Mobilization of Starch after Submergence of Air-Grown Rice Coleoptiles. Implications for Growth and Gravitropism

Submergence of air-grown rice seedlings (Oryza sativa L. var. Sasanishiki) induces elongation of the coleoptile. We investigated whether rapid underwater extension is associated with a loss of starch. After 1 d of submergence the starch content was reduced by 70%. This loss of reserve carbohydrate was accompanied by a 38% increase in the concentration of glucose in the cell sap of the coleoptiles. The submerged (starch-depleted) coleoptiles had a slower negative gravitropism than the air-grown controls, although the rate of elongation in the horizontal position was not impaired. We conclude that the submergence-induced mobilization of starch provides substrates and osmotica for the rapidly growing cells. In addition, our results indicate that a full complement of starch is necessary for normal gravitropism in the rice coleoptile.     

水稻耐淹涝性状的遗传分析和SSR标记的研究

淹涝胁迫对水稻生产造成了严重影响, 发掘可应用于耐淹涝辅助选择的分子标记(MAS), 将有助于水稻耐淹涝性状的遗传改良。应用耐淹涝材料FR13A和淹涝敏感材料IR39595-503-2-1-2为亲本做正反交获得F₁和F₂代群体。对正反交的F₁群体的耐淹涝性状进行遗传分析, 发现正反交的F₁代群体在耐淹涝性状上没有显著差异, 说明耐淹涝性状是核基因控制。从两次淹涝处理中F₂代群体的分离情况来看, 来源于FR13A的耐淹特性表现出数量-质量性状遗传的特点。当淹涝胁迫压力比较轻时表现为数量性状遗传, 具有微效多基因的作用。当淹涝胁迫压力增大时, 表现为主效基因控制的质量性状。在SSR分析中, 187对SSR引物中有73对引物在两亲本间有明显的差异, 差异率为39%。用这73对差异引物, 对F2群体进行多态筛选, 结果筛选到一个与耐淹涝性状连锁的标记RM219, 验证了耐淹涝性状确实由主效基因Sub1控制, 因此, RM219在水稻耐淹涝育种中具有利用价值。     

Reorientation of Microfibrils and Microtubules at the Outer Epidermal Wall of Maize Coleoptiles During Auxin-Mediated Growth

Auxin-mediated elongation growth of isolated subapical coleoptile segments of maize (Zea mays L.) is controlled by the extensibility of the outer cell wall of the outer epidermis (Kutschera et al., 1987). Here we investigate the hypothesis that auxin controls the extensibility of this wall by changing the orientation of newly deposited microfibrils through a corresponding change in the orientation of cortical microtubules. On the basis of electron micrographs it is shown that cessation of growth after removal of the endogenous source of auxin is correlated with a relative increase of longitudinally orientated microfibrils and microtubules at the inner wall surface. Conversely, reinduction of growth by exogenous auxin is correlated with a relative increase of transversely orientated microfibrils and microtubules at the inner wall surface. These changes can be detected 30–60 min after the removal and addition of auxin, respectively. The functional significance of directional changes of newly desposited wall microfibrils for the control of elongation growth is discussed.     

水稻萌发耐淹性的遗传分析

水稻(Oryza sativa)萌发耐淹性受到复杂的网络调控, 其分子机制不同于苗期耐淹性的相关机制, 萌发耐淹性的强弱影响着直播稻的成苗率。通过对256份水稻核心种质的萌发耐淹性评估, 发现粳稻和籼稻之间的萌发耐淹性差异并不显著, 都存在广泛的遗传变异。利用以粳稻R0380为供体亲本, 籼稻RP2334为轮回亲本的170个高代回交自交系构建含146个分子标记的连锁图谱, 以低氧胚芽鞘长度为性状指标, 通过复合区间作图法检测到影响萌发耐淹性的4个QTLs(quantitative trait loci), 分别定位于第2(2个)、3(1个)和8号(1个)染色体。贡献率最大的QTL为qGS2.2, 其值为17.34%, 增效等位基因来自轮回亲本籼稻RP2334; 其余3个QTLs的增效等位基因均来自供体亲本粳稻R0380, 贡献率分别为12.86%、9.37%和14.60%。     

Dynamic Aspects of Alcoholic Fermentation of Rice Seedlings in Response to Anaerobiosis and to Complete Submergence: Relationship to Submergence Tolerance

