An enhancer trap system to track developmental dynamics in Marchantia polymorpha.

A combination of streamlined genetics, experimental tractability and relative morphological simplicity compared to vascular plants makes the liverwort Marchantia polymorpha an ideal model system for studying many aspects of plant biology. Here we describe a transformation vector combining a constitutive fluorescent membrane marker with a nuclear marker that is regulated by nearby enhancer elements and use this to produce a library of enhancer trap lines for Marchantia. Screening gemmae from these lines allowed the identification and characterization of novel marker lines, including markers for rhizoids and oil cells. The library allowed the identification of a margin tissue running around the thallus edge, highlighted during thallus development. The expression of this marker is correlated with auxin levels. We generated multiple markers for the meristematic apical notch region, which have different spatial expression patterns, reappear at different times during meristem regeneration following apical notch excision and have varying responses to auxin supplementation or inhibition. This reveals that there are proximodistal substructures within the apical notch that could not be observed otherwise. We employed our markers to study Marchantia sporeling development, observing meristem emergence as defining the protonema-to-prothallus stage transition, and subsequent production of margin tissue during the prothallus stage. Exogenous auxin treatment stalls meristem emergence at the protonema stage but does not inhibit cell division, resulting in callus-like sporelings with many rhizoids, whereas pharmacologically inhibiting auxin synthesis and transport does not prevent meristem emergence. This enhancer trap system presents a useful resource for the community and will contribute to future Marchantia research.

Agrobacterium-Mediated Transient Transformation of Marchantia Liverworts.

Agrobacterium-mediated transient gene expression is a rapid and useful approach for characterizing functions of gene products in planta. However, the practicability of the method in the model liverwort Marchantia polymorpha has not yet been thoroughly described. Here we report a simple and robust method for Agrobacterium-mediated transient transformation of Marchantia thalli and its applicability. When thalli of M. polymorpha were co-cultured with Agrobacterium tumefaciens carrying ß-glucuronidase (GUS) genes, GUS staining was observed primarily in assimilatory filaments and rhizoids. GUS activity was detected 2 days after infection and saturated 3 days after infection. We were able to transiently co-express fluorescently tagged proteins with proper localizations. Furthermore, we demonstrate that our method can be used as a novel pathosystem to study liverwort-bacteria interactions. We also provide evidence that air chambers support bacterial colonization.

An evolutionarily conserved NIMA-related kinase directs rhizoid tip growth in the basal land plant Marchantia polymorpha.

Tip growth is driven by turgor pressure and mediated by the polarized accumulation of cellular materials. How a single polarized growth site is established and maintained is unclear. Here, we analyzed the function of NIMA-related protein kinase 1 (MpNEK1) in the liverwort Marchantia polymorpha In the wild type, rhizoid cells differentiate from the ventral epidermis and elongate through tip growth to form hair-like protrusions. In Mpnek1 knockout mutants, rhizoids underwent frequent changes in growth direction, resulting in a twisted and/or spiral morphology. The functional MpNEK1-Citrine protein fusion localized to microtubule foci in the apical growing region of rhizoids. Mpnek1 knockouts exhibited increases in both microtubule density and bundling in the apical dome of rhizoids. Treatment with the microtubule-stabilizing drug taxol phenocopied the Mpnek1 knockout. These results suggest that MpNEK1 directs tip growth in rhizoids through microtubule organization. Furthermore, MpNEK1 expression rescued ectopic outgrowth of epidermal cells in the Arabidopsis thaliana nek6 mutant, strongly supporting an evolutionarily conserved NEK-dependent mechanism of directional growth. It is possible that such a mechanism contributed to the evolution of the early rooting system in land plants.

Chytrid rhizoid morphogenesis resembles hyphal development in multicellular fungi and is adaptive to resource availability.

