotes to investigate protistan community complexity in two contrasting anoxic marine ecosystems (Framvaren Fjord, Norway; Cariaco deep-sea basin, Venezuela). Both sampling sites have previously been scrutinized for protistan diversity by traditional clone library construction and Sanger sequencing. By comparing these clone library data with 454 amplicon library data, we assess the efficiency of high-throughput tag sequencing strategies. We here present a novel, highly conservative bioinformatic analysis pipeline for the processing of large tag sequence data sets.     

Density-dependent regulation in populations of potato-colonizing aphids

Scarcity of long-term (over 30 years) data series represents a major challenge for an accurate estimation of the role of density-dependent processes in population regulation. We analyzed population densities of the wingless parthenogenic morphs of buckthorn aphid (BA), Aphis nasturtii Kaltenbach, potato aphid (PA), Macrosiphum euphorbiae (Thomas), and green peach aphid (GPA), Myzus persicae (Sulzer) from 1949 to 2003 for signs of density-dependent regulation. We found strong evidence of density-dependent regulation, with detection of density dependence being fairly consistent among the different statistical techniques. Direct density dependence was detected for the populations of all three species. There was also evidence of delayed density dependence for PA. The periodicity of population fluctuations for BA and GPA was 6.1 years and 3.9 years, respectively. The periodicity for PA was not explicit, being highly variable throughout the time series. Effects of density-independent weather factors were relatively minor compared to density-dependent regulation. The BA populations experienced a significant reduction in both density and annual oscillations starting in 1995, while GPA populations experienced a similar reduction in 1991. No such change was apparent for PA. The most likely explanation for the observed phenomenon is a change in the composition of the lady beetle community following the establishment of two alien coccinellid species, and/or changes in insecticide use by commercial growers in the area of the study.     

Spatial Distribution and Molecular Diversity of Archaeal Communities in the Extreme Hypersaline Meromictic Brâncoveanu Lake (Transylvanian Basin,Romania)

Dating from the Middle Miocene, the massive halite deposits lying beneath the Transylvanian Basin (Central Romania) have been valuable mineral resources quarried for millennia. Among the numerous hypersaline pit lakes that resulted from this mining, Brâncoveanu Lake is unique by its extreme salinity. Assessment of physicochemical variables, water chemistry and trophic status indicated that Brâncoveanu Lake is a permanently stratified, pH-neutral, NaCl-rich and eutrophied system. We investigated the abundance, molecular diversity and vertical distribution of archaeal community by culture-independent approaches. Additionally, the most relevant environmental parameters shaping the archaeal community composition were evaluated by statistical methods. Archaea appeared to largely outnumber Bacteria; altogether the great prevalence of Halobacteriaceae-related sequences could imply a major contribution of this group to the biogeochemical carbon turnover. The fairly distinct composition of archaeal communities reflects the lake's physicochemical stratification. Among the limnological factors, salinity and oxygen showed a significant impact on determining the composition and structure of archaeal assemblages. Furthermore, Brâncoveanu Lake might harbor novel microorganisms such as members of the recently described phylum Nanohaloarchaea. Overall, this study reported the occurrence of halophilic Archaea in a little explored hydrogeochemical system and provided a better insight into geomicrobiology of meromictic hypersaline pit lakes.     

Potassium channel blocker crafted by α-hairpinin scaffold engineering

The α-Hairpinins are a family of plant defense peptides with a common fold presenting two short α-helices stabilized by two invariant S–S-bridges. We have shown previously that substitution of just two amino acid residues in a wheat α-hairpinin Tk-AMP-X2 leads to Tk-hefu-2 that features specific affinity to voltage-gated potassium channels K_V1.3. Here, we utilize a combined molecular modeling approach based on molecular dynamics simulations and protein surface topography technique to improve the affinity of Tk-hefu-2 to K_V1.3 while preserving its specificity. An important advance of this work compared with our previous studies is transition from the analysis of various physicochemical properties of an isolated toxin molecule to its consideration in complex with its target, a membrane-bound ion channel. As a result, a panel of computationally designed Tk-hefu-2 derivatives was synthesized and tested against K_V1.3. The most active mutant Tk-hefu-10 showed a half-maximal inhibitory concentration of ∼150 nM being >10 times more active than Tk-hefu-2 and >200 times more active than the original Tk-hefu. We conclude that α-hairpinins provide an attractive disulfide-stabilized scaffold for the rational design of ion channel inhibitors. Furthermore, the success rate can be considerably increased by the proposed “target-based” iterative strategy of molecular design.     

Adult cannibalism in an oligophagous herbivore,the Colorado potato beetle

Cannibalism, or intraspecific predation, can play a major role in changing individual fitness and population processes. In insects, cannibalism frequently occurs across life stages, with cannibals consuming a smaller or more vulnerable stage. Predation of adult insects on one another is considered to be uncommon. We investigated adult cannibalism in the Colorado potato beetle, Leptinotarsa decemlineata (Say), which is an oligophagous herbivore specializing on plants in family Solanaceae, and an important agricultural pest. Under laboratory conditions, starvation and crowding encouraged teneral adults to feed upon each other, which reduced their weight loss during the period of starvation. However, pupae were attacked and consumed before adults. Injured beetles had a higher probability of being cannibalized than intact beetles. Males were more frequently attacked than females, but that appeared to be a function of their smaller size rather than other gender‐specific traits. Cannibalizing eggs at a larval stage did not affect beetle propensity to cannibalize adults at an adult stage. When given a choice between conspecific adults and mealworms, the beetles preferred to eat conspecifics. Cannibalistic behavior, including adult cannibalism, could be important for population persistence in this species.     

Interleukin-11 binds specific EF-hand proteins via their conserved structural motifs

Interleukin-11 (IL-11) is a hematopoietic cytokine engaged in numerous biological processes and validated as a target for treatment of various cancers. IL-11 contains intrinsically disordered regions that might recognize multiple targets. Recently we found that aside from IL-11RA and gp130 receptors, IL-11 interacts with calcium sensor protein S100P. Strict calcium dependence of this interaction suggests a possibility of IL-11 interaction with other calcium sensor proteins. Here we probed specificity of IL-11 to calcium-binding proteins of various types: calcium sensors of the EF-hand family (calmodulin, S100B and neuronal calcium sensors: recoverin, NCS-1, GCAP-1, GCAP-2), calcium buffers of the EF-hand family (S100G, oncomodulin), and a non-EF-hand calcium buffer (α-lactalbumin). A specific subset of the calcium sensor proteins (calmodulin, S100B, NCS-1, GCAP-1/2) exhibits metal-dependent binding of IL-11 with dissociation constants of 1–19 μM. These proteins share several amino acid residues belonging to conservative structural motifs of the EF-hand proteins, ‘black’ and ‘gray’ clusters. Replacements of the respective S100P residues by alanine drastically decrease its affinity to IL-11, suggesting their involvement into the association process. Secondary structure and accessibility of the hinge region of the EF-hand proteins studied are predicted to control specificity and selectivity of their binding to IL-11. The IL-11 interaction with the EF-hand proteins is expected to occur under numerous pathological conditions, accompanied by disintegration of plasma membrane and efflux of cellular components into the extracellular milieu.