AtPLC2, a gene encoding phosphoinositide-specific phospholipase C,is constitutively expressed in vegetative and floral tissues in Arabidopsis thaliana

A cDNA encoding a phosphoinositide-specific phospholipase C (PI-PLC) from the higher plant Arabidopsis thaliana was cloned and characterized.The gene corresponding to this cDNA is designated AtPLC2. The overall structure of the predicted AtPLC2 protein is similar to those of plant PI-PLCs and mammalian -type PI-PLCs. Northern blot analysis revealed that AtPLC2 is expressed constitutively whereas AtPLC1S, another gene for PI-PLC of Arabidopsis, is induced by environmental stresses such as dehydration and salinity, indicating that the function of AtPLC2 is distinct from that of AtPLC1S. The AtPLC2 mRNA was detected in vegetative and floral tissues. We determined the positions of these two PI-PLCs genes on Arabidopsis chromosomes by RFLP mapping using P1 genomic clones.     

Preparation of polyclonal antibody specific for AtPLC4, an Arabidopsis phosphatidylinositol-specific phospholipase C in rabbits

Phosphoinositide-specific phospholipase Cs (PI-PLCs) are important enzymes in eukaryotes, which catalyze the hydrolysis of phosphatidylinositol 4,5-bisphosphate into the two second messengers inositol 1,4,5-trisphosphate and diacylglycerol. The Arabidopsis genome contains nine putative PI-PLC genes. AtPLC4, an abiotic stress induced gene, has been reported to encode an active PI-PLC isoform. However, the exact roles of putative AtPLC4 in plant remain to be elicited. The first 108 amino acid residues of the N-terminal of AtPLC4, referred to as AtPLC4 N, was expressed as a recombinant protein in Escherichia coli and used as antigen in generating antibody. Purified recombinant proteins including AtPLC1 to AtPLC5, AtPLC8, AtPLC9 and AtPLC4 N were transferred onto the same blot to test specificity of the prepared antibody. Western blot result shows that only AtPLC4 and AtPLC4 N can be recognized by the antibody. The antibody recognized a protein of approximately 68kDa in the plasma membrane fraction and cytosolic fractions prepared from Arabidopsis thaliana plants. This corresponds very well with the calculated molecular weight of AtPLC4. The results suggest that AtPLC4 may encode a plasma membrane-associated protein.     

N-terminal EF-hand-like domain is required for phosphoinositide-specific phospholipase C activity in Arabidopsis thaliana

Phosphoinositide-specific phospholipase C's (PI-PLCs) are ubiquitous in eukaryotes, from plants to animals, and catalyze the hydrolysis of phosphatidylinositol 4,5-bisphosphate into the two second messengers inositol 1,4,5-trisphosphate and diacylglycerol. In animals, four distinct subfamilies of PI-PLCs have been identified, and the three-dimensional structure of one rat isozyme, PLC-delta1, determined. Plants appear to contain only one gene family encoding PI-PLCs. The catalytic properties of plant PI-PLCs are very similar to those of animal enzymes. However, very little is known about the regulation of plant PI-PLCs. All plant PI-PLCs comprise three domains, X, Y and C2, which are also conserved in isoforms from animals and yeast. We here show that one PI-PLC isozyme from Arabidopsis thaliana, AtPLC2, is predominantly localized in the plasma membrane, and that the conserved N-terminal domain may represent an EF-hand domain that is required for catalytic activity but not for lipid binding.     

Unique and overlapping expression patterns among the Arabidopsis 1-amino-cyclopropane-1-carboxylate synthase gene family members