Key to the ecological prominence of fungi is their distinctive cell biology, our understanding of which has been principally based on dikaryan hyphal and yeast forms. The early-diverging Chytridiomycota (chytrids) are ecologically important and a significant component of fungal diversity, yet their cell biology remains poorly understood. Unlike dikaryan hyphae, chytrids typically attach to substrates and feed osmotrophically via anucleate rhizoids. The evolution of fungal hyphae appears to have occurred from rhizoid-bearing lineages and it has been hypothesized that a rhizoid-like structure was the precursor to multicellular hyphae. Here, we show in a unicellular chytrid, Rhizoclosmatium globosum, that rhizoid development exhibits striking similarities with dikaryan hyphae and is adaptive to resource availability. Rhizoid morphogenesis exhibits analogous patterns to hyphal growth and is controlled by ß-glucan-dependent cell wall synthesis and actin polymerization. Chytrid rhizoids growing from individual cells also demonstrate adaptive morphological plasticity in response to resource availability, developing a searching phenotype when carbon starved and spatial differentiation when interacting with particulate organic matter. We demonstrate that the adaptive cell biology and associated developmental plasticity considered characteristic of hyphal fungi are shared more widely across the Kingdom Fungi and therefore could be conserved from their most recent common ancestor.

Macroalgal-bacterial interactions: identification and role of thallusin in morphogenesis of the seaweed Ulva (Chlorophyta).

Macroalgal microbiomes have core functions related to biofilm formation, growth, and morphogenesis of seaweeds. In particular, the growth and development of the sea lettuce Ulva spp. (Chlorophyta) depend on bacteria releasing morphogenetic compounds. Under axenic conditions, the macroalga Ulva mutabilis develops a callus-like phenotype with cell wall protrusions. However, co-culturing with Roseovarius sp. (MS2) and Maribacter sp. (MS6), which produce various stimulatory chemical mediators, completely recovers morphogenesis. This ecological reconstruction forms a tripartite community which can be further studied for its role in cross-kingdom interactions. Hence, our study sought to identify algal growth- and morphogenesis-promoting factors (AGMPFs) capable of phenocopying the activity of Maribacter spp. We performed bioassay-guided solid-phase extraction in water samples collected from U. mutabilis aquaculture systems. We uncovered novel ecophysiological functions of thallusin, a sesquiterpenoid morphogen, identified for the first time in algal aquaculture. Thallusin, released by Maribacter sp., induced rhizoid and cell wall formation at a concentration of 11 pmol l-1. We demonstrated that gametes acquired the iron complex of thallusin, thereby linking morphogenetic processes with intracellular iron homeostasis. Understanding macroalgae-bacteria interactions permits further elucidation of the evolution of multicellularity and cellular differentiation, and development of new applications in microbiome-mediated aquaculture systems.

Convergent evolution of water-conducting cells in Marchantia recruited the ZHOUPI gene promoting cell wall reinforcement and programmed cell death.

A key adaptation of plants to life on land is the formation of water-conducting cells (WCCs) for efficient long-distance water transport. Based on morphological analyses it is thought that WCCs have evolved independently on multiple occasions. For example, WCCs have been lost in all but a few lineages of bryophytes but, strikingly, within the liverworts a derived group, the complex thalloids, has evolved a novel externalized water-conducting tissue composed of reinforced, hollow cells termed pegged rhizoids. Here, we show that pegged rhizoid differentiation in Marchantia polymorpha is controlled by orthologs of the ZHOUPI and ICE bHLH transcription factors required for endosperm cell death in Arabidopsis seeds. By contrast, pegged rhizoid development was not affected by disruption of MpNAC5, the Marchantia ortholog of the VND genes that control WCC formation in flowering plants. We characterize the rapid, genetically controlled programmed cell death process that pegged rhizoids undergo to terminate cellular differentiation and identify a corresponding upregulation of conserved putative plant cell death effector genes. Lastly, we show that ectopic expression of MpZOU1 increases production of pegged rhizoids and enhances drought tolerance. Our results support that pegged rhizoids evolved independently of other WCCs. We suggest that elements of the genetic control of developmental cell death are conserved throughout land plants and that the ZHOUPI/ICE regulatory module has been independently recruited to promote cell wall modification and programmed cell death in liverwort rhizoids and in the endosperm of flowering plant seed.

Microbial diversity and their extracellular enzyme activities among different soil particle sizes in mossy biocrust under N limitation in the southeastern Tengger Desert, China.

