水稻氮饥饿诱导基因的克隆与表达分析

为了解水稻(Oryza sativa L.)对氮饥饿反应的分子与基因背景,利用RaSH策略构建了水稻氮(N)饥饿诱导cDNA文库.通过反向Northern筛选该文库,获得氮饥饿诱导的18个功能已知基因和2个功能未知基因.这些已知基因涉及碳代谢、次生代谢产物合成、蛋白质分解代谢、激素代谢、信号转导、生长调控过程及转录因子.这些基因表现出不同的时空表达模式.研究结果表明了植物对氮饥饿反应涉及互相关联的多种生理与分子机理,提供了相关的一些基因信息.     

水稻氮饥饿诱导基因的克隆与表达分析

为了解水稀(Oryza sativa L.)对氮饥饿反应的分子与基因背景,利用RaSH策略构建了水稻氮(N)饥饿诱导cDNA文库。通过反向Northern筛选该文库,获得氮饥饿诱导的18个功能已知基因和2个功能未知基因。这些已知基因涉及碳代谢、次生代谢产物合成、蛋白质分解代谢、激素代谢、信号转导、生长调控过程及转录因子。这些基因表现出不同的时空表达模式。研究结果表明了植物对氮饥饿反应涉及互相关联的多种生理与分子机理,提供了相关的一些基因信息。     

Nitrogen Partitioning During Early Development of Supernodulating Soybean (Glycine max [L.] Merrill) Mutants and their Wild-Type Parent

The relative importance of fixed N², cotyledonary N, and nitratefor growth of seedlings of soybean cv. Bragg and two of itsnitrate tolerant supernodulating (nts) mutants (intermediatents 1116 and extreme nts1007) was investigated during symbioticdevelopment in the presence of nitrate (3.0 mol m^–3) using¹⁵N techniques. Newly-fixed N₂ and nitrate were both major sourcesof N for nodule development and nitrate principally supportedearly shoot and root growth in Bragg. In the nts mutants, however,all plant parts and nodules in particular, relied more on Nstored in the cotyledons. This resulted in later nodule maturityand a period of prolonged N-starvation for the seedlings ofthe extreme supernodulator, and could be responsible for theirsubsequently lowered biomass accumulation compared to the parentcultivar. Key words: Nodules, N partitioning, supernodulating soybeans     

Effect of nitrogen deficiency on antioxidant status and Cd toxicity in rice seedlings

Effect of nitrogen (N) deficiency on antioxidant status and Cd toxicity in rice seedlings was investigated. N deficiency resulted in a reduction of shoot growth but not root growth. The contents of N-containing compounds such as nitrate, chlorophyll, and protein decreased in leaves of rice seedlings grown under N deficiency. Accumulation of abscisic acid and H₂O₂ in leaves was induced by N deficiency. The content of ascorbate and the activities of ascorbate peroxidase, glutathione reductase, and catalase in N-deficient leaves were lower than their respective control leaves. However, glutathione content was not affected and superoxide dismutase activity was increased by N deficiency. Cd toxicity in N-deficient seedlings was more pronounced than that in N-sufficient ones. Pretreatment with ascorbate or L-galactono-1,4-lactone, a biosynthetic precursor of ascorbate resulted in a reduction of Cd toxicity enhanced by N deficiency. N deficiency also resulted in an enhancement of Cd uptake in rice seedlings. The possible mechanism of Cd toxicity enhanced by N deficiency is discussed.     

