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Cortical cell size regulates root metabolic cost
SUMMARY It has been hypothesized that vacuolar occupancy in mature root cortical parenchyma cells regulates root metabolic cost and thereby plant fitness under conditions of drought, suboptimal nutrient availability, and increased soil mechanical impedance. However, the mechanistic role of vacuoles in reducing root metabolic cost was unproven. Here we provide evidence to support this hypothesis. We first show that root cortical cell size is determined by both cortical cell diameter and cell length. Significant genotypic variation for both cortical cell diameter (~1.1‐ to 1.5‐fold) and cortical cell length (~ 1.3‐ to 7‐fold) was observed in maize and wheat. GWAS and QTL analyses indicate cortical cell diameter and length are heritable and under independent genetic control. We identify candidate genes for both phenes. Empirical results from isophenic lines contrasting for cortical cell diameter and length show that increased cell size, due to either diameter or length, is associated with reduced root respiration, nitrogen content, and phosphorus content.RootSlice, a functional‐structural model of root anatomy, predicts that an increased vacuolar: cytoplasmic ratio per unit cortical volume causes reduced root respiration and nutrient content. Ultrastructural imaging of cortical parenchyma cells with varying cortical diameter and cortical cell length confirms thein silicopredictions and shows that an increase in cell size is correlated with increased vacuolar volume and reduced cytoplasmic volume. Vacuolar occupancy and its relationship with cell size merits further investigation as a phene for improving crop adaptation to edaphic stress.
Structural dissection of ergosterol metabolism reveals a pathway optimized for membrane phase separation
Sterols are among the most abundant lipids in eukaryotic cells yet are synthesized through notoriously long metabolic pathways. It has been proposed that the molecular evolution of such pathways must have required each step to increase the capacity of its product to condense and order phospholipids. Here, we carry out a systematic analysis of the ergosterol pathway that leverages the yeast vacuole’s capacity to phase separate into ordered membrane domains. In the post-synthetic steps specific to ergosterol biosynthesis, we find that successive modifications act to oscillate ordering capacity, settling on a level that supports phase separation while retaining fluidity of the resulting domains. Simulations carried out with each intermediate showed how conformers in the sterol’s alkyl tail are capable of modulating long-range ordering of phospholipids, which could underlie changes in phase behavior. Our results indicate that the complexity of sterol metabolism could have resulted from the need to balance lipid interactions required for membrane organization.
The Legionella Effector SdjA Is a Bifunctional Enzyme That Distinctly Regulates Phosphoribosyl Ubiquitination
Legionella pneumophila promotes its survival and replication in phagocytes by actively modulating cellular processes using effectors injected into host cells by its Dot/Icm type IV secretion system. Many of these effectors function to manipulate the ubiquitin network of infected cells, thus contributing to the biogenesis of the Legionella-containing vacuole (LCV), which is permissive for bacterial replication. Among these, members of the SidE effector family (SidEs) catalyze ubiquitination of functionally diverse host proteins by a mechanism that is chemically distinct from the canonical three-enzyme cascade. The activity of SidEs is regulated by two mechanisms: reversal of the phosphoribosyl ubiquitination by DupA and DupB and direct inactivation by SidJ, which is a calmodulin-dependent glutamylase. In many L. pneumophila strains, SidJ belongs to a two-member protein family. Its homolog SdjA appears to function differently from SidJ despite the high-level similarity in their primary sequences. Here, we found that SdjA is a bifunctional enzyme that exhibits distinct activities toward members of the SidE family. It inhibits the activity of SdeB and SdeC by glutamylation. Unexpectedly, it also functions as a deglutamylase that reverses SidJ-induced glutamylation on SdeA. Our results reveal that an enzyme can catalyze two completely opposite biochemical reactions, which highlights the distinct regulation of phosphoribosyl ubiquitination by the SidJ effector family.
