Same but different — pseudo-pectin in the charophytic alga Chlorokybus atmophyticus

All land‐plant cell walls possess hemicelluloses, cellulose and anionic pectin. The walls of their cousins, the charophytic algae, exhibit some similarities to land plants’ but also major differences. Charophyte ‘pectins’ are extractable by conventional land‐plant methods, although they differ significantly in composition. Here, we explore ‘pectins’ of an early‐diverging charophyte, Chlorokybus atmophyticus, characterising the anionic polysaccharides that may be comparable to ‘pectins’ in other streptophytes. Chlorokybus ‘pectin’ was anionic and upon acid hydrolysis gave GlcA, GalA and sulphate, plus neutral sugars (Ara≈Glc>Gal>Xyl); Rha was undetectable. Most Gal was the l‐enantiomer. A relatively acid‐resistant disaccharide was characterised as β‐d‐GlcA‐(1→4)‐l‐Gal. Two Chlorokybus ‘pectin’ fractions, separable by anion‐exchange chromatography, had similar sugar compositions but different sulphate‐ester contents. No sugars were released from Chlorokybus ‘pectin’ by several endo‐hydrolases [(1,5)‐α‐l‐arabinanase, (1,4)‐β‐d‐galactanase, (1,4)‐β‐d‐xylanase, endo‐polygalacturonase] and exo‐hydrolases [α‐ and β‐d‐galactosidases, α‐(1,6)‐d‐xylosidase]. ‘Driselase’, which hydrolyses most land‐plant cell wall polysaccharides to mono‐ and disaccharides, released no sugars except traces of starch‐derived Glc. Thus, the Ara, Gal, Xyl and GalA of Chlorokybus ‘pectin’ were not non‐reducing termini with configurations familiar from land‐plant polysaccharides (α‐l‐Araf, α‐ and β‐d‐Galp, α‐ and β‐d‐Xylp and α‐d‐GalpA), nor mid‐chain residues of α‐(1→5)‐l‐arabinan, β‐(1→4)‐d‐galactan, β‐(1→4)‐d‐xylan or α‐(1→4)‐d‐galacturonan. In conclusion, Chlorokybus possesses anionic ‘pectic’ polysaccharides, possibly fulfilling pectic roles but differing fundamentally from land‐plant pectin. Thus, the evolution of land‐plant pectin since the last common ancestor of Chlorokybus and land plants is a long and meandering path involving loss of sulphate, most l‐Gal and most d‐GlcA; re‐configuration of Ara, Xyl and GalA; and gain of Rha

Agrobacterium-mediated gene delivery and transient expression in the red macroalga Chondrus crispus

Molecular resources and transgenic studies in red algae are lagging behind those for green algae. The Agrobacterium-mediated gene-transfer method routinely used in plant transformation has not been fully utilised in the red algae, which, as an important source of phycocolloids, warrant more studies. In this regard, a stepwise methodology was developed for Agrobacterium-mediated transformation of the carrageenophyte Chondrus crispus using pCAMBIA 1301 and a construct featuring a codon-optimized beta-glucuronidase (GUS) reporter gene driven by the endogenous Chondrus actin promoter. The effects of several factors on transformation efficiency were investigated. An intimate association of Chondrus and bacterial cells was observed using scanning electron microscopy. GUS transient expression within Chondrus cortical and medullary cells with both expression cassettes testified to the amenability of Chondrus to Agrobacterium-mediated transformation. Darker staining, indicative of higher GUS activity, was observed with the Chondrus-specific construct, suggesting its superiority over the pCAMBIA 1301. Presence of acetosyringone, the wounding method and the type of co-cultivation medium significantly affected the transformation outcome and efficiency. The Agrobacterium-mediated transient expression presented here constitutes a first step towards tailoring a transformation strategy for Chondrus, which can serve to facilitate further transgenic studies in this important red alga