Proteomics analysis reveals multiple regulatory mechanisms in response to selenium in rice

Selenium (Se) shows both beneficial and toxic effects on plant growth. Rice (Oryza sativa L.) seedlings cultivated under lower concentrations of sodium selenite showed enhanced growth, whereas higher concentrations of sodium selenite repressed seedling growth. To acquire detailed regulatory mechanisms underlying these effects, a comparative proteomics study using 2-dimensional gel electrophoresis and MALDI-TOF/TOF MS was performed. By comparison of gel images between Se treatments and control, 66 and 97 differentially expressed proteins were identified in shoot and root, respectively under at least one of the Se treatment concentrations. Gene Ontology and Clustering analysis reveal primary metabolism, photosynthesis and redox homeostasis are the most highly affected biological processes by Se treatments. Lower Se treatments (2 and 6 mg/L sodium selenite) activated antioxidative system, enhanced photosynthesis and primary metabolism. However, higher Se treatment (10 mg/L sodium selenite) damaged photosynthesis apparatus, inhibited photosynthesis and primary metabolism. Protein ubiquitination and phosphorylation may also play important roles in Se response in rice. In conclusion, our study provided novel insights into Se response in rice at the proteome level, which are expected to be highly useful for dissecting the Se response pathways in higher plants and for producing Se enriched rice cultivars in the future.     

Physiological studies and proteomic analysis for differentially expressed proteins and their possible role in the root of N-efficient rice (Oryza sativa L.)

The root proteome of nitrogen-efficient and nitrogen-inefficient rice cultivars was compared in this study in order to investigate the differential expression of proteins under deficient (1 mM), low (10 mM) and high (25 mM) levels of nitrogen (N). Nitrogen use efficiency (NUE) was assessed by biochemical assays such as N-uptake kinetics and activities of N-assimilation enzymes. Two-dimensional gel electrophoresis and MALDI–TOF–MS analysis resulted in the identification of 504 protein spots (210 and 294 spots in cvs. Rai Sudha and Munga Phool, respectively). A positive correlation was observed between physiological parameters and the concentration of a number of root proteins. Sixty-three spots showed a significant cultivar × N-treatment effect on the level of expression. Functional aspects of eleven spots with major alterations in expression over control were critically analyzed. The data suggest that glutamine synthetase, cysteine proteinase inhibitor-I, porphobilinogen deaminase (fragment) and ferritin were involved in conferring N efficiency to the N-efficient rice cultivars/genotypes. Interestingly, these proteins are involved directly or indirectly in N assimilation. Such studies should help us in identifying and understanding the structural or functional protein(s) involved in the response to the level of nitrogen fertilization.     

土壤施钙诱导水稻幼苗抗低温和抗病生理机制研究

试验发现,以干土重1％的CaO混土处理培育水稻秧苗,能显著提高幼苗的抗低温和抗立枯病能力．对其生理机制研究表明,该处理对水稻幼苗体内活性氧清除酶系统具有显著影响,与空白对照处理相比。施钙处理的水稻幼苗根部和地上部SOD活性增强;根部POD活性显著增强,地上部POD活性下降;根部和地上部CAT活性则先下降,后上升;根部和地上部的可溶性蛋白含量均有所上升．POD同工酶PAGE电泳结果表明,CaO处理的水稻幼苗地上部POD同工酶谱带明显减弱和减少,而根部POD同工酶谱带增强和增多．这些结果揭示了土壤施加CaO可提高水稻幼苗抗低温和抗病能力的部分原因．     

Regulation of the α-expansin gene OsEXPA8 expression affects root system architecture in transgenic rice plants

Expansins are cell wall proteins implicated in the control of plant growth via loosening of the extracellular matrix, and are encoded by a large gene family. However, data linked to loss of function of single genes which support the role of expansins in root growth remain limited. In this study, we used RNA interference to examine the biological functions of the rice α-expansin gene OsEXPA8. Repression of OsEXPA8 expression in rice impaired the root system architecture and plant growth significantly, leading to shorter primary roots and fewer lateral roots. Accordingly, the cell size of the root vascular bundle reduced drastically. Notably, OsEXPA8 silencing impaired root hair elongation; moreover, plant height was clearly reduced. Transient expression of OsEXPA8-GFP in onion epidermal cells verified that OsEXPA8 is located on the cell wall. OsEXPA8 was expressed predominantly in the root and shoot of one-week-old rice seedlings, and highly induced by NaCl but suppressed by nitrate and phosphate starvation. In addition, atomic force microscopy was used to explore alterations in cell wall nanomechanics caused by OsEXPA8 protein reduction, which showed that the wall stiffness (Young’s modulus) of OsEXPA8-silenced suspension cells was increased significantly. Taken together, our results suggest that OsEXPA8 is critical for root system architecture, which supports the hypothesis that expansins are involved in enhancing plant growth by mediating cell wall loosening.     