Adaptive Optical Two-Photon Fluorescence Microscopy Probes Cellular Organization of Ocular Lenses In Vivo
The mammalian ocular lens is an avascular multicellular organ that grows continuously throughout life. Traditionally, its cellular organization is investigated using dissected lenses, which eliminates in vivo environmental and structural support. Therefore, in vivo optical imaging methods for studying lenses in their native context in live animals are urgently needed. Here, we demonstrated that two-photon fluorescence microscopy can visualize lens cells in vivo. To maintain subcellular resolution at depth, we used adaptive optics to correct aberrations owing to ocular and lens tissues, which led to substantial signal and resolution improvements. Imaging lens cells up to 980 µm deep, we observed novel cellular organizations including suture-associated voids, enlarged vacuoles, and large cavities, contrary to the conventional view of a highly ordered organization. We tracked these features longitudinally over weeks and observed the incorporation of new cells during growth. Taken together, noninvasive longitudinal in vivo imaging of lens morphology using adaptive optics two-photon fluorescence microscopy will allow us to observe the development or alterations of lens cellular organization in living animals directly.
Uptake and speciation of zinc in edible plants grown in smelter contaminated soils
Heavy metal accumulation in edible plants grown in contaminated soils poses a major environmental risk to humans and grazing animals. This study focused on the concentration and speciation of Zn in different edible plants grown in soils contaminated with smelter wastes (Spelter, WV, USA) containing high levels of the metals Zn, Cu, Pb, Cd. Their accumulation was examined in different parts (roots, stem, and leaves) of plants and as a function of growth stage (dry seed, sprouting seed, cotyledon, and leaves) in the root vegetables radish, the leafy vegetable spinach and the legume clover. Although the accumulation of metals varied significantly with plant species, the average metal concentrations were [Zn] > [Pb] > [Cu] > [Cd]. Metal uptake studies were complemented with bulk and micro X-ray absorption spectroscopy (XAS) at Zn K-edge and micro X-ray fluorescence (μXRF) measurements to evaluate the speciation and distribution of Zn in these plant species. Dynamic interplay between the histidine and malate complexation of Zn was observed in all plant species. XRF mapping of spinach leaves at micron spatial resolution demonstrated the accumulation of Zn in vacuoles and leaf tips. Radish root showed accumulation of Zn in root hairs, likely as ZnS nanoparticles. At locations of high Zn concentration in spinach leaves, μXANES suggests Zn complexation with histidine, as opposed to malate in the bulk leaf. These findings shed new light on the dynamic nature of Zn speciation in plants.
Genome-Wide Association Study Reveals Complex Genetic Architecture of Cadmium and Mercury Accumulation and Tolerance Traits in Medicago truncatula
Heavy metals are an increasing problem due to contamination from human sources that and can enter the food chain by being taken up by plants. Understanding the genetic basis of accumulation and tolerance in plants is important for reducing the uptake of toxic metals in crops and crop relatives, as well as for removing heavy metals from soils by means of phytoremediation. Following exposure of Medicago truncatula seedlings to cadmium (Cd) and mercury (Hg), we conducted a genome-wide association study using relative root growth (RRG) and leaf accumulation measurements. Cd and Hg accumulation and RRG had heritability ranging 0.44 – 0.72 indicating high genetic diversity for these traits. The Cd and Hg trait associations were broadly distributed throughout the genome, indicated the traits are polygenic and involve several quantitative loci. For all traits, candidate genes included several membrane associated ATP-binding cassette transporters, P-type ATPase transporters, oxidative stress response genes, and stress related UDP-glycosyltransferases. The P-type ATPase transporters and ATP-binding cassette protein-families have roles in vacuole transport of heavy metals, and our findings support their wide use in physiological plant responses to heavy metals and abiotic stresses. We also found associations between Cd RRG with the genes CAX3 and PDR3, two linked adjacent genes, and leaf accumulation of Hg associated with the genes NRAMP6 and CAX9. When plant genotypes with the most extreme phenotypes were compared, we found significant divergence in genomic regions using population genomics methods that contained metal transport and stress response gene ontologies. Several of these genomic regions show high linkage disequilibrium (LD) among candidate genes suggesting they have evolved together. Minor allele frequency (MAF) and effect size of the most significant SNPs was negatively correlated with large effect alleles being most rare. This is consistent with purifying selection against alleles that increase toxicity and abiotic stress. Conversely, the alleles with large affect that had higher frequencies that were associated with the exclusion of Cd and Hg. Overall, macroevolutionary conservation of heavy metal and stress response genes is important for improvement of forage crops by harnessing wild genetic variants in gene banks such as the Medicago HapMap collection.