Introduction: Mossy biocrust represents a stable stage in the succession of biological soil crust in arid and semi-arid areas, providing a microhabitat that maintains microbial diversity. However, the impact of mossy biocrust rhizoid soil and different particle sizes within the mossy biocrust layer and sublayer on microbial diversity and soil enzyme activities remains unclear. Methods: This study utilized Illumina MiSeq sequencing and high-throughput fluorometric technique to assess the differences in microbial diversity and soil extracellular enzymes between mossy biocrust rhizoid soil and different particle sizes within the mossy biocrust sifting and sublayer soil. Results: The results revealed that the total organic carbon (TOC), total nitrogen (TN), ammonium (NH4+) and nitrate (NO3-) in mossy biocrust rhizoid soil were the highest, with significantly higher TOC, TN, and total phosphorus (TP) in mossy biocrust sifting soil than those in mossy biocrust sublayer soil. Extracellular enzyme activities (EAAs) exhibited different responses to various soil particle sizes in mossy biocrust. Biocrust rhizoid soil (BRS) showed higher C-degrading enzyme activity and lower P-degrading enzyme activity, leading to a significant increase in enzyme C: P and N: P ratios. Mossy biocrust soils were all limited by microbial relative nitrogen while pronounced relative nitrogen limitation and microbial maximum relative carbon limitation in BRS. The diversity and richness of the bacterial community in the 0.2 mm mossy biocrust soil (BSS0.2) were notably lower than those in mossy biocrust sublayer, whereas the diversity and richness of the fungal community in the rhizoid soil were significantly higher than those in mossy biocrust sublayer. The predominant bacterial phyla in mossy biocrust were Actinobacteriota, Protebacteria, Chloroflexi, and Acidobacteriota, whereas in BSS0.2, the predominant bacterial phyla were Actinobacteriota, Protebacteria, and Cyanobacteria. Ascomycota and Basidiomycota were dominant phyla in mossy biocrust. The bacterial and fungal community species composition exhibited significant differences. The mean proportions of Actinobacteriota, Protebacteria, Chloroflexi, Acidobacteriota, Acidobacteria, Cyanobacteria, and Bacteroidota varied significantly between mossy biocrust rhizoid and different particle sizes of mossy biocrust sifting and sublayer soil (p < 0.05). Similarly, significant differences (p < 0.05) were observed in the mean proportions of Ascomycota, Basidiomycota, and Glomeromycota between mossy biocrust rhizoid and different particle sizes within the mossy biocrust sifting and sublayer soil. The complexity and connectivity of bacterial and fungal networks were higher in mossy biocrust rhizoid soil compared with different particle sizes within the mossy biocrust sifting and sublayer soil. Discussion: These results offer valuable insights to enhance our understanding of the involvement of mossy biocrust in the biogeochemical cycle of desert ecosystems.

Combined effects of cadmium and zinc on growth, tolerance, and metal accumulation in Chara australis and enhanced phytoextraction using EDTA.

Chara australis (R. Br.) is a macrophytic alga that can grow in and accumulate Cd from artificially contaminated sediments. We investigated the effects of Zn independently and in combination with Cd on C. australis growth, metal tolerance, and uptake. Plant growth was reduced at concentrations ≥ 75 mg Zn (kg soil)⁻¹. Zn also increased the concentration of glutathione in the plant, suggesting alleviation of stress. Phytotoxic effects were observed at ≥ 250 mg added Zn (kg soil)⁻¹. At 1.5mg Zn (kg soil)⁻¹, the rhizoid bioconcentration factor (BCF) was >1.0 for both Cd and Zn. This is a criterion for hyperaccumulator status, a commonly used benchmark for utility in remediation of contaminated soils by phytoextraction. There was no significant interaction between Cd and Zn on accumulation, indicating that Chara should be effective at phytoextraction of mixed heavy metal contamination in sediments. The effects of the chelator, ethylenediaminetetraacetic acid (EDTA), were also tested. Moderate levels of EDTA increased Cd and Zn accumulation in rhizoids and Cd BCF of shoots, enhancing Chara's potential in phytoremediation. This study demonstrates for the first time the potential of macroalgae to remove metals from sediments in aquatic systems that are contaminated with a mixture of metals.

Establishment and application of novel culture methods in Marchantia polymorpha: persistent tip growth is required for substrate penetration by rhizoids.