Rice plants are severely damaged by complete submergence. Thisis a problem in rice farming and could be the result, in part,of tissue anoxia imposed by a reduced availability of oxygen.To investigate this possibility we monitored alcoholic fermentationproducts as markers for tissue anaerobiosis using sensitivelaser-based spectroscopy able to sense ethanol and acetaldehydedown to 3 nl l^–1 and 0·1 nl l^–1,respectively. Acetaldehyde emission began within 0·5 hof imposing an oxygen-free gas phase environment followed closelyby ethanol. As treatment progressed, ethanol output increasedand came to exceed acetaldehyde emission as this stabilizedconsiderably after approx. 3 h. On re-entry of air, a sharppost-anaerobic peak of acetaldehyde production was observed.This was found to be diagnostic of a preceding anoxic episodeof 0·5 h or more. When anaerobiosis was lengthenedby up to 14 h, the size of the post-anaerobic acetaldehydeoutburst increased. After de-submergence from oxygen-free water,a similarly strong but slower post-anaerobic acetaldehyde upsurgewas seen, which was accompanied by an increase in ethanol emission.Light almost, but not completely, eliminated fermentation inanaerobic surroundings and also the post-anaerobic or post-submergencepeaks in acetaldehyde production. All photosynthetically generatedoxygen was consumed within the plant. There was no substantialdifference in acetaldehyde and ethanol output between FR13Aand the less submergence-tolerant line CT6241 under any submergencetreatment. In some circumstances, submergence damaged CT6241more than FR13A even in the absence of vigorous fermentation.We conclude that oxygen deprivation may not always determinethe extent of damage caused to rice plants by submergence undernatural conditions.     

Submergence tolerance of rice – A new glasshouse method for the experimental submergence of plants

A new method is described for evaluation of submergence tolerance of rice ( Oryza sativa L.) plants. Responses of a range of cultivars corresponded with known differences in field performance. The method 1) allows fast and effective determination of submergence tolerance, 2) allows screening of many plants in a small glasshouse area, 3) provides for recovery of superior plants for seed collection, 4) allows manipulation of many environmental variables to mimic the natural submergence environment, and 5) uses simple, inexpensive, readily available equipment. Physiological studies performed with this method gave results similar to those obtained from field studies and showed that submergence tolerance increased in older plants; it decreased with increasing depth, increasing temperature and with high or low light levels. The system is ideal for the rapid evaluation of rice germplasm under controlled conditions and physiological studies on the mechanism of rice submergence tolerance.     

Small GTP-Binding Protein Gene Is Associated with QTL for Submergence Tolerance in Rice

Small GTP-binding proteins play critical roles in signal transduction in mammalian and plant systems. In this study, sequence variation of a small GTP-binding protein identified in the subgenomic region was analyzed. The major quantitative trait locus (QTL) controlling submergence tolerance on the 6.5-cM region of chromosome 9 was previously mapped, sequenced, and annotated. One of the most interesting candidate genes located in this QTL was a 5.2-kb sequence, which included a coding sequence consisting of two exons and a promoter. The deduced amino acid sequence corresponded to a 24.8 kD protein consisting of 226 amino acids, with 98% identity to RGP1, a small GTP-binding protein involved in a signal pathway responding to hormones, such as cytokinin and ethylene. According to the amino acid sequence, a putative small G-protein was classified as a small Ras-related GTP-binding protein. DNA gel blot analysis showed that the putative gene encoding the Ras-related GTP-binding protein was present as a single copy in the rice genome. Comparison of genomic sequences from several rice cultivars tolerant to submergence identified single nucleotide polymorphisms located in the TATA box of the Ras promoter region. Linkage analysis showed that the putative gene for GTP-binding protein was tightly linked to the peak of the QTL previously mapped on the long arm of chromosome 9. The single strand conformation polymorphism of the putative GTP-binding protein gene can be used for allele discrimination and marker assisted selection for tolerance to flash flooding.     