Mechanisms of substrate recognition by a typhoid toxin secretion-associated muramidase
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Legionella pneumophila modulates host energy metabolism by ADP-ribosylation of ADP/ATP translocases
The intracellular pathogen Legionella pneumophila delivers more than 330 effectors into host cells by its Dot/Icm type IV secretion system. Those effectors are essential for the biogenesis of the Legionella-containing vacuole (LCV) that permits its intracellular survival and replication. It has long been documented that the LCV is associated with mitochondria and a number of Dot/Icm effectors have been shown to target to this organelle. Yet, the biochemical function and host cell target of most of these effectors remain unknown. Here, we found that the Dot/Icm substrate Ceg3 (Lpg0080) is a mono-ADP-ribosyltransferase that localizes to the mitochondria in host cells where it attacks ADP/ATP translocases (ANTs) by ADP-ribosylation and blunts their ADP/ATP exchange activity. The modification occurs on the second arginine residue in the -RRRMMM- element, which is conserved among all known ADP/ATP carriers from different organisms. Our results reveal modulation of host energy metabolism as a novel virulence mechanism for L. pneumophila.
Fatty change of the granular pneumocyte.
Cytoplasmic alterations and fat vacuole formation in pneumocytes of guinea pigs exposed to severe hypoxia in low pressure chamber
Effect of lunar materials on plant tissue culture.
Lunar material collected during the Apollo 11, 12, 14, and 15 missions has been used to treat 12 species of higher plant tissue cultures. Biochemical and morphological studies have been conducted on several of these species. Tobacco tissue cultures treated with 0.22 g of lunar material exhibited increased greening more complex chloroplasts, less cytoplasmic vacuolation and greater vesiculation. Pine tissue cultures reacted to treatment by an increased deposition of tannin-like materials. The percentage of dry weight and soluble protein was increased in cultures treated with either lunar or terrestrial rock materials.
Microanalytical identification of barium sulphate crystals in statoliths of Chara Rhizoids
In contrast to higher plants, Chara rhizoids contain statolith vacuoles filled with biocrystallites of BaS04 in the form of rods composed of globular subunits ca. 7 nm in diameter. The revelation of the crystallites under electron microscopy is dependent on the fixative; best structural preservation was observed after fixation in a buffered glutaraldehyde + acrolein solution; 0s04 and KMnO4 partially dissolved both the biocrystallites and synthetic BaS04.
Ultrastructural studies of the mitochondriae in the striated muscles of birds with regard to experimental hypokinesis
Electron microscopic studies were carried out on the mitochrondria of the transversely striated muscles with regard to experimental hypokinesia. As compared to the central group the mitochondria of m. pectoralis thoracicus and the m. iliotibialis posterior in hypokinetic birds reveal marked changes. In filamentous and ovoid mitochondria, vacuoles can be observed which in some cases produced larger light formations with following disappearance of the cristae and destruction of mitochondria. Fat particles located at the poles of the altered mitochondria, sporadically occurring also laterally, presented another finding. The Z-lines of the sarcomere did not form a continuous line, but were somewhat shifted.
Morphological changes in neurons of the hind limb reflex arc during long term immobilization
Twelve adult rabbits were immobilized for 9 to 31 days, followed by histological study of the nerve processes of lumbar vertebra 7 and sacral vertebra 1, the sciatic nerve and the motor endings of the thigh muscles. In the spinal ganglia, dystrophic changes of increasing severity with immobilization time were found, including pericellular edema, vacuolized neuroplasm, pycnotic changes, cytolysis and destruction. Chromatophilic matter decreased and was partly bleached, and amitotic division occurred. A portion of the sciatic nerve fibers were argentophilic, and some fragmentary decomposition occurred. Considerable dystrophic changes occurred in the motor nerve endings.
Ultrastructure of the root cap of Arabidopsis Thaliana L. Heynh under spaceflight conditions
Peculiarities of the ultrastructural organization of Arabidopsis root cap cells grown from the stage of two cotyledonous leaves in the Svetoblok-1 apparatus aboard the Salyut 6 research orbital station and in the laboratory are assessed. It is established that under conditions of real space flight vacuolization of the root cap cells increses considerably compared to the control variant. Changes in the topography and ulstrastructure of amyloplasts as well as lysis of cell walls are observed in the material under study. An assumption is advanced on analogous cell responses observed at the ultrastructural level to weightlessness and clinostatic conditions.