1-Aminocyclopropane-1-carboxylate synthase (ACS) catalyzes the rate-limiting step in the ethylene biosynthetic pathway in plants. The Arabidopsis genome encodes nine ACS polypeptides that form eight functional (ACS2, ACS4-9, and ACS11) homodimers and one nonfunctional (ACS1) homodimer. Transgenic Arabidopsis lines were constructed expressing the beta-glucuronidase (GUS) and green fluorescence protein (GFP) reporter genes from the promoter of each of the gene family members to determine their patterns of expression during plant development. All genes, except ACS9, are expressed in 5-d-old etiolated or light-grown seedlings yielding distinct patterns of GUS staining. ACS9 expression is detected later in development. Unique and overlapping expression patterns were detected for all the family members in various organs of adult plants. ACS11 is uniquely expressed in the trichomes of sepals and ACS1 in the replum. Overlapping expression was observed in hypocotyl, roots, various parts of the flower (sepals, pedicle, style, etc.) and in the stigmatic and abscission zones of the silique. Exogenous indole-3-acetic acid (IAA) enhances the constitutive expression of ACS2, 4, 5, 6, 7, 8, and 11 in the root. Wounding of hypocotyl tissue inhibits the constitutive expression of ACS1 and ACS5 and induces the expression of ACS2, 4, 6, 7, 8, and 11. Inducers of ethylene production such as cold, heat, anaerobiosis, and Li(+) ions enhance or suppress the expression of various members of the gene family in the root of light-grown seedlings. Examination of GUS expression in transverse sections of cotyledons reveals that all ACS genes, except ACS9, are expressed in the epidermis cell layer, guard cells, and vascular tissue. Similar analysis with root tip tissue treated with IAA reveals unique and overlapping expression patterns in the various cell types of the lateral root cap, cell division, and cell expansion zones. IAA inducibility is gene-specific and cell type-dependent across the root tip zone. This limited comparative exploration of ACS gene family expression reveals constitutive spatial and temporal expression patterns of all gene family members throughout the growth period examined. The unique and overlapping gene activity pattern detected reveals a combinatorial code of spatio-temporal coexpression among the various gene family members during plant development. This raises the prospect that functional ACS heterodimers may be formed in planta.     

番茄AT-hook基因家族的鉴定及胁迫条件下的表达分析

AT-hook蛋白家族在植物生长发育、器官构建及逆境胁迫和激素信号应答中发挥重要作用。本研究在番茄基因组范围内,利用生物信息学方法对番茄AT-hook基因家族的成员、分布、结构和功能进行分析。结果表明,番茄AT-hook家族包含32个成员,分为3种类型,其中类型Ⅰ含有13个成员;遗传进化分析表明番茄AT-hook基因成员与拟南芥家族基因具有相似分类。利用实时荧光定量PCR对番茄32个基因开展组织表达分析,结果表明AT-hook基因具有表达差异,主要在根和花中表达较高。氧化胁迫分析结果表明,32个基因受ABA、SA、盐、高温和低温诱导表达,其中部分基因显著上调或下调表达,很可能参与了番茄逆境胁迫条件下的防御应答反应。本研究结果将为番茄AT-hook家族基因的深入研究提供依据,为进一步解析番茄AT-hook基因的功能奠定基础。     

The Arabidopsis XET-related gene family: environmental and hormonal regulation of expression

Enzymes that modify cell wall components most likely play critical roles in altering size, shape, and physical properties of plant cells. Regulation of such modifying activity is expected to be important during morphogenesis and in eliciting developmental and physiological alterations that arise in response to environmental conditions. Previous work has shown that the Arabidopsis TCH4 gene encodes a xyloglucan endotransglycosylase (XET) which acts on the major hemicellulose of the plant cell wall. The expression of TCH4 is dramatically upregulated in response to several environmental stimuli (including touch, wind, darkness, heat shock, and cold shock) as well as the growth-enhancing hormones, auxin and brassinosteroids. This paper reports the presence of an extensive X ET ,related (XTR) gene family in Arabidopsis. In addition to TCH4, this family includes two previously identified genes, EXT and Meri-5, and at least five additional genes. The cDNAs of the XTR family share between 46 and 79% sequence identity and the predicted XTR proteins share from 37 to 84% identity. All eight proteins include potential N-terminal signal sequences and most have a conserved motif (DEIDFEFLG) that is also found in Bacillusβ-glucanase and may be important for enzyme activity. The members of the XTR gene family are differentially sensitive to environmental and hormonal stimuli. Magnitude and kinetics of regulation are distinct for the different genes. Differential regulation of expression of this complex gene family suggests a recruitment of related, yet distinct, cell wall-modifying enzymes that may control the properties of cell walls and tissues during development and in response to environmental cues.     

Comprehensive analysis of CCCH-type zinc finger gene family in citrus (Clementine mandarin) by genome-wide characterization