Molecular Genetics of Submergence Tolerance in Rice: QTL Analysis of Key Traits

Flash flooding of young rice plants is a common problem forrice farmers in south and south-east Asia. It severely reducesgrain yield and increases the unpredictability of cropping.The inheritance and expression of traits associated with submergencestress tolerance at the seedling stage are physiologically andgenetically complex. We exploited naturally occurring differencesbetween certain rice lines in their tolerance to submergenceand used quantitative trait loci (QTL) mapping to improve understandingof the genetic and physiological basis of submergence tolerance.Three rice populations, each derived from a single cross betweentwo cultivars differing in their response to submergence, wereused to identify QTL associated with plant survival and variouslinked traits. These included total shoot elongation under water,the extent of stimulation of shoot elongation caused by submergence,a visual submergence tolerance score, and leaf senescence underdifferent field conditions, locations and years. Several majorQTL determining plant survival, plant height, stimulation ofshoot elongation, visual tolerance score and leaf senescenceeach mapped to the same locus on chromosome 9. These QTL weredetected consistently in experiments across all years and inthe genetic backgrounds of all three mapping populations. SecondaryQTL influencing tolerance were also identified and located onchromosomes 1, 2, 5, 7, 10 and 11. These QTL were specific toparticular traits, environments, or genetic backgrounds. Allidentified QTL contributed to increased submergence tolerancethrough their effects on decreased underwater shoot elongationor increased maintenance of chlorophyll levels, or on both.These findings establish the foundations of a marker-assistedscheme for introducing submergence tolerance into agriculturallydesirable cultivars of rice.     

Responses by Coleoptiles of Intact Rice Seedlings to Anoxia: K+ Net Uptake from the External Solution and Translocation from the Caryopses

This study evaluated the effects of anoxia on K⁺ uptake andtranslocation in 3–4-d-old, intact, rice seedlings (Oryzasativa L. cv. Calrose). Rates of net K⁺ uptake from the mediumover 24 h by coleoptiles of anoxic seedlings were inhibitedby 83–91 %, when compared with rates in aerated seedlings.Similar uptake rates, and degree of inhibition due to anoxia,were found for Rb⁺ when supplied over 1·5–2 h,starting 22 h after imposing anoxia. The Rb⁺ uptake indicatedthat intact coleoptiles take up ions directly from the externalsolution. Monovalent cation (K⁺ and Rb⁺) net uptake from thesolution was inhibited by anoxia to the same degree for thecoleoptiles of intact seedlings and for coleoptiles excised,‘aged’, and supplied with exogenous glucose. Transportof endogenous K⁺ from caryopses to coleoptiles was inhibitedless by anoxia than net K⁺ uptake from the solution, the inhibitionbeing 55 % rather than 87 %. Despite these inhibitions,osmotic pressures of sap (_sap) expressed from coleoptiles ofseedlings exposed to 48 h of anoxia, with or without exogenousK⁺, were 0·66 ± 0·03 MPa; however,the contributions of K⁺ to _sap were 23 and 16 %, respectively.After 24 h of anoxia, the K⁺ concentrations in the basal10 mm of the coleoptiles of seedlings with or without exogenousK⁺, were similar to those in aerated seedlings with exogenousK⁺. In contrast, K⁺ concentrations had decreased in aeratedseedlings without exogenous K⁺, presumably due to ‘dilution’by growth; fresh weight gains of the coleoptile being 3·6-to 4·7-fold greater in aerated than in anoxic seedlings.Deposition rates of K⁺ along the axes of the coleoptiles werecalculated for the anoxic seedlings only, for which we assessedthe elongation zone to be only the basal 4 mm. K⁺ depositionin the basal 6 mm was similar for seedlings with or withoutexogenous K⁺, at 0·6–0·87 µmolg^–1 f. wt h^–1. Deposition rates in zones above6 mm from the base were greater for seedlings with, thanwithout, exogenous K⁺; the latter were sometimes negative. Weconclude that for the coleoptiles of rice seedlings, anoxiainhibits net K⁺ uptake from the external solution to a muchlarger extent than K⁺ translocation from the caryopses. Furthermore,K⁺ concentrations in the elongation zone of the coleoptilesof anoxic seedlings were maintained to a remarkable degree,contributing to maintenance of _sap in cells of these elongatingtissues.     

Anoxic Responses of Seedling Roots of Rice Cultivars Varying in Tolerance to Submergence

Differences in tolerance to submergence and anoxia exhibited by cultivar-specific rice (Oryza sativa L.) extend to the primary root tips and axes of 3-day-old seedlings. This paper considers the physiological mechanisms which might account for rice root intolerance to anoxia, particularly those implicated in pH regulation and sugar metabolism in relation to hypoxic acclimation. Hypoxic treatment and the presence of glucose during anoxia did not permit root tips and axes of intolerant cultivars to survive 24-h anoxia. The absence of typical glycolytic and fermentative enzyme induction together with no improvement of ethanol production and energy status during anoxia suggest that intolerant cultivars are not capable of hypoxic acclimation at the level of energy and sugar metabolism. However, root tip survival was enhanced in buffered medium after hypoxic treatment, suggesting a relationship between hypoxic treatment and improved pH regulation.     

