Protoplasts: a useful research system for plant cell biology, especially dedifferentiation

As protoplasts have the characteristics of no cell walls, rapid population growth, and synchronicity, they are useful tools for research in many fields, especially cellular biology (Table 1). This article is an overview that focuses on the application of protoplasts to investigate the mechanisms of dedifferentiation, including changes in hormone signals, epigenetic changes, and organelle distribution during the dedifferentiation process. The article also emphasizes the wide range of uses for protoplasts in studying protein positions and signaling during different stresses. The examples provided help to show that protoplast systems, for example the mesophyll protoplast system of Arabidopsis, represent promising tools for studying developmental biology. Meanwhile, specific analysis of protoplast, which comes from different tissue, has specific advantages and limitations (Table 2), and it can provide recommendations to use this system.     

An optimized B lymphocyte stimulator (BLyS) antagonist peptide inhibits the interaction of BLyS with BCMA

B lymphocyte stimulator (BLyS) antagonists are new therapeutic reagents for treating the autoimmune diseases. Peptibodies can inhibit the bioactivity of BLyS, the same as other BLyS antagonists: decoyed BLyS receptors and anti-BLyS antibodies. In this study, a new optimized BLyS antagonist peptide was designed according to our previous work by the computer-aided homology modeling. Competitive ELISA showed that the peptide at 100 μg/ml could inhibit 54 % of the BCMA-Fc binding to BLyS. To maintain its stability and spatial conformation, the peptide was fused to human IgG1 Fc to form a peptide-Fc fusion protein—a novel peptibody by gene engineering. ELISA indicated that the peptibody could bind with BLyS in dosage-dependent manner as BCMA-Fc did. This study highlights the possibility of designing and optimizing BLyS antagonist peptides with high biopotency by the computer-aided design. Thus, these peptides could neutralize BLyS activity and be potential antagonists to treat autoimmune diseases related with BLyS overexpression.     

Genetic manipulation revealing an unusual N-terminal region in a stand-alone non-ribosomal peptide synthetase involved in the biosynthesis of ramoplanins

Ramoplanins produced by Actinoplanes are new structural class of lipopeptide and are currently in phase III clinical trials for the prevention of vancomycin-resistant enterococcal infections. The depsipeptide structures of ramoplanins are synthesized by non-ribosomal peptide synthetases (NRPS). Romo-orf17, a stand-alone NRPS, is responsible for the recruitment of Thr into the linear NRPS pathways for which the corresponding adenylation domain is absent. Here, systematical gene inactivation and complementation have been carried out in a Actinoplanes sp. using homologous recombination and site-specific integration methods. A hybrid gene coding for the N-terminal region of the stand-alone NRPS and the A-PCP domains of a heterologous NRPS restored production of ramoplanins. The results elucidate the unusual N-terminal region which is essential for the biosynthesis of ramoplanins.     

Arbuscular mycorrhizal fungi-enhanced resistance against Phytophthora sojae infection on soybean leaves is mediated by a network involving hydrogen peroxide, jasmonic acid, and the metabolism of carbon and nitrogen

The arbuscular mycorrhizal fungi (AMF) enhance the resistance to pathogen infection in host plant. However, it is unclear how the AMF are involved in the systemic acquired resistance of host plant against pathogen. Here, an experiment was carried out to clarify the role of the AMF in soybean’s defense against the infection from pathogen Phytophthora sojae. It was found that the AMF contributed to the resistance of soybean against Phytophthora sojae by the release of hydrogen peroxide and by the accumulation of jasmonic acid in response to pathogenic invasion. Furthermore, the trade of nitrogen (N) from the fungus for carbon from the host was accelerated in the AM symbiosis in the defense reaction, which was indicated by the increased soluble sugar level, NO content and enzyme activities involved in N metabolism in the AM symbiosis.     