A NIMA-related protein kinase, MpNEK1, directs tip growth of rhizoids through microtubule depolymerization in a liverwort Marchantia polymorpha. The Mpnek1 knockouts were shown to develop curly and spiral rhizoids due to the fluctuated direction of growth. Still, physiological roles and mechanisms of MpNEK1-dependent rhizoid tip growth remain to be clarified. Here, we developed novel culture methods to further study rhizoid growth of M. polymorpha, in which plants were grown on vertical plates. We applied the established methods to investigate MpNEK1 function in rhizoid growth. Rhizoids of the wild-type and Mpnek1 plants grew toward the gravity. The aerial rhizoids were longer in Mpnek1 than in the wild type. When the rhizoids were grown on the surface of a cellophane sheet, rhizoid length was comparable between the wild type and Mpnek1, whereas Mpnek1 developed more rhizoids compared to the wild type. We also applied gellan gum, which is more transparent than agar, to analyze rhizoids grown in the medium. Rhizoids of Mpnek1 displayed defect on entering into the solid medium. These results suggest that Mpnek1 rhizoids have the deficiency in invasive tip growth. Thus, stable directional growth is important for rhizoids to get into the soil to anchor plant body and to adsorb water and nutrients. Collectively, our newly designed growth systems are valuable for analyzing rhizoid growth.

Three-dimensionally visualized rhizoid system of moss, Physcomitrium patens, by refraction-contrast X-ray micro-computed tomography.

Land plants have two types of shoot-supporting systems, root system and rhizoid system, in vascular plants and bryophytes. However, since the evolutionary origin of the systems is different, how much they exploit common systems or distinct systems to architect their structures is largely unknown. To understand the regulatory mechanism of how bryophytes architect the rhizoid system responding to environmental factors, we have developed the methodology to visualize and quantitatively analyze the rhizoid system of the moss, Physcomitrium patens, in 3D. The rhizoids having a diameter of 21.3 µm on the average were visualized by refraction-contrast X-ray micro-computed tomography using coherent X-ray optics available at synchrotron radiation facility SPring-8. Three types of shape (ring-shape, line and black circle) observed in tomographic slices of specimens embedded in paraffin were confirmed to be the rhizoids by optical and electron microscopy. Comprehensive automatic segmentation of the rhizoids, which appeared in three different form types in tomograms, was tested by a method using a Canny edge detector or machine learning. The accuracy of output images was evaluated by comparing with the manually segmented ground truth images using measures such as F1 score and Intersection over Union, revealing that the automatic segmentation using machine learning was more effective than that using the Canny edge detector. Thus, machine learning-based skeletonized 3D model revealed quite dense distribution of rhizoids. We successfully visualized the moss rhizoid system in 3D for the first time.

Ultrastructure of rhizoids in the marine bryozoan Dendrobeania fruticosa (Gymnolaemata: Cheilostomata).

Bryozoans form colonies of iterated modules, termed zooids, and display varying degrees of polymorphism. Polymorphic colonies comprise autozooids (or feeding zooids) and heteromorphic zooids, among which the most common types are avicularia and kenozooids. Kenozooids differ in shape, size, and presumed function. Among this diversity, there are rhizoids, which serve to attach colonies to the substrate or to lift them above it. To date, only general data on anatomy of kenozooids at light microscopy level are available. Here, we present the first description of the ultrastructure of the holdfast-like rhizoids of the cheilostome bryozoan Dendrobeania fruticosa. The rhizoid wall is composed of a single-layered epidermis, which produces the ectocyst. The voluminous cavity is acoelomate: it has no special cellular lining, nor any signs of an extracellular matrix toward the epidermis. It is traversed by delicate branching funicular strands that originate from the pore plate. The only cells in contact with the epidermis are the cells of the funicular system and the storage cells. The pore plate between the rhizoid and autozooid includes a variable number of communication pores. Each pore is plugged with a rosette complex, which includes a cincture cell and four special cells extending through the pore. The limiting cells are absent, and the special cells are in direct contact with the funicular strands. Cell contacts between special cells are absent; moreover, there are spaces between their proximal lobes filled with a heterogeneous matrix similar to that in the lumen of the funicular strands. Such matrix is also found outside of the extracellular matrix surrounding the special cells. These findings allow us to suggest that nutrient transport most likely occurs between, rather than through, the special cells. However, further studies are needed to understand how the rosette complex functions.

