Azolla--a model organism for plant genomic studies

The aquatic ferns of the genus Azolla are nitrogen-fixing plants that have great potentials in agricultural production and environmental conservation. Azolla in many aspects is qualified to serve as a model organism for genomic studies because of its importance in agriculture, its unique position in plant evolution, its symbiotic relationship with the N2-fixing cyanobacterium, Anabaena azollae, and its moderate-sized genome. The goals of this genome project are not only to understand the biology of the Azolla genome to promote its applications in biological research and agriculture practice but also to gain critical insights about evolution of plant genomes. Together with the strategic and technical improvement as well as cost reduction of DNA sequencing, the deciphering of their genetic code is imminent.     

Targeted Gene Knockouts Reveal Overlapping Functions of the Five Physcomitrella patens FtsZ Isoforms in Chloroplast Division,Chloroplast Shaping,Cell Patterning,Plant Development,and Gravity Sensing

Chloroplasts and bacterial cells divide by binary fission. The key protein in this constriction division is FtsZ, a self-assembling GTPase similar to eukaryotic tubulin. In prokaryotes, FtsZ is almost always encoded by a single gene, whereas plants harbor several nuclear-encoded FtsZ homologs. In seed plants, these proteins group in two families and all are exclusively imported into plastids. In contrast, the basal land plant Physcomitrella patens, a moss, encodes a third FtsZ family with one member. This protein is dually targeted to the plastids and to the cytosol. Here, we report on the targeted gene disruption of all ftsZ genes in R patens. Subsequent analysis of single and double knockout mutants revealed a complex interaction of the different FtsZ isoforms not only in plastid division, but also in chloroplast shaping, cell patterning, plant development, and gravity sensing. These results support the concept of a plastoskeleton and its functional integration into the cytoskeleton, at least in the moss R patens.     

Comparative Genomic Study of the Thioredoxin Family in Photosynthetic Organisms with Emphasis on Populus trichocarpa

The recent genome sequencing of Populus trichocarpa and Vitis vinifera, two models of woody plants, of Sorghum bicolor, a model of monocot using C4 metabolism, and of the moss Physcomitrella patens, together with the availability of photosynthetic organism genomes allows performance of a comparative genomic study with organisms having different ways of life, reproduction modes, biological traits, and physiologies. Thioredoxins （Trxs） are small ubiq- uitous proteins involved in the reduction of disulfide bridges in a variety of target enzymes present in all sub-cellular compartments and involved in many biochemical reactions. The genes coding for these enzymes have been identified in these newly sequenced genomes and annotated. The gene content, organization and distribution were compared to other photosynthetic organisms, leading to a refined classification. This analysis revealed that higher plants and bryo- phytes have a more complex family compared to algae and cyanobacteria and to non-photosynthetic organisms, since poplar exhibits 49 genes coding for typical and atypical thioredoxins and thioredoxin reductases, namely one-third more than monocots such as Oryza sativa and S. bicolor. The higher number of Trxs in poplar is partially explained by gene duplication in the Trx m, h, and nucleoredoxin classes. Particular attention was paid to poplar genes with emphasis on Trx-like classes called Clot, thioredoxin-like, thioredoxins of the lilium type and nucleoredoxins, which were not described in depth in previous genomic studies.     

Clues to the signals for chloroplast photo-relocation from the lifetimes of accumulation and avoidance responses

Chloroplast photo-relocation movement is crucial for plant survival; however, the mechanism of this phenomenon is still poorly understood. Especially, the signal that goes from photoreceptor to chloroplast is unknown, although the photoreceptors(phototropin 1 and 2) have been identified and an actin structure(chloroplast actin filaments) has been characterized that is specific for chloroplast movement. Here,in gametophytes of the fern Adiantum capillus-veneris,gametophores of the moss Physcomiterella patens, and leaves of the seed plant Arabidopsis thaliana, we sought to characterize the signaling system by measuring the lifetime of the induced response. Chloroplast movements were induced by microbeam irradiation with high-intensity blue light and recorded. The lifetime of the avoidance state was measured as a lag time between switching off the beam and the loss of avoidance behavior, and that of the accumulation state was measured as the duration of accumulation behavior following the extinction of the beam. The lifetime for the avoidance response state is approximately 3–4 min and that for the accumulation response is 19–28 min. These data suggest that the two responses are based on distinct signals.     

Recycling Isolation of Plant DNA,A Novel Method

DNA is one of the most basic and essential genetic materials in the field of molecular biology.To date,isolation of sufficient and good-quality DNA is still a challenge for many plant species,though various DNA extraction methods have been published.In the present paper,a recycling DNA extraction method was proposed.The key step of this method was that a single plant tissue sample was recycled for DNA extraction for up to four times,and correspondingly four DNA precipitations(termed as the 1st,2nd,3rd and 4th DNA sample, respectively) were conducted.This recycling step was integrated into the conventional CTAB DNA extraction method to establish a recycling CTAB method.This modified CTAB method was tested in eight plant species,wheat,sorghum,barley,corn,rice,Brachypodium distachyon,Miscanthus sinensis and tung tree.The results showed that high-yield and good-quality DNA samples could be obtained by using this new method in all the eight plant species.The DNA samples were good templates for PCR amplification of both ISSR and SSR markers.The recycling method can be used in multiple plant species and can be integrated with multiple conventional DNA isolation methods,and thus is an effective and universal DNA isolation method.     

