Plant Cell Wall Matrix Polysaccharide Biosynthesis

The wall of an expanding plant cell consists primarily of cellulose microfibrils embedded in a matrix of hemicellulosic and pectic polysaccharides along with small amounts of structural and enzymatic proteins. Matrix polysaccharides are synthesized in the Golgi and exported to the cell wall by exocytosis, where they intercalate among cellulose microfibrils, which are made at the plasma membrane and directly deposited into the cell wall. Involvement of Golgi glucan synthesis in auxin-induced cell expansion has long been recognized; however, only recently have the genes corresponding to glucan synthases been identified. Biochemical purification was unsuccessful because of the labile nature and very low abundance of these enzymes. Mutational genetics also proved fruitless. Expression of candidate genes identified through gene expression profiling or comparative genomics in heterologous systems followed by functional characterization has been relatively successful. Several genes from the cellulose synthase-like （Csl） family have been found to be involved in the synthesis of various hemicellulosic glycans. The usefulness of this approach, however, is limited to those enzymes that probably do not form complexes consisting of unrelated proteins. Nonconventional approaches will continue to incrementally unravel the mechanisms of Golgi polysaccharide biosynthesis.     

培育具有安全选择标记或无选择标记的转基因植物

转基因植物中选择标记的安全性已成为植物基因工程研究的热点之一。从两个方面可以解决转基因植物中的选择标记问题。一是选用安全的正向选择标记,主要是与糖代谢和激素代谢相关的基因。二是构建能去除选择标记基因的转化系统,主要有共转化系统、双T-DNA边界载体系统、位点特异性重组系统和转座子系统等。这些植物基因工程的方法将有助于培育安全的转基因植物。 Abstract:The bio-safety of selective markers in transgenic plants has been a hot spot in the field of plant genetic engineering.To solve the problem of selective markers in the transgenic plants,two means of producing transgenic plants have been developed.One is the utilization of bio-safe positive selective markers which are genes mainly related to metabolism of auxins and carbohydrates.The other is the establishment of transformation systems allowing marker genes to be eliminated from the transgenic plants,which include co-ransformation,double T-DNA border vectors,site-specific recombination and transposition.All these approaches of plant genetic engineering will benefit breeding transgenic plants with bio-safety.     

Analyses of Sexual Reproductive Success in Transgenic and/or Mutant Plants

The pistil, the female reproductive organ of plants, is a key player in the success of sexual plant reproduction. Ultimately, the production of fruits and seeds depends on the proper pistil development and function. Therefore, the identification and characterization of pistil expressed genes is essential for a better understanding and manipulation of the plant reproduction process. For studying the function of pistil expressed genes, transgenic and/or mutant plants for the genes of interest are used. The present article provides a review of methods already exploited to analyze sexual reproductive success. We intend to supply useful information and to guide future experiments in the study of genes affecting pistil development and function.     

Recent Advances in Cloning and Characterization of Disease Resistance Genes in Rice

Rice diseases caused by fungi, bacteria and viruses are one of the major constraints for sustainable rice （Oryza sativa L.） production worldwide. The use of resistant cultivars is considered the most economical and effective method to control rice diseases. In the last decade, a dozen resistance genes against the fungal pathogen Magnaporthe grisea and the bacterial pathogen Xanthomonas oryzae pv. oryzae have been cloned. Approximately half of them encode nuclear binding site （NBS） and leucine rich repeat （LRR）-containing proteins, the most common type of cloned plant resistance genes. Interestingly, four of them encode novel proteins which have not been identified in other plant species, suggesting that unique mechanisms might be involved in rice defense responses. This review summarizes the recent advances in cloning and characterization of disease resistance genes in rice and presents future perspectives for in-depth molecular analysis of the function and evolution of rice resistance genes and their interaction with avirulence genes in pathogens.     

植物碱性亮氨酸拉链(bZIP)蛋白的研究进展(二)———DNA结合特性、基因表达、功能及应用

植物碱性亮氨酸拉链(bZIP)蛋白在高等植物基因表达与调控中起重要作用。本文介绍了植物bZIP蛋白与DNA结合特性,探讨了它们的基因表达和功能,综述了它们在分子生物学和基因工程研究中的应用。 Abstract:Plant basic leucine zipper (bZIP) proteins play an important role in the expression and regulation of higher plant genes.The DNA-binding properties of plant bZIP protein are introduced first in this article.Then their expression and function are discussed.Finally,their application to the studies of molecular biology and genetic engineering is reviewed.     

植物基因工程表达载体的改进和优化策略

外源基因在转基因植物中表达效率低一直是令相关研究者困扰的一个问题.转基因植物的生物安全性最近在全球范围内开始引起世人的担忧.本文简要介绍了近年来在植物表达载体构建方面所采用的一些新策略,这些策略有助于增强外源基因的表达水平、提高生物工程体的安全性。 Abstract:The low expression level of foreign genes in transgenic plants is a puzzling question for researchers in plant genetic engineering.The biosafety concern is now causing a growing public wariness of transgenic plants around the world.This article reviews the new strategies currently used in construction of plant expression vector for plant genetic engineering.These strategies are important for obtaining better expression of foreign genes and developing safer transgenic plants.     