The Role of the Epidermis in the Control of Elongation Growth in Stems and Coleoptiles

The peripheral cell wall(s) of stems and coleoptiles are 6 to 20 times thicker than the walls of the inner tissues. In coleoptiles, the outer wall of the outer epidermis shows a multilayered, helicoidal cellulose architecture, whereas the walls of the parenchyma and the outer wall of the inner epidermis are unilayered. In hypocotyls and epicotyls both the epidermal and some subepidermal walls are multilayered, helicoidal structures. The walls of the internal tissues (inner cortex, pith) are unilayered, with cellulose microfibrils oriented primarily transversely. Peeled inner tissues rapidly extend in water, whereas the outer cell layer(s) contract on isolation. This indicates that the peripheral walls limit elongation of the intact organ. Experiments with the pressure microprobe indicate that the entire organ can be viewed as a giant, turgid cell: the extensible inner tissues exert a pressure (turgor) on the peripheral wall(s), which bear the longitudinal wall stress of the epidermal and internal cells. Numerous studies have shown that auxin induces elongation of isolated, intact sections by loosening of the growth-limiting peripheral cell wall(s). Likewise, the effect of light on reduction of stem elongation and cell wall extensibility in etiolated seedlings is restricted to the peripheral cell layers of the organ. The extensible inner tissues provide the driving force (turgor pressure), whereas the rigid peripheral wall(s) limit, and hence control, the rate of organ elongation.     

Preferential Induction of Alcohol Dehydrogenase in Coleoptiles of Rice Seedlings Germinated in Submergence Condition

Difference in the growth response to submergence between coleoptiles and roots of rice (Oryza sativa L.) was investigated in 9-d-old rice seedlings. The coleoptile length in the submergence condition was much greater than that in aerobic condition, whereas the root length in the submergence condition was less than that in the aerobic condition. Alcohol dehydrogenase (ADH) activity in the coleoptiles in the submergence condition was much greater than that in the aerobic condition, but ADH activity in the roots in the submergence condition increased slightly. These results suggest that the preferential ADH induction in rice seedlings may contribute to the difference in the growth response between the coleoptiles and roots under low oxygen conditions.     

Growth promotion and an increase in cell wall extensibility by silicon in rice and some other Poaceae seedlings

The effect of silicon on organ growth and its mechanisms of action were studied in rice (Oryza sativa L. cv. Koshihikari), oat (Avena sativa L. cv. Victory), and wheat (Triticum aestivum L. cv. Daichino-Minori) seedlings grown in the dark. Applying silicon in the form of silicic acid to these seedlings via culture solution resulted in growth promotion of third (rice) or second (oat and wheat) leaves. The optimal concentration of silicon was 5–10 mM. No growth promotion was observed in early organs, such as coleoptiles or first leaves. In silicon-treated rice third leaves, the epidermal cell length increased, especially in the basal regions, without any effect on the number of cells, showing that silicon promoted cell elongation but not cell division. Silicon also increased the cell wall extensibility significantly in the basal regions of rice third leaves. These results indicate that silicon stimulates growth of rice and some other Poaceae leaves by increasing cell wall extensibility. Received: July 31, 2001 / Accepted: September 18, 2001     

Exogenous Nitrate as a Terminal Acceptor of Electrons in Rice (Oryza sativa) Coleoptiles and Wheat (Triticum aestivum) Roots under Strict Anoxia

To elucidate the physiological role of exogenous nitrate under anaerobic conditions, we studied the effect of 10 mM KNO₃ on the mitochondrial ultrastructure in rice (Oryza sativa L.) coleoptiles and in wheat (Triticum aestivum L.) roots, detached from four-day-old seedlings, under strict anoxia. In wheat roots, following 6-h-long anoxia in the absence of exogenous nitrate, the mitochondrial membranes were partially degraded and, after 9 h under anoxia, the mitochondrial membranes and the membranes of other organelles were completely destroyed. In rice coleoptiles, the partial membrane degradation was observed only after 24 h and their complete breakdown after 48 h of anaerobiosis. In the presence of exogenous nitrate, no membrane destruction was noticed even after 9 and 48 h of anaerobiosis in wheat roots and rice coleoptiles, respectively. These results indicate that exogenous nitrate exerts protective action as a terminal electron acceptor, alternative to the molecular oxygen. Our findings are compared with the results of other researchers concerning the adverse or favorable nitrate action on plant growth, metabolism, and energy status under anaerobic stress.     