Induction of apoptosis in purified animal and plant nuclei by Xenopus egg extracts

We have developed a cell-free system that can trigger the nuclei purified from mouse liver and suspensioncultured carrot cells to undergo apoptosis as defined by the formation of apoptotic bodies and nucleosomal DNA fragments.The effects of different divalent cations and cycloheximide on DNA cleavage in this system were assessed.The fact that nuclei of plant cells can be induced to undergo apoptosis in a cell-free animal system suggests that animals and plants share a common signal transduction pathway triggering in the initiation stage of apoptosis.     

Flavonoids:New Roles for Old Molecules

Flavonoids are ubiquitous in the plant kingdom and have many diverse functions including defense, UV protection, auxin transport inhibition, allelopathy, and flower coloring. Interestingly, these compounds also have considerable biological activity in plant, animal and bacterial systems-such broad activity is accomplished by few compounds. Yet, for all the research over the last three decades, many of the cellular targets of these secondary metabolites are unknown. The many mutants available in model plant ...     

Cellulose synthase ‘class specific regions’ are intrinsically disordered and functionally undifferentiated

Cellulose synthases(CESAs) are glycosyltransferases that catalyze formation of cellulose microfibrils in plant cell walls.Seed plant CESA isoforms cluster in six phylogenetic clades, whose non-interchangeable members play distinct roles within cellulose synthesis complexes(CSCs).A ‘class specific region’(CSR), with higher sequence similarity within versus between functional CESA classes, has been suggested to contribute to specific activities or interactions of different isoforms.We investigated CESA isoform specificity in the moss,Physcomitrella patens(Hedw.) B.S.G.to gain evolutionary insights into CESA structure/function relationships.Like seed plants, P.patens has oligomeric rosette-type CSCs, but the Pp CESAs diverged independently and form a separate CESA clade.We showed that P.patens has two functionally distinct CESAs classes, based on the ability to complement the gametophore-negative phenotype of a ppcesa5 knockout line.Thus, non-interchangeable CESA classes evolved separately in mosses and seed plants.However, testing of chimeric moss CESA genes for complementation demonstrated that functional classspecificity is not determined by the CSR.Sequence analysis and computational modeling showed that the CSR is intrinsically disordered and contains predicted molecular recognition features, consistent with a possible role in CESA oligomerization and explaining the evolution of class-specific sequences without selection for class-specific function.     

Heterologous expression of Hp BHY and Cr BKT increases heat tolerance in Physcomitrella patens

Heat stress can restrict plant growth,development,and crop yield.As essential plant antioxidants,carotenoids play significant roles in plant stress resistance.b-carotene hydroxylase(BHY)and b-carotene ketolase(BKT),which catalyze the conversions of b-carotene to zeaxanthin and b-carotene to canthaxanthin,respectively,are key enzymes in the carotenoid biosynthetic pathway,but little is known about their potential functions in stress resistance.Here,we investigated the roles of b-carotene hydroxylase and b-carotene ketolase during heat stress in Physcomitrella patens through expressing a b-carotene ketolase gene from Chlamydomonas reinhardtii(Cr BKT)and a b-carotene hydroxylase gene from Haematococcus pluvialis(Hp BHY)in the moss P.patens.In transgenic moss expressing these genes,carotenoids content increased(especially lutein content),and heat stress tolerance increased,with reduced leafy tissue necrosis.To investigate the mechanism of this heat stress resistance,we measured various physiological indicators and found a lower malondialdehyde level,higher peroxidase and superoxide dismutase activities,and higher endogenous abscisic acid and salicylate content in the transgenic plants in response to high-temperature stress.These results demonstrate that Cr BKT and Hp BHY increase plant heat stress resistance through the antioxidant and damage repair metabolism,which is related to abscisic acid and salicylate signaling.     

Evolutionary crossroads in developmental biology: Physcomitrella patens

The moss Physcomitrella patens has recently emerged as a powerful genetically tractable model plant system. As a member of the bryophytes, P. patens provides a unique opportunity to study the evolution of a myriad of plant traits, such as polarized cell growth, gametophyte-to-sporophyte transitions, and sperm-to-pollen transition. The availability of a complete genome sequence, together with the ability to perform gene targeting efficiently in P. patens has spurred a flurry of elegant reverse genetic studies in this plant model that address a variety of key questions in plant developmental biology.     