MpFEW RHIZOIDS1 miRNA-Mediated Lateral Inhibition Controls Rhizoid Cell Patterning in Marchantia polymorpha.

Lateral inhibition patterns differentiated cell types among equivalent cells during development in bacteria, metazoans, and plants. Tip-growing rhizoid cells develop among flat epidermal cells in the epidermis of the early-diverging land plant Marchantia polymorpha. We show that the majority of rhizoid cells develop individually, but some develop in linear, one-dimensional groups (chains) of between 2 and 7 rhizoid cells in wild-type plants. The distribution of rhizoid cells can be accounted for within a simple cellular automata model of lateral inhibition. The model predicted that in the absence of lateral inhibition, two-dimensional rhizoid cell groups (clusters) form. These can be larger than those formed with lateral inhibition. M. polymorpha rhizoid differentiation is positively regulated by the ROOT HAIR DEFECTIVE SIX-LIKE1 (MpRSL1) basic-helix-loop-helix (bHLH) transcription factor, which is directly repressed by the FEW RHIZOIDS1 (MpFRH1) microRNA (miRNA). To test if MpFRH1 miRNA acts during lateral inhibition, we generated loss-of-function (lof) mutants without the MpFRH1 miRNA. Two-dimensional clusters of rhizoids develop in Mpfrh1lof mutants as predicted by the model for plants that lack lateral inhibition. Furthermore, two-dimensional clusters of up to 9 rhizoid cells developed in the Mpfrh1lof mutants compared to a maximum number of 7 observed in wild-type groups. The higher steady-state levels of MpRSL1 mRNA in Mpfrh1lof mutants indicate that MpFRH1-mediated lateral inhibition involves the repression of MpRSL1 activity. Together, the modeling and genetic data indicate that MpFRH1 miRNA mediates lateral inhibition by repressing MpRSL1 during pattern formation in the M. polymorpha epidermis.

Evolution of the plant body plan.

Land plants evolved about 470 million years ago or even earlier, in a biological crust-dominated terrestrial flora. The origin of land plants was probably one of the most significant events in Earth's history, which ultimately contributed to the greening of the terrestrial environment and opened up the way for the diversification of both plant and non-plant lineages. Fossil and phylogenetic evidence suggest that land plants have evolved from fresh-water charophycean algae, which were physiologically, genetically, and developmentally potentiated to make the transition to land. Since all land plants have biphasic life cycles, in contrast to the haplontic life cycle of Charophytes, the evolution of land plants was linked to the origin of a multicellular sporophytic phase. Land plants have evolved complex body plans in a way that overall complexity increased toward the tip of the land plant tree of life. Early forms were unbranched, with terminal sporangia and simple rhizoid rooting structures but without vasculature and leaves. Later on, branched forms with lateral sporangia appeared and paved the route for the evolution for indeterminacy. Finally, leaves and roots evolved to enable efficient nutrient transport to support a large plant body. The fossil record also suggests that almost all plant organs, such as leaves and roots, evolved multiple times independently over the course of land plant evolution. In this review, we summarize the current knowledge on the evolution of the land plant body plan by combining evidence of the fossil record, phylogenetics, and developmental biology.

Evolution and co-option of developmental regulatory networks in early land plants.

Land plants evolved from an ancestral alga from which they inherited developmental and physiological characters. A key innovation of land plants is a life cycle with an alternation of generations, with both haploid gametophyte and diploid sporophyte generations having complex multicellular bodies. The origins of the developmental genetic programs patterning these bodies, whether inherited from an algal ancestor or evolved de novo, and whether programs were co-opted between generations, are largely open questions. We first provide a framework for land plant evolution and co-option of developmental regulatory pathways and then examine two cases in more detail.

A soil bacterium alters sex determination and rhizoid development in gametophytes of the fern Ceratopteris richardii.

