A Calmodulin C-Lobe Ca2+-Dependent Switch Governs Kv7 Channel Function

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Methane and carbon dioxide dynamics within four vernal pools in Maine,USA

Vernal pools are small, seasonal wetlands that are a common landscape feature contributing to biodiversity in northeastern North American forests. Basic information about their biogeochemical functions, such as carbon cycling, is limited. Concentrations of dissolved methane (CH₄) and carbon dioxide (CO₂) and other water chemistry parameters were monitored weekly at the bottom and surface of four vernal pools in central and eastern Maine, USA, from April to August 2016. The vernal pools were supersaturated with respect to CH₄ and CO₂ at all sampling dates and locations. Concentrations of dissolved CH₄ and CO₂ ranged from 0.4 to 210 μmol L^?1 and 72–2300 μmol L^?1, respectively. Diffusive fluxes of CH₄ and CO₂ into the atmosphere ranged from 0.2 to 73 mmol m^?2 d^?1, and 30–590 mmol m^?2 d^?1, respectively. During the study period, the four vernal pools emitted 0.1–5.8 kg C m^?2 and 9.6–120 kg C m^?2 as CH₄ and CO₂, respectively. The production fluxes (production rates normalized to surface area) of CH₄ and CO₂ ranged from ? 0.02 to 0.66 and 0.40–4.6 g C m^?2 d^?1, respectively, and increased significantly over the season. Methane concentrations were best predicted by alkalinity, ortho-phosphate and depth, while CO₂ concentrations were best predicted with only alkalinity. Alkalinity as a predictor variable highlights the importance of anaerobic respiration in production of both gases. Our study pools had large concentrations and effluxes of CH₄ and CO₂ compared to permanently inundated wetlands, indicating vernal pools are metabolically active sites and may be important contributors to the global carbon budget.     

Assessment of the influence of intrinsic environmental and geographical factors on the bacterial ecology of pit latrines

Improving the rate and extent of faecal decomposition in basic forms of sanitation such as pit latrines would benefit around 1.7 billion users worldwide, but to do so requires a major advance in our understanding of the biology of these systems. As a critical first step, bacterial diversity and composition was studied in 30 latrines in Tanzania and Vietnam using pyrosequencing of 16S rRNA genes, and correlated with a number of intrinsic environmental factors such as pH, temperature, organic matter content/composition and geographical factors. Clear differences were observed at the operational taxonomic unit, family and phylum level in terms of richness and community composition between latrines in Tanzania and Vietnam. The results also clearly show that environmental variables, particularly substrate type and availability, can exert a strong structuring influence on bacterial communities in latrines from both countries. The origins and significance of these environmental differences are discussed. This work describes the bacterial ecology of pit latrines in combination with inherent latrine characteristics at an unprecedented level of detail. As such, it provides useful baseline information for future studies that aim to understand the factors that affect decomposition rates in pit latrines.     

Identifying the core microbial community in the gut of fungus‐growing termites

Gut microbes play a crucial role in decomposing lignocellulose to fuel termite societies, with protists in the lower termites and prokaryotes in the higher termites providing these services. However, a single basal subfamily of the higher termites, the Macrotermitinae, also domesticated a plant biomass‐degrading fungus (Termitomyces), and how this symbiont acquisition has affected the fungus‐growing termite gut microbiota has remained unclear. The objective of our study was to compare the intestinal bacterial communities of five genera (nine species) of fungus‐growing termites to establish whether or not an ancestral core microbiota has been maintained and characterizes extant lineages. Using 454‐pyrosequencing of the 16S rRNA gene, we show that gut communities have representatives of 26 bacterial phyla and are dominated by Firmicutes, Bacteroidetes, Spirochaetes, Proteobacteria and Synergistetes. A set of 42 genus‐level taxa was present in all termite species and accounted for 56–68% of the species‐specific reads. Gut communities of termites from the same genus were more similar than distantly related species, suggesting that phylogenetic ancestry matters, possibly in connection with specific termite genus‐level ecological niches. Finally, we show that gut communities of fungus‐growing termites are similar to cockroaches, both at the bacterial phylum level and in a comparison of the core Macrotermitinae taxa abundances with representative cockroach, lower termite and higher nonfungus‐growing termites. These results suggest that the obligate association with Termitomyces has forced the bacterial gut communities of the fungus‐growing termites towards a relatively uniform composition with higher similarity to their omnivorous relatives than to more closely related termites.     