CmPEX6, a Gene Involved in Peroxisome Biogenesis,Is Essential for Parasitism and Conidiation by the Sclerotial Parasite Coniothyrium minitans

Coniothyrium minitans is a sclerotial parasite of the plant-pathogenic fungus Sclerotinia sclerotiorum, and conidial production and parasitism are two important aspects for commercialization of this biological control agent. To understand the mechanism of conidiation and parasitism at the molecular level, we constructed a transfer DNA (tDNA) insertional library with the wild-type strain ZS-1. A conidiation-deficient mutant, ZS-1TN22803, was uncovered through screening of this library. This mutant could produce pycnidia on potato dextrose agar (PDA), but most were immature and did not bear conidia. Moreover, this mutant lost the ability to parasitize or rot the sclerotia of S. sclerotiorum. Analysis of the tDNA flanking sequences revealed that a peroxisome biogenesis factor 6 (PEX6) homolog of Saccharomyces cerevisiae, named CmPEX6, was disrupted by the tDNA insertion in this mutant. Targeted gene replacement and gene complementation tests confirmed that a null mutation of CmPEX6 was responsible for the phenotype of ZS-1TN22803. Further analysis showed that both ZS-1TN22803 and the targeted replacement mutants could not grow on PDA medium containing oleic acid, and they produced much less nitric oxide (NO) and hydrogen peroxide (H₂O₂) than wild-type strain ZS-1. The conidiation of ZS-1TN22803 was partially restored by adding acetyl-CoA or glyoxylic acid to the growth media. Our results suggest that fatty acid β-oxidation, reactive oxygen and nitrogen species, and possibly other unknown pathways in peroxisomes are involved in conidiation and parasitism by C. minitans.     

Responses of Active Ammonia Oxidizers to Eutrophication and Oxygen Statuses in Taihu Freshwater Sediments

Abstract

Here, we employed DNA-based stable isotope probing (SIP) and molecular biology methods to investigate active ammonia oxidizer communities in suboxic sediments (0 to –2?cm) at the micromolar oxygen level and layers (–2 to –5?cm) at nanomolar oxygen concentrations from meso-eutrophic and light-eutrophic locations in Taihu Lake. The results revealed that ammonia-oxidizing archaea (AOA) were less active in the anoxic layer of meso-eutrophic sites, while ammonia-oxidizing bacteria (AOB) were less active in suboxic sediments of light-eutrophic sites after 8?weeks of incubation. The active AOA in the meso- and light-eutrophic sediments belonged to the Nitrosopumilus, Nitrosotalea, and Nitrososphaera clusters and the Nitrosopumilus and Nitrososphaera clusters, respectively, with Nitrosopumilus cluster as the predominant AOA, which took up a higher ratio in the light-eutrophic and suboxic layers than their counterparts. The advantageous active AOB were numerically predominated by the Nitrosomonas cluster in the suboxic layers, and the Nitrosospira cluster in the anoxic layers, respectively, both of which were distributed in diverse frequencies in different eutrophication statuses. The role and community composition diversities of active ammonia oxidizers in freshwater sediments were attributed to the different eutrophication (including nitrogen and organic carbon content) and oxygen statuses.     

The Mechanism,Thermodynamic and Kinetic Characteristics of the Microbial Reduction of Goethite Mediated by Anthraquinone-2-Sulfonate

The microbial reduction process of goethite by Shewanella decolorationis S12 was evaluated. The results showed the electron shuttle, anthraquinone-2-sulfonate (AQS), could enhance the microbial reduction. The thermodynamic and kinetic characteristics of goethite reduction by microorganisms were influenced by AQS, concentrations of iron oxide, and electron donor. Transformation between oxidized and reduced species of the electron shuttle during the microbial reduction could be newly noticed. Two interactive steps, biotic and abiotic, were involved in the microbial reduction of Fe (III) oxide mediated by electron shuttle.     

Arabidopsis CROLIN1, a Novel Plant Actin-binding Protein,Functions in Cross-linking and Stabilizing Actin Filaments

Higher order actin filament structures are necessary for cytoplasmic streaming, organelle movement, and other physiological processes. However, the mechanism by which the higher order cytoskeleton is formed in plants remains unknown. In this study, we identified a novel actin-cross-linking protein family (named CROLIN) that is well conserved only in the plant kingdom. There are six isovariants of CROLIN in the Arabidopsis genome, with CROLIN1 specifically expressed in pollen. In vitro biochemical analyses showed that CROLIN1 is a novel actin-cross-linking protein with binding and stabilizing activities. Remarkably, CROLIN1 can cross-link actin bundles into actin networks. CROLIN1 loss of function induces pollen germination and pollen tube growth hypersensitive to latrunculin B. All of these results demonstrate that CROLIN1 may play an important role in stabilizing and remodeling actin filaments by binding to and cross-linking actin filaments.