Gametophytes of the fern Ceratopteris richardii develop into either hermaphrodites or males. As hermaphrodites develop, they secrete antheridiogen, or ACE, into the environment, inducing male development in undifferentiated gametophytes. Hermaphrodites are composed of archegonia, antheridia, rhizoids and a notch meristem, while males consist of antheridia and rhizoids. Much of the research on sexual and morphological development concerns gametophytes grown in sterile environments. Using biochemical and molecular techniques we identify a soil bacterium and explore its effects on sexual and rhizoid development. Hermaphrodite and male gametophytes were exposed to this bacterium and the effects on sexual development, rhizoid length and rhizoid number were explored. The bacterium was identified as a pseudomonad, Pseudomonas nitroreducens. Gametophytes grown in the presence of the pseudomonad were more likely to develop into hermaphrodites across all gametophyte densities. Across all gametophyte sizes, hermaphrodites had rhizoids that were 2.95× longer in the presence of the pseudomonad while males had rhizoids that were 2.72× longer in the presence of the pseudomonad. Both hermaphrodite and male gametophytes developed fewer rhizoids in the presence of the pseudomonad. Control hermaphrodites produced 1.23× more rhizoids across all gametophyte sizes. For male gametophytes grown in the absence of the pseudomonad, the rate of increase in the number of rhizoids was greater with increasing size in the control than the rate of increase in males grown in the presence of the pseudomonad. The pseudomonad may be acting on gametophyte sexual development via several potential mechanisms: degradation of ACE, changes in nutrient availability or phytohormone production. The pseudomonad may also increase rhizoid number through production of phytohormones or changes in nutrient availability.

Evolution of Cell Wall Polymers in Tip-Growing Land Plant Gametophytes: Composition, Distribution, Functional Aspects and Their Remodeling.

During evolution of land plants, the first colonizing species presented leafy-dominant gametophytes, found in non-vascular plants (bryophytes). Today, bryophytes include liverworts, mosses, and hornworts. In the first seedless vascular plants (lycophytes), the sporophytic stage of life started to be predominant. In the seed producing plants, gymnosperms and angiosperms , the gametophytic stage is restricted to reproduction. In mosses and ferns, the haploid spores germinate and form a protonema, which develops into a leafy gametophyte producing rhizoids for anchorage, water and nutrient uptakes. The basal gymnosperms (cycads and Ginkgo) reproduce by zooidogamy. Their pollen grains develop a multi-branched pollen tube that penetrates the nucellus and releases flagellated sperm cells that swim to the egg cell. The pollen grain of other gymnosperms (conifers and gnetophytes) as well as angiosperms germinates and produces a pollen tube that directly delivers the sperm cells to the ovule (siphonogamy). These different gametophytes, which are short or long-lived structures, share a common tip-growing mode of cell expansion. Tip-growth requires a massive cell wall deposition to promote cell elongation, but also a tight spatial and temporal control of the cell wall remodeling in order to modulate the mechanical properties of the cell wall. The growth rate of these cells is very variable depending on the structure and the species, ranging from very slow (protonemata, rhizoids, and some gymnosperm pollen tubes), to a slow to fast-growth in other gymnosperms and angiosperms. In addition, the structural diversity of the female counterparts in angiosperms (dry, semi-dry vs wet stigmas, short vs long, solid vs hollow styles) will impact the speed and efficiency of sperm delivery. As the evolution and diversity of the cell wall polysaccharides accompanied the diversification of cell wall structural proteins and remodeling enzymes, this review focuses on our current knowledge on the biochemistry, the distribution and remodeling of the main cell wall polymers (including cellulose, hemicelluloses, pectins, callose, arabinogalactan-proteins and extensins), during the tip-expansion of gametophytes from bryophytes, pteridophytes (lycophytes and monilophytes), gymnosperms and the monocot and eudicot angiosperms.

An Evolutionarily Conserved Receptor-like Kinases Signaling Module Controls Cell Wall Integrity During Tip Growth.

Rooting cells and pollen tubes-key adaptative innovations that evolved during the colonization and subsequent radiation of plants on land-expand by tip growth. Tip growth relies on a tight coordination between the protoplast growth and the synthesis/remodeling of the external cell wall. In root hairs and pollen tubes of the seed plant Arabidopsis thaliana, cell wall integrity (CWI) mechanisms monitor this coordination through the Malectin-like receptor kinases (MLRs), such as AtANXUR1 and AtFERONIA, that act upstream of the AtMARIS PTI1-like kinase. Here, we show that rhizoid growth in the early diverging plant, Marchantia polymorpha, is also controlled by an MLR and PTI1-like signaling module. Rhizoids, root hairs, and pollen tubes respond similarly to disruption of MLR and PTI1-like encoding genes. Thus, the MLR and PTI1-like signaling module that controls CWI during tip growth is conserved between M. polymorpha and A. thaliana, suggesting that it was active in the common ancestor of land plants.