Biogeochemical Hotspots in Forested Landscapes: The Role of Vernal Pools in Denitrification and Organic Matter Processing

Quantifying spatial and temporal heterogeneity in ecosystem processes presents a challenge for conserving ecosystem function across landscapes. In particular, many ecosystems contain small features that play larger roles in ecosystem processes than their size would indicate; thus, they may represent “hotspots” of activity relative to their surroundings. Biogeochemical hotspots are characterized as small features within a landscape that show comparatively high chemical reaction rates. In northeastern forests in North America, vernal pools are abundant, small features that typically fill in spring with snow melt and precipitation and dry by the end of summer. Ephemeral flooding alters soil moisture and the depth of the soil’s oxic/anoxic boundary, which may affect biogeochemical processes. We studied the effects of vernal pools on leaf-litter decomposition rates, soil enzyme activity, and denitrification in vernal pools to assess whether they function as biogeochemical hotspots. Our results indicate that seasonal inundation enhanced leaf-litter decomposition, denitrification, and enzyme activity in vernal pools relative to adjacent forest sites. Leaves in seasonally flooded areas decomposed faster than leaves in terra firme forest sites. Flooding also influenced the C, N, and P stoichiometry of decomposing leaf litter and explained the variance in microbial extracellular enzyme activity for phosphatase, β-d-glucosidase, and β-N-acetylglucosaminidase. Additionally, denitrification rates were enhanced by seasonal flooding across all of the study pools. Collectively, these data suggest that vernal pool ecosystems may function as hotspots of leaf-litter decomposition and denitrification and play a significant role in decomposition and nutrient dynamics relative to their size.     

Targeted transgene integration in plant cells using designed zinc finger nucleases

Targeted transgene integration in plants remains a significant technical challenge for both basic and applied research. Here it is reported that designed zinc finger nucleases (ZFNs) can drive site-directed DNA integration into transgenic and native gene loci. A dimer of designed 4-finger ZFNs enabled intra-chromosomal reconstitution of a disabled gfp reporter gene and site-specific transgene integration into chromosomal reporter loci following co-transformation of tobacco cell cultures with a donor construct comprised of sequences necessary to complement a non-functional pat herbicide resistance gene. In addition, a yeast-based assay was used to identify ZFNs capable of cleaving a native endochitinase gene. Agrobacterium delivery of a Ti plasmid harboring both the ZFNs and a donor DNA construct comprising a pat herbicide resistance gene cassette flanked by short stretches of homology to the endochitinase locus yielded up to 10% targeted, homology-directed transgene integration precisely into the ZFN cleavage site. Given that ZFNs can be designed to recognize a wide range of target sequences, these data point toward a novel approach for targeted gene addition, replacement and trait stacking in plants.     

Communication between accessory factors and the Cre recombinase at hybrid psi-loxP sites

By placing loxP adjacent to the accessory sequences from the Xer/psi multimer resolution system, we have imposed topological selectivity and specificity on Cre/loxP recombination. In this hybrid recombination system, the Xer accessory protein PepA binds to psi accessory sequences, interwraps them, and brings the loxP sites together such that the product of recombination is a four-node catenane. Here, we investigate communication between PepA and Cre by varying the distance between loxP and the accessory sequences, and by altering the orientation of loxP. The yield of four-node catenane and the efficiency of recombination in the presence of PepA varied with the helical phase of loxP with respect to the accessory sequences. When the orientation of loxP was reversed, or when half a helical turn was added between the accessory sequences and loxP, PepA reversed the preferred order of strand exchange by Cre at loxP. The results imply that PepA and the accessory sequences define precisely the geometry of the synapse formed by the loxP sites, and that this overcomes the innate preference of Cre to initiate recombination on the bottom strand of loxP. Further analysis of our results demonstrates that PepA can stimulate strand exchange by Cre in two distinct synaptic complexes, with the C-terminal domains of Cre facing either towards or away from PepA. Thus, no specific PepA-recombinase interaction is required, and correct juxtaposition of the loxP sites is sufficient to activate Cre in this system.