Plant Terpenoids： Biosynthesis and Ecological Functions

Among plant secondary metabolites terpenolds are a structurally most diverse group; they function as phytoalexins In plant direct defense, or as signals In Indirect defense responses which involves herbivores and their natural enemies. In recent years, more and more attention has been paid to the Investigation of the ecological role of plant terpenolds. The biosynthesis pathways of monoterpenes, sesquiterpenes, and diterpenes Include the synthesis of C5 precursor isopentenyl diphosphate （IPP） and Its allylic isomer dlmethylallyl dlphosphate （DMAPP）, the synthesis of the immediate diphosphate precursors, and the formation of the diverse terpenoids. Terpene synthases （TPSs） play a key role In volatile terpene synthesis. By expression of the TPS genes, significant achievements have been made on metabolic engineering to Increase terpenoid production. This review mainly summarizes the recent research progress In elucidating the ecological role of terpenoids and characterization of the enzymes Involved in the terpenold biosynthesis. Spatial and temporal regulations of terpenoids metabolism are also discussed.     

植物纤维素生物合成及其相关酶类

纤维素是由成千上万个D-葡萄糖分子通过β-1,4糖苷键连接的具有一定立体构象的链状聚合物,其葡萄糖残基约为2000～25000个。它是细胞壁的主要组成成分之一。植物,大多数藻类,一些细菌和真菌甚至有些动物都能合成纤维素。它作为世界上最丰富的,具有巨大商业价值的生物多聚体,几十年来一直受到人们的重视,成为人们的研究热点。尽管如此,这方面的研究仍比较滞后,人们对纤维素生物合成途径及其相关酶类还是知之甚少。近几年来,随着基因组学的发展,关于纤维素的生物合成及相关基因表达调控的研究也成绩斐然,现就高等植物纤维素生物合成途径及其相关的纤维素合成酶的最新研究进展作一介绍。     

细菌纤维素合酶的基本特性及作用机理

细菌纤维素是一种天然的生物质高分子聚合物。相较于植物纤维素,其具有更高的纯度和优异的力学性能。有望作为一种绿色的新型高分子材料被广泛应用。细菌纤维素合酶作为合成细菌纤维素的关键酶,其主导细菌纤维素的合成过程。因此,对其合成机理的探索有助于实现细菌纤维素大量生产和广泛应用。本文从细菌纤维素合酶的基本特性出发,综述了菌种筛选、提升产量和合酶的细胞定位等内容;围绕纤维素合酶的作用机理阐述了体外合成方法的影响因素,以及利用该方法探究各亚基相关作用的现状。以此探究细菌纤维素合酶的合成机制,并提出了当前研究中存在的问题。同时,展望了该领域未来的研究方向,以期通过对合成机理的探讨为细菌纤维素的大规模应用提供理论基础。     

Cellulose biosynthesis: current views and evolving concepts

* AIMS: To outline the current state of knowledge and discuss the evolution of various viewpoints put forth to explain the mechanism of cellulose biosynthesis. * SCOPE: Understanding the mechanism of cellulose biosynthesis is one of the major challenges in plant biology. The simplicity in the chemical structure of cellulose belies the complexities that are associated with the synthesis and assembly of this polysaccharide. Assembly of cellulose microfibrils in most organisms is visualized as a multi-step process involving a number of proteins with the key protein being the cellulose synthase catalytic sub-unit. Although genes encoding this protein have been identified in almost all cellulose synthesizing organisms, it has been a challenge in general, and more specifically in vascular plants, to demonstrate cellulose synthase activity in vitro. The assembly of glucan chains into cellulose microfibrils of specific dimensions, viewed as a spontaneous process, necessitates the assembly of synthesizing sites unique to most groups of organisms. The steps of polymerization (requiring the specific arrangement and activity of the cellulose synthase catalytic sub-units) and crystallization (directed self-assembly of glucan chains) are certainly interlinked in the formation of cellulose microfibrils. Mutants affected in cellulose biosynthesis have been identified in vascular plants. Studies on these mutants and herbicide-treated plants suggest an interesting link between the steps of polymerization and crystallization during cellulose biosynthesis. * CONCLUSIONS: With the identification of a large number of genes encoding cellulose synthases and cellulose synthase-like proteins in vascular plants and the supposed role of a number of other proteins in cellulose biosynthesis, a complete understanding of this process will necessitate a wider variety of research tools and approaches than was thought to be required a few years back.     

Reduced cellulose synthesis invokes lignification and defense responses in Arabidopsis thaliana

The cell wall determines the shape of plant cells and is also the primary interface for pathogen interactions. The structure of the cell wall can be modified in response to developmental and environmental cues, for example to strengthen the wall and to create barriers to pathogen ingress. The ectopic lignin 1-1 and 1-2 (eli1-1 and eli1-2) mutations lead to an aberrant deposition of lignin, a complex phenylpropanoid polymer. We show that the eli1 mutants occur in the cellulose synthase gene CESA3 in Arabidopsis thaliana and cause reduced cellulose synthesis, providing further evidence for the function of multiple CESA subunits in cellulose synthesis. We show that reduced levels of cellulose synthesis, caused by mutations in cellulose synthase genes and in genes affecting cell expansion, activate lignin synthesis and defense responses through jasmonate and ethylene and other signaling pathways. These observations suggest that mechanisms monitoring cell wall integrity can activate lignification and defense responses.     