The root form of ferredoxin-NADP oxidoreductase is expressed in rice coleoptiles for the assimilation of nitrate

Two ferredoxin-dependent proteins, nitrite reductase and glutamate synthase, play a role in nitrate assimilation during the anaerobic germination of rice (Oryza sativa L.). This paper reports the expression of the root form of ferredoxin-NADP⁺ oxidoreductase (FNR), the protein responsible for providing reduced ferredoxin in rice coleoptiles. Using an antibody against FNR, a protein with the expected molecular mass for root FNR (35 kDa) was recognized by Western blot analysis in extracts from aerobic and anaerobic coleoptiles. The enzyme is synthesized de novo, as shown by immunoprecipitation of the radiolabeled 35-kDa protein from anaerobic seedlings grown in the presence of [³⁵S]methionine. Northern blot analysis with specific probes for root and leaf FNR showed the presence of mRNA for the root form but not for the leaf form, in both aerobic and anaerobic rice coleoptiles. The inductive effect of exogenous nitrate on the expression of FNR is further evidence for the presence of the root type of FNR in rice coleoptiles. The importance of the expression of root FNR during the anaerobic development of rice seedlings is discussed. Received: 7 October 1996 / Accepted: 22 January 1997     

Carbon–Nitrogen Interaction Modulates Plant Growth and Expression of Metabolic Genes in Rice

Carbon (C) and nitrogen (N) interact to coordinate their metabolism in achieving C:N homeostasis in all cellular organisms. Plant shoots and roots take up C and N, respectively, and the coordinated C and N assimilation is essential for normal plant growth and development. In this study, rice was used as a model system for the investigation of molecular mechanisms underlying C–N interactions and coordination in cereal species. We investigated the growth response of rice seedlings to a wide range of exogenous C:N availabilities and established balanced exogenous C:N that was optimal for rice seedling growth. To assess correlations between the modulation of plant growth and the regulation of metabolic gene expression by C:N availabilities, we examined the expression of PEPC, PK, NR, GS, and GOGAT in rice seedlings treated with four C:N availabilities: low C/low N, low C/high N, high C/low N, and high C/high N. It was found that their expression was subjected to complex regulation by C:N availabilities. Our results demonstrate that growth of shoot and root rice seedlings is regulated by C–N interaction and growth modulations are associated with changes in metabolic gene expression. Our findings suggest that rice is a useful model system for the investigation of regulation mechanisms responsible for C–N interaction and coordination in plants.     

Seedling vigour and the early growth of transplanted rice (Oryza sativa)