Report of prochloron research, IPE-7 (Palau, February 1982)
Various aspects of Prochloron research are discussed. At suitable low-tide periods about 5-6 new sites were surveyed as possible convenient sources of symbiotic didemnids. The Kanori Channel site peviously surveyed during IPE-VI remains by far the best, in terms of species, quantities and accessibility. Prochloron from the six major species of symbiotic didemnids was compared serum, cell size and vacuolation, etc. Tadpoles from Lissoclinum patella colonies were observed emerging from cloacal apertures; about 400 were collected. All but 4 carried a girdle of symbiotic Prochloron cells (about 40,000 per larva). Observations were made on cell viability indicated that a marked increase in protoplasm viscosity of the cell contents was associated with cell death. Living cells, in 5 microlitres of buffered sea water under a coverslip, when pressed with a 2 kg weight for 10 seconds, attempts made to culture Prochloron in sea-water media.
Distribution of calmodulin in corn seedlings - Immunocytochemical localization in coleoptiles and root apices
Immunofluorescence techniques have been used to study the distribution of calmodulin in several tissues in etiolated corn (Zea mays, var. Bear Hybrid) seedlings. Uniform staining was seen in the background cytoplasm of most cell types. Cell walls and vacuoles were not stained. In coleoptile mesophyll cells the nucleoplasm of most nuclei was stained as was the stroma of most amyloplasts. The lumen border of mature tracheary elements in coleoptiles also stained. In the rootcap the most intensely stained regions were the cytoplasms of columella cells and of the outermost cells enmeshed in the layer of secreted slime. Nuclei in the rootcap cells did not stain distinctly, but those in all cell types of the root meristem did. Also in the root meristem, the cytoplasm of metaxylem elements stained brightly. These results are compared and contrasted with previous data on the localization of calmodulin in pea root apices and epicotyls and discussed in relation to current hypotheses on mechanisms of gravitropism.
Simulated Microgravity Induced Cytoskeletal Rearrangements are Modulated by Protooncogenes
Microgravity is the environment living systems encounter during space flight and gravitational unloading is the effect of this environment on living systems. The cell, being a multiphasic chemical system, is a useful starting point to study the potential impact of gravity unloading on physiological function. In the absence of gravity, sedimentation of organelles including chromosomes, mitochondria, nuclei, the Golgi apparatus, vacuoles, and the endoplasmic reticulum may be affected. Most of these organelles, however, are somewhat held in place by cytoskeleton. Hansen and Igber suggest that intermediate filaments act to stabilize the nuleus against rotational movement, and integrate cell and nuclear structure. The tensegrity theory supports the idea that mechanical or physical forces alters the cytoskeletal structures of a cell resulting in the changes in cell: matrix interactions and receptor-signaling coupling. This type of stress to the cytoskeleton may be largely responsible regulating cell shape, growth, movement and metabolism. Mouse MC3T3 El cells under microgravity exhibited significant cytoskeletal changes and alterations in cell growth. The alterations in cytoskeleton architecture may be due to changes in the expression of actin related proteins or integrins. Philopott and coworkers reported on changes in the distribution of microtubule and cytoskeleton elements in the cells of heart tissue from space flight rats and those centrifuged at 1.7g. Other researchers have showed that microgravity reduced EGF-induced c-fos and c-jun expression compared to 1 g controls. Since c-fos and c-jun are known regulators of cell growth, it is likely that altered signal transduction involving protooncogenes may play a crucial role in the reduced growth and alterations in cytoskeletal arrangements found during space flight. It is clear that a microgravity environment induces a number of changes in cell shape, cell surface molecules, gene expression, and cytoskeletal reorganization. However the underlying mechanism for these cellular changes have not been clearly defined. We examined alterations in endothelial migration, and cytoskeleton architecture (microfilamentous f-actin and vimentin-rich- intermediate filaments) following wounding under simulated microgravity. We also examined the possibility that altered signal transduction pathways, involving protooncogenes, may play a crucial role in microgravity-induced retardation of cell migration and alterations in cytoskeletal organization. We hypothesize that, based on the tensegrity theory, cytoskeletal organization respond to gravitational unloading and through this response, cell behavior, function and gene expression are modified.