Antioxidant and Cytoprotective Activities of Enzymatic Extracts from Rhizoid of Laminaria japonica.

Rhizoid of Laminaria japonica was hydrolyzed with proteases and carbohydrases to obtain antioxidant materials. Oxygen radical absorbance capacity (ORAC) of the enzymatic extracts was evaluated and the Protamex extract (PE) exhibited the highest ORAC value. PE also potently scavenged 2,2-diphenyl-1-picrylhydrazyl radical, 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulphonic) acid cation radical, and hydrogen peroxide (H2O2) and had good reducing power. PE inhibited hydroxyl radical-induced DNA scission by measuring the conversion of supercoiled pBR322 plasmid DNA to the open circular form. The cytoprotective effect of PE against H2O2-induced hepatic cell damage was also investigated. PE showed a dose-dependent cytoprotective effect in cultured hepatocytes by inhibiting intracellular reactive oxygen species scavenging activity. In addition, PE up-regulated the expression of heme oxygenase-1, which is a cytoprotective enzyme, by activating translocation of nuclear factor-erythroid 2-related factor 2. Taken together, the enzymatic extract of rhizoid of L. japonica, particularly PE, may be useful for antioxidant additives.

The Mechanism Forming the Cell Surface of Tip-Growing Rooting Cells Is Conserved among Land Plants.

To discover mechanisms that controlled the growth of the rooting system in the earliest land plants, we identified genes that control the development of rhizoids in the liverwort Marchantia polymorpha. 336,000 T-DNA transformed lines were screened for mutants with defects in rhizoid growth, and a de novo genome assembly was generated to identify the mutant genes. We report the identification of 33 genes required for rhizoid growth, of which 6 had not previously been functionally characterized in green plants. We demonstrate that members of the same orthogroup are active in cell wall synthesis, cell wall integrity sensing, and vesicle trafficking during M. polymorpha rhizoid and Arabidopsis thaliana root hair growth. This indicates that the mechanism for constructing the cell surface of tip-growing rooting cells is conserved among land plants and was active in the earliest land plants that existed sometime more than 470 million years ago [1, 2].

Structure and function of CrACA1, the major PM-type Ca2+-ATPase, expressed at the peak of the gravity-directed trans-cell calcium current in spores of the fern Ceratopteris richardii.

Spores of the fern Ceratopteris richardii have proven to be a valuable single-cell system for studying gravity responses. The earliest cellular change directed by gravity in these cells is a trans-cell calcium current, which peaks near 10 h after the spores are induced to germinate. This current is needed for gravity-directed axis alignment, and its peak is coincident with the time period when gravity polarises the direction of subsequent nuclear migration and rhizoid growth. Transcriptomic analysis of genes expressed at the 10-h time point revealed several that encode proteins likely to be key components that either drive the current or regulate it. Notable among these is a plasma membrane (PM)-type Ca(2+) ATPase, CrACA1, whose activity pumping Ca(2+) out of cells is regulated by gravity. This report provides an initial characterisation of the structure and expression of this protein, and demonstrates its heterologous function complementing the K616 mutant of yeast, which is deficient in PM-type Ca(2+) pump activity. Gravity-induced changes in the trans-cell Ca(2+) current occur within seconds, a result consistent with the hypothesis that the force of gravity can rapidly alter the post-translational state of the channels and pumps that drive this current across spore cells. This report identifies a transporter likely to be a key driver of the current, CrACA1, and characterises the role of this protein in early germination and gravity-driven polarity fixation through analysis of expression levels, functional complementation and pharmacological treatments. These data, along with newly available transcriptomic data obtained at the 10-h time point, indicate that CrACA1 is present, functional and likely a major contributing component of the trans-cell Ca(2+) efflux. CrACA1 is not necessary for polar axis alignment, but pharmacological perturbations of it disrupt rhizoid development. These data support and help refine the post-translational modification model for gravity responses.