Previous studies suggest that the positive response of transplanted rice (Oryza sativa L.) to nursery fertiliser application was due to increased seedling vigour or possibly to increased nutrient content. This paper presents results of two glasshouse experiments designed to test the hypothesis that seedling vigour was responsible for the response of transplanted seedlings to nursery treatments. The aim of the present study was to explore the concept of seedling vigour of transplanted rice and to determine what plant attributes conferred vigour on the seedlings. Seedling vigour treatments were established by subjecting seedlings to short-term submergence (0, 1 and 2 days/week) in one experiment and to leaf clipping or root pruning and water stress in another to determine their effect on plant growth after transplanting. Submerging seedlings increased plant height but depressed shoot and root dry matter and root:shoot ratio of the seedling at 28 days after sowing. After transplanting these seedlings, prior submergence depressed shoot dry matter at 40 days. Nursery nutrient application increased plant height, increased root and shoot dry matter, but generally decreased root:shoot ratio. Pruning up to 60% of the roots at transplanting decreased shoot and root dry matter, P concentration in leaves at panicle initiation (PI) and straw dry matter and grain yield at maturity. By contrast, pruning 30% of leaves depressed shoot and root dry matter by 30% at PI, and root dry matter and straw and grain yield by 20% at maturity. The combined effects of leaf clipping and root pruning on shoot, root and straw dry matter were largely additive. It is concluded that the response of rice yield to nursery treatments is largely due to increased seedling vigour and can be effected by a range of nutritional as well as non-nutritional treatments of seedlings that increase seedling dry matter, nutrient content, and nutrient concentration. Impairment of leaf growth and to a lesser extent root growth in the nursery depressed seedling vigour after transplanting. However, rather than increasing stress tolerance, seedling vigour was more beneficial when post transplant growth was not limited by nutrient or water stresses.     

Root growth dynamics of Aleppo pine (Pinus halepensis Mill.) seedlings in relation to shoot elongation, plant size and tissue nitrogen concentration

Large and high nitrogen (N) concentration seedlings frequently have higher survival and growth in Mediterranean forest plantations than seedlings with the opposite traits, which has been linked to the production of deeper and larger root systems in the former type of seedlings. This study assessed the influence of seedling size and N concentration on root growth dynamics and its relation to shoot elongation in Aleppo pine (Pinus halepensis Mill.) seedlings. We cultivated seedlings that differed in size and tissue N concentration that were subsequently transplanted into transparent methacrylate tubes in the field. The number of roots, root depth, and the root and shoot elongation rate (length increase per unit time) were periodically measured for 10 weeks. At the end of the study, we also measured the twig water potential (ψ) and the mass of plant organs. New root mass at the end of the study increased with seedling size, which was linked to the production of a greater number of new roots of lower specific length rather than to higher elongation rate of individual roots. Neither plant size nor N concentration affected root depth. New root mass per leaf mass unit, shoot elongation rate, and pre-dawn ψ were reduced with reduction in seedling size, while mid-day ψ and the root relative growth rate were not affected by seedling size. N concentration had an additive effect on plant size on root growth but its overall effect was less important than seedling size. Shoot and roots had an antagonistic elongation pattern through time in small seedlings, indicating that the growth of both organs depressed each other and that they competed for the same resources. Antagonism between shoot and root elongation decreased with plant size, disappearing in large and medium seedlings, and it was independent of seedling N concentration. We conclude that root and shoot growth but not rooting depth increased with plant size and tissue N concentration in Aleppo pine seedlings. Since production of new roots is critical for the establishment of planted seedlings, higher absolute root growth in large seedlings may increase their transplanting performance relative to small seedlings. The lack of antagonism between root and shoot growth in large seedlings suggests that these plants can provide resources to sustain simultaneous growth of both organs.     

Proteomic Analysis of Salt Stress Responses in Rice Shoot

To gain a better understanding of the mechanism of rice (Oryza sativa L.) in response to salt stress, we performed a proteomics analysis of rice in response to 250 mM NaCl treatment using shoots of 3-day-old nascent seedlings. The changes of protein patterns were monitored with two-dimensional gel electrophoresis. Of 57 protein spots showing changes in abundance in response to salt stress, 52 were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The identified proteins were classified into eight functional categories. Several novel salt stress-responsive proteins, including protein synthesis inhibitor I, photosystem II stability/assembly factor HCF136, trigger factor-like protein and cycloartenol-C24-methyltransferase are upregulated upon salt stress. In order to figure out the different and similar molecular mechanism among salt and other stresses, regulation of some salt responsive proteins under other abiotic stress (cold and dehydration) and abscisic acid application was also analyzed. The possible molecular mechanism of rice seedlings in response to salinity and other stresses were discussed.