Insights into the Evolution of Multicellularity from the Sea Lettuce Genome

We report here the 98.5 Mbp haploid genome (12,924 protein coding genes) of Ulva mutabilis, a ubiquitous and iconic representative of the Ulvophyceae or green seaweeds. Ulva’s rapid and abundant growth makes it a key contributor to coastal biogeochemical cycles; its role in marine sulfur cycles is particularly important because it produces high levels of dimethylsulfoniopropionate (DMSP), the main precursor of volatile dimethyl sulfide (DMS). Rapid growth makes Ulva attractive biomass feedstock but also increasingly a driver of nuisance “green tides.” Ulvophytes are key to understanding the evolution of multicellularity in the green lineage, and Ulva morphogenesis is dependent on bacterial signals, making it an important species with which to study cross-kingdom communication. Our sequenced genome informs these aspects of ulvophyte cell biology, physiology, and ecology. Gene family expansions associated with multicellularity are distinct from those of freshwater algae. Candidate genes, including some that arose following horizontal gene transfer from chromalveolates, are present for the transport and metabolism of DMSP. The Ulva genome offers, therefore, new opportunities to understand coastal and marine ecosystems and the fundamental evolution of the green lineage

Optical Atomic Clock Comparison through Turbulent Air

We use frequency comb-based optical two-way time-frequency transfer (O-TWTFT) to measure the optical frequency ratio of state-of-the-art ytterbium and strontium optical atomic clocks separated by a 1.5 km open-air link. Our free-space measurement is compared to a simultaneous measurement acquired via a noise-cancelled fiber link. Despite non-stationary, ps-level time-of-flight variations in the free-space link, ratio measurements obtained from the two links, averaged over 30.5 hours across six days, agree to $6\times10^{-19}$, showing that O-TWTFT can support free-space atomic clock comparisons below the $10^{-18}$ level

A genetic cause of Alzheimer disease: mechanistic insights from Down syndrome

Down syndrome, caused by an extra copy of chromosome 21, is associated with a greatly increased risk of early onset Alzheimer disease. It is thought that this risk is conferred by the presence of three copies of the gene encoding amyloid precursor protein (APP), an Alzheimer risk factor, although the possession of extra copies of other chromosome 21 genes may also play a role. Further study of the mechanisms underlying the development of Alzheimer disease in Down syndrome could provide insights into the mechanisms that cause dementia in the general population

The effects of spatial scale and isoscape on consumer isotopic niche width

1. The mean and variance of ecological variables are dependent on sampling attributes such as the coverage of environmental heterogeneity (sampling extent) and spatial scale. Trophic niche width is often approximated by bulk tissue stable isotopes of C and N, i.e. the population isotopic niche. However, recent studies suggest that environmental heterogeneity (experienced by individuals) may be more important in defining the isotopic niche width than trophic variability. We hypothesised that isotopic niche width will increase monotonically with spatial scale, largely produced by environmental variation, e.g. nutrient source. 2. To refine this hypothesis, by describing the shapes of isotope scaling curves, we explored a previously published dataset describing three Chilean intertidal species representing different feeding guilds (grazing snails, suspension feeding mussel). We tested these hypotheses on a new, larger dataset describing three functionally-analogous intertidal species from Northern Ireland. We generated isotopic variance-area curves from a spatially-explicit bootstrap and investigated the scale-dependency of environment-isotope relationships, including wave exposure and sub-habitat heterogeneity. 3. Spatial scale explained 50% of the variance in population isotopic niche widths (bivariate C-N ellipse area) by simple, non-linear relationships. Finer scales ( 40% was explained by modelling linear coefficients. A ẟ15N baseline gradient, or isoscape, dominated ẟ15N variance scaling patterns, from sheltered, terrestrially-influenced embayments to exposed, pelagic-dominated coastline. Consumer ẟ13C variance had a weaker scale-dependence, plateauing at mesoscales (> 20 km lag). 4. We show that isotopic niche width is strongly dependent on sampling spatial extent, which controls the environmental heterogeneity experienced by individual consumers. Environmental heterogeneity must be accounted for before isotopic niche width can be considered to accurately represent trophic niche width. Studies conducted at different spatial scales are likely to identify different environment-isotope relationships. 5. We recommend that spatial scale should be incorporated into sampling designs explicitly, easiest by maintaining a consistent lag distance or area within which populations are sampled. Identified isoscapes can be de-trended, where necessary.Table S4. Northern Irish environment and consumers' stable isotope dataThis file contains all consumer (ME = Mytilus, LL = Littorina littorea, PV = Patella vulgata) stable isotope values alongside environmental variables for the site.FEreddinST4.csvTable S5. Northern Irish macroalgal stable isotope data.This file contains macroalgal (Laminaria digitata, Ulva spp., Fucus serratus) stable isotope values.FEreddinST5.csvR code for isotope scaling curves and plottingDirectories need changing. All files imported are also made in this R code from the original data files. The file 'Covariogrammodels.csv' is attached separately for consistencyIsotope scaling curves.RCovariogram model parametersThis file is needed to plot the covariograms, but is also made in 'Isotope scaling curves.R'Covariogrammodels.csvR code for modelling the effect of sample sizeFiles needed for this code are either made within it or in 'Isotope scaling curves.R' from the original data.SampleSizeModelling.