Plant callose synthase complexes

Synthesis of callose (-1,3-glucan) in plants has been a topic of much debate over the past several decades. Callose synthase could not be purified to homogeneity and most partially purified cellulose synthase preparations yielded -1,3-glucan in vitro, leading to the interpretation that cellulose synthase might be able to synthesize callose. While a rapid progress has been made on the genes involved in cellulose synthesis in the past five years, identification of genes for callose synthases has proven difficult because cognate genes had not been identified in other organisms. An Arabidopsis gene encoding a putative cell plate-specific callose synthase catalytic subunit (CalS1) was recently cloned. CalS1 shares high sequence homology with the well-characterized yeast -1,3-glucan synthase and transgenic plant cells over-expressing CalS1 display higher callose synthase activity and accumulate more callose. The callose synthase complex exists in at least two distinct forms in different tissues and interacts with phragmoplastin, UDP-glucose transferase, Rop1 and, possibly, annexin. There are 12 CalS isozymes in Arabidopsis, and each may be tissue-specific and/or regulated under different physiological conditions responding to biotic and abiotic stresses.     

Identification of a new gene in an operon for cellulose biosynthesis in Acetobacter xylinum

DNA sequencing of the region downstream of the cellulose synthase catalytic subunit gene of Acetobacter xylinum led to the identification of an open reading frame coding for a polypeptide of 86 kDa. The deduced amino acid sequence of this polypeptide matches from position 27 to 40 with the N-terminal amino acid sequence determined for a 93 kDa polypeptide that copurifies with the cellulose synthase catalytic subunit during purification of cellulose synthase. The cellulose synthase catalytic subunit gene and the gene encoding the 93 kDa polypeptide, along with other genes probably, are organized as an operon for cellulose biosynthesis in which the first gene is the catalytic subunit gene and the second gene codes for the 93 kDa polypeptide. The function of the 93 kDa polypeptide is not clear at present, however it appears to be tightly associated with the cellulose synthase catalytic subunit. Sequence analysis of the polypeptide shows that it is a membrane protein with a signal sequence at the N-terminal end and a transmembrane helix in the C-terminal region for anchoring it into the membrane.     

Integrative approaches to determining Csl function

While there is an ever-increasing amount of information regarding cellulose synthase catalytic subunits (CesA) and their role in the formation of the cell wall, the remainder of the enzymes that synthesize structural cell wall polysaccharides are unknown. The completion of the Arabidopsis genome and the wealth of the sequence information from other plant genome projects provide a rich resource for determining the identity of these enzymes. Arabidopsis contains six families of genes related to cellulose synthase, the cellulose synthase-like (Csl) genes. Our laboratory is taking a multidisciplinary approach to determine the function of the Csl genes, incorporating genomic, genetic and biochemical data. Information from expressed sequence tag (EST) projects has revealed the presence of Csl genes in all plant species with a significant number of ESTs. Certain Csl families appear to be missing from some species. For example, no examples of CslG ESTs have been found in rice or maize. Microarray data and reporter constructs are being used to determine the expression pattern of the CesA and Csl genes in Arabidopsis. Mutations and insertion events have been identified in a majority of the genes in the Arabidopsis CesA superfamily and are being characterized by phenotypic and biochemical analysis. While we cannot yet link the function of any of the Csl genes to their respective products, the expression and localization of these genes is consistent with the expected expression pattern of polysaccharide synthases that contribute to the primary cell wall.     

Three AtCesA6‐like members enhance biomass production by distinctively promoting cell growth in Arabidopsis

Cellulose is an abundant biopolymer and a prominent constituent of plant cell walls. Cellulose is also a central component to plant morphogenesis and contributes the bulk of a plant's biomass. While cellulose synthase (CesA) genes were identified over two decades ago, genetic manipulation of this family to enhance cellulose production has remained difficult. In this study, we show that increasing the expression levels of the three primary cell wall AtCesA6‐like genes (AtCesA2, AtCesA5, AtCesA6), but not AtCesA3, AtCesA9 or secondary cell wall AtCesA7, can promote the expression of major primary wall CesA genes to accelerate primary wall CesA complex (cellulose synthase complexes, CSCs) particle movement for acquiring long microfibrils and consequently increasing cellulose production in Arabidopsis transgenic lines, as compared with wild‐type. The overexpression transgenic lines displayed changes in expression of genes related to cell growth and proliferation, perhaps explaining the enhanced growth of the transgenic seedlings. Notably, overexpression of the three AtCesA6‐like genes also enhanced secondary cell wall deposition that led to improved mechanical strength and higher biomass production in transgenic mature plants. Hence, we propose that overexpression of certain AtCesA genes can provide a biotechnological approach to increase cellulose synthesis and biomass accumulation in transgenic plants.