植物分子生物学研究极具前景的模式系统--小立碗藓

小立碗藓作为植物分子生物学研究极具前景的模式系统已日益受到人们的重视,它的生活史周期短,易于培养,转基因植株易于分析,核基因组容易和外源DNA发生同源重组,这些特点使它成为研究基因功能的良好材料.一些成功的基因敲除和基因破坏已经在小立碗藓中实现,这些基因的功能也通过小立碗藓转化植株的特点得以证实.小立碗藓标签突变文库已经建立,其应用为小立碗藓基因的进一步研究打下了基础.关于小立碗藓的ESTs数据库已经建立,已有67 000条ESTs信息.     

植物分子生物学研究极具前景的模式系统——小立碗藓

小立碗藓作为植物分子生物学研究极具前景的模式系统已日益受到人们的重视,它的生活史周期短,易于培养,转基因植株易于分析,核基因组容易和外源DNA 发生同源重组,这些特点使它成为研究基因功能的良好材料。一些成功的基因敲除和基因破坏已经在小立碗藓中实现,这些基因的功能也通过小立碗藓转化植株的特点得以证实。小立碗藓标签突变文库已经建立,其应用为小立碗藓基因的进一步研究打下了基础。关于小立碗藓的ESTs 数据库已经建立,已有67 000 条ESTs 信息。     

有前景的模式植物小立碗藓的研究新进展

小立碗藓是在分子生物学研究方面有广阔应用前景的模式植物。该文主要综述了有关小立碗藓在功能基因组学、进化和适应性及植物生理等方面最新的研究进展。     

Exploration and identification of Physcomitrella patens moss peptides

In the current study the isolation and identification of Physcomitrella patens (Hedw.) B.S.G. moss peptides are described. Physcomitrella patens moss is actively used in recent years as a model organism to study the biology of plants. Protoplasts, protonemata and gametophores of the moss are demonstrated for the first time to contain diverse small peptides. From gametophores was isolated and identified 58 peptides that are fragments of 14 proteins, and from protonemata - 49 peptides, fragments of 15 proteins. It was found that the protonemata and gametophores Ph. patens, which are the successive stages of development of this plant, significantly different from each other as a peptide composition and the spectrum of the precursor protein of identified peptides. Isolation of protoplasts of the enzymatic destruction of cell wall protonemata accompanied by massive degradation of intracellular proteins, many of whom are proteins of photosynthesis, which is a characteristic response of plants to stress the impact of environmental factors. A total of moss protoplasts were isolated and identified 323 peptides that are fragments of 79 proteins.     

Efficient gene targeting in the moss Physcomitrella patens

The moss Physcomitrella patens is used as a genetic model system to study plant development, taking advantage of the fact that the haploid gametophyte dominates in its life cycle. Transformation experiments designed to target three single-copy genomic loci were performed to determine the efficiency of gene targeting in this plant. Mean transformation rates were 10-fold higher with the targeting vectors and molecular evidence for the integration of exogenous DNA into each targeted locus by homologous recombination is provided. The efficiency of gene targeting determined in these experiments is above 90%, which is in the range of that observed in yeast and several orders of magnitude higher than previous reports of gene targeting in plants. Thus, gene knock-out and allele replacement approaches are directly accessible to study plant development in the moss Physcomitrella patens . Moreover, efficient gene targeting has so far only been observed in lower eukaryotes such as protozoa, yeasts and filamentous fungi, and, as shown here the first example from the plant kingdom is a haplobiontic moss. This suggests a possible correlation between efficient gene targeting and haplo-phase in eukaryotes.     

Cloning of the PpMSH-2 cDNA of Physcomitrella patens, a moss in which gene targeting by homologous recombination occurs at high frequency

In the moss Physcomitrella patens integrative transformants from homologous recombination are obtained at an efficiency comparable to that found for yeast. This property, unique in the plant kingdom, allows the knockout of specific genes. It also makes the moss a convenient model to study the regulation of homologous recombination in plants. We used degenerate oligonucleotides designed from AtMSH2 from Arabidopsis thaliana and other known MutS homologues to isolate the P. patens MSH2 (PpMSH2) cDNA. The deduced sequence of the PpMSH2 protein is respectively 60.8% and 59.6% identical to the maize and A. thaliana MSH2. Phylogenic studies show that PpMSH2 is closely related to the group of plant MSH2 proteins. Southern analysis reveals that the gene exists as a single copy in the P. patens genome.     

Moss systems biology en route: phytohormones in Physcomitrella development

The moss Physcomitrella patens has become a powerful model system in modern plant biology. Highly standardized cell culture techniques, as well as the necessary tools for computational biology, functional genomics and proteomics have been established. Large EST collections are available and the complete moss genome will be released soon. A simple body plan and the small number of different cell types in Physcomitrella facilitate the study of developmental processes. In the filamentous juvenile moss tissue, developmental decisions rely on the differentiation of single cells. Developmental steps are controlled by distinct phytohormones and integration of environmental signals. Especially the phytohormones auxin, cytokinin, and abscisic acid have distinct effects on early moss development. In this article, we review current knowledge about phytohormone influences on early moss development in an attempt to fully unravel the complex regulatory signal transduction networks underlying the developmental decisions of single plant cells in a holistic systems biology approach.