The CCCH-type zinc finger proteins comprise a large gene family of regulatory proteins and are widely distributed in eukaryotic organisms. The CCCH proteins have been implicated in multiple biological processes and environmental responses in plants. Little information is available, however, about CCCH genes in plants, especially in woody plants such as citrus. The release of the whole-genome sequence of citrus allowed us to perform a genome-wide analysis of CCCH genes and to compare the identified proteins with their orthologs in model plants. In this study, 62 CCCH genes and a total of 132 CCCH motifs were identified, and a comprehensive analysis including the chromosomal locations, phylogenetic relationships, functional annotations, gene structures and conserved motifs was performed. Distribution mapping revealed that 54 of the 62 CCCH genes are unevenly dispersed on the nine citrus chromosomes. Based on phylogenetic analysis and gene structural features, we constructed 5 subfamilies of 62 CCCH members and integrative subfamilies from citrus, Arabidopsis, and rice, respectively. Importantly, large numbers of SNPs and InDels in 26 CCCH genes were identified from Poncirus trifoliata and Fortunella japonica using whole-genome deep re-sequencing. Furthermore, citrus CCCH genes showed distinct temporal and spatial expression patterns in different developmental processes and in response to various stress conditions. Our comprehensive analysis of CleC3Hs is a valuable resource that further elucidates the roles of CCCH family members in plant growth and development. In addition, variants and comparative genomics analyses deepen our understanding of the evolution of the CCCH gene family and will contribute to further genetics and genomics studies of citrus and other plant species.     

Genome-wide identification and evolutionary analysis of Arabidopsis sm genes family

Sm proteins are members of a family of small proteins that are widespread in biosphere and found associated with RNA metabolism. To date, to our knowledge, only Arabidopsis SAD1 gene has been studied functionally in plant. In this study, 42 Sm genes are identified through comprehensive analysis in Arabidopsis. And a complete overview of this gene family is presented, including the gene structures, phylogeny, chromosome locations, selection pressure and expression. The results reveal that gene duplication contributes to the expansion of the Sm gene family in Arabidopsis genome, diverse expression patterns suggest their functional differentiation and divergence analysis indicates purifying selection as a key role in evolution. Our comparative genomics analysis of Sm genes will provide the first step towards the future experimental research on determining the functions of these genes.     

The multigenic family of thioredoxin h in Arabidopsis thaliana: specific expression and stress response

In the model plant Arabidopsis thaliana, cytosolic thioredoxins h (TRXh) are encoded by a multigenic family of eight genes. Genomic studies have revealed that a number of these genes are duplicated genes originating from a common ancestor. This multiplicity of thioredoxin h genes raises questions of the specificity of plant thioredoxins and the function of such a large multigenic family in plant. The results from studies using northern blots, semi-quantitative RT-PCR and transgenic promoter–GUS fusions provide strong evidence that the members of the AtTRXh gene family show expression levels that vary among different plant organs. Moreover, distinct AtTRXh genes are induced in response to pathogenic elicitors. Together, our data suggest that the members of the multigenic family of AtTRXh may not have redundant functions.     

Genome-Wide Survey and Expression Analysis Suggest Diverse Roles of Glutaredoxin Gene Family Members During Development and Response to Various Stimuli in Rice

Glutaredoxins (GRXs) are glutathione-dependent oxidoreductase enzymes involved in a variety of cellular processes. In this study, our analysis revealed the presence of 48 genes encoding GRX proteins in the rice genome. GRX proteins could be classified into four classes, namely CC-, CGFS-, CPYC- and GRL-type, based on phylogenetic analysis. The classification was supported with organization of predicted conserved putative motifs in GRX proteins. We found that expansion of this gene family has occurred largely via whole genome duplication events in a species-specific manner. We explored rice oligonucleotide array data to gain insights into the function of GRX gene family members during various stages of development and in response to environmental stimuli. The comprehensive expression analysis suggested diverse roles of GRX genes during growth and development in rice. Some of the GRX genes were expressed in specific organs/developmental stages only. The expression of many of rice GRX genes was influenced by various phytohormones, abiotic and biotic stress conditions, suggesting an important role of GRX proteins in response to these stimuli. The identification of GRX genes showing differential expression in specific tissues or in response to environmental stimuli provide a new avenue for in-depth characterization of selected genes of importance.     

The ALDH gene superfamily of Arabidopsis

Aldehyde dehydrogenases (ALDHs) represent a protein superfamily of NAD(P)(+)-dependent enzymes that oxidize a wide range of endogenous and exogenous aliphatic and aromatic aldehydes. The Arabidopsis genome contains 14 unique ALDH sequences encoding members of nine ALDH families, including eight known families and one novel family (ALDH22) that is currently known only in plants. Here, we identify members of the ALDH gene superfamily in Arabidopsis; provide a revised, unified nomenclature for these ALDH genes; analyze the molecular relationship among Arabidopsis ALDH genes and compare them to ALDH genes from other species, including prokaryotes and mammals; and describe the role of ALDHs in cytoplasmic male sterility, plant defense and abiotic stress tolerance.