Monitoring apoptosis in intact cells by high‐resolution magic angle spinning 1 H NMR spectroscopy

Apoptosis maintains an equilibrium between cell proliferation and cell death. Many diseases, including cancer, develop because of defects in apoptosis. A known metabolic marker of apoptosis is a notable increase in 1H NMR‐observable resonances associated with lipids stored in lipid droplets. However, standard one‐dimensional NMR experiments allow the quantification of lipid concentration only, without providing information about physical characteristics such as the size of lipid droplets, viscosity of the cytosol, or cytoskeletal rigidity. This additional information can improve monitoring of apoptosis‐based cancer treatments in intact cells and provide us with mechanistic insight into why these changes occur. In this paper, we use high‐resolution magic angle spinning (HRMAS) 1H NMR spectroscopy to monitor lipid concentrations and apparent diffusion coefficients of mobile lipid in intact cells treated with the apoptotic agents cisplatin or etoposide. We also use solution‐state NMR spectroscopy to study changes in lipid profiles of organic solvent cell extracts. Both NMR techniques show an increase in the concentration of lipids but the relative changes are 10 times larger by HRMAS 1H NMR spectroscopy. Moreover, the apparent diffusion rates of lipids in apoptotic cells measured by HRMAS 1H NMR spectroscopy decrease significantly as compared with control cells. Slower diffusion rates of mobile lipids in apoptotic cells correlate well with the formation of larger lipid droplets as observed by microscopy. We also compared the mean lipid droplet displacement values calculated from the two methods. Both methods showed shorter displacements of lipid droplets in apoptotic cells. Our results demonstrate that the NMR‐based diffusion experiments on intact cells discriminate between control and apoptotic cells. Apparent diffusion measurements in conjunction with 1H NMR spectroscopy‐derived lipid signals provide a novel means of following apoptosis in intact cells. This method could have potential application in enhancing drug discovery by monitoring drug treatments in vitro, particularly for agents that cause portioning of lipids such as apoptosis

Bioloistic delivery of Ca2+ dyes into plant and algal cells.

In eukaryotes, changes in cytosolic Ca2+ concentrations ([Ca2+]cyt) are associated with a number of environmental and developmental stimuli. However, measuring [Ca2+]cyt changes in single plant or algal cells is often problematic. Although a wide range of Ca2+-sensitive fluorescent dyes is available, they are often difficult to introduce into plant cells. Micro-injection is the most robust method for dye loading, but is time-consuming, technically demanding, and unsuitable in many cell types. To overcome these problems, we have adapted biolistic techniques to load Ca2+-sensitive dyes into guard cells of the flowering plant, Commelina communis, cells of the green alga Chlamydomonas reinhardtii, and zygotes of the brown alga, Fucus serratus. Using this approach, we have been able to monitor [Ca2+]cyt changes in response to various stimuli, including a novel [Ca2+]cyt response in C. reinhardtii. The method allows the use of free acid and dextran-conjugated dyes. Biolistic loading of differentiated plant cells is easier, quicker, and more widely applicable than micro-injection, and should broaden the study of plant signal transduction