Light-induced increase in the contents of ferulic and diferulic acids in cell walls of Avena coleoptiles: its relationship to growth inhibition by light

White fluorescent light (5 W m⁻²) inhibited Avena coleoptile growth. Light caused in increase in minimum stress relaxation time and a decrease in extensibility (strain/load) of coleoptile cell walls. Light increased the contents of ferulic acid (FA) and diferulic acid (DFA) ester-linked to the hemicellulose I in cell walls. These changes in the phenolic contents correlated with those of the mechanical properties of cell walls, suggesting that light stimulates the formation of DFA in hemicellulose I, making cell walls rigid, and thus results in growth inhibition. The ratio of DFA to FA was almost constant in the dark, but decreased in light, although it was almost constant in Oryza coleoptiles either in the dark or in light (Tan et al. 1992). From this fact, it is speculated that in the light condition, the formation of DFA in cell walls is limited in the step of the peroxidase catalyzed coupling reaction to produce DFA, while in the dark it is limited in the step of the feruloylation of hemicellulose I.     

Correlation between cell wall extensibility and the content of diferulic and ferulic acids in cell walls of Oryza sativa coleoptiles grown under water and in air

Rice ( Oryza sativa L. cv. Sasanishiki) coleoptiles grown under water achieved greater length than those grown either in air or under water with constant air bubbling. The extensibility of cell walls in coleoptiles grown under water was larger than that in the other treatments. Per unit length of the coleoptile, the content of ferulic and diferulic acids ester-linked to hemicelluloses was higher in air and bubbling type coleoptiles than in water type ones. The extensibility of the coleoptile cell walls correlated with the content of diferulic acids per unit length and per hemicellulose, suggesting that the enhancement of the formation of diferulic acid bridges in hemicelluloses in air or under water with air bubbling makes the cell walls mechanically rigid; thereby inhibiting cell elongation in rice coleoptiles. In addition, the ratio of diferulic acid to ferulic acid was almost constant irrespective of coleoptile age, zone and growth conditions, suggesting that the feruloylation of hemicelluloses is rate-limiting in the formation of diferulic acid bridges in the cell walls of rice coleoptiles.     

Overexpression of OsJAC1, a Lectin Gene, Suppresses the Coleoptile and Stem Elongation in Rice

Lectin plays an Important role In defense signaling In plants, but its function In plant growth and development is not well known. Previously, we cloneds rice （Oryza sativa L.） gene OsJAC1 encoding s msnnose-blndlng Jscslln-relsted lectln, and found that OsJAC1 was Jssmonlc acid （JA） Inducible. Here we cloned the promoter of OsJAC1, and GUS activity was detected In young roots, coleoptlles, sheaths, leaves, nodes of stems, stems, rschlses, pistils, stsmens and lemmss of OsJAC1：：GUS trsnsgenlc rice, suggesting that OsJAC1 Is s constitutive expression gene In rice. Moreover, OsJAC1-overexpressed （Ubi：：OsJAC1） rice showed dwarfism with shorter coleptlles resulting from the failure of cell elongation of coleoptlles. In addition, compared with coleoptlles of wild-type plants, those of OsJAC1 overexpresslon rice were more sensitive to JA treatment. These data revealed that, besides Its roles in defense response, lectin plays an Important role in rice growth and development.     

Genome-Wide Identification of DUF26 Domain-Containing Genes in Dongxiang Wild Rice and Analysis of Their Expression Responses under Submergence

The DUF26 domain-containing protein is an extracellular structural protein, which plays an important role in signal transduction. Dongxiang wild rice (Oryza rufipogon Griff.) is the northern-most common wild rice in China. Using domain analysis, 85 DUF26 domain-containing genes were identified in Dongxiang wild rice (DXWR) and further divided into four categories. The DUF26 domain-containing genes were unevenly distributed on chromosomes, and there were 18 pairs of tandem repeats. Gene sequence analysis showed that there were significant differences in the gene structure and motif distribution of the DUF26 domain in different categories. Motifs 3, 8, 9, 13, 14, 16, and 18 were highly conserved in all categories. It was also found that there were eight plasmodesmata localization proteins (PDLPs) with a unique motif 19. Collinearity analysis showed that DXWR had a large number of orthologous genes with wheat, maize, sorghum and zizania, of which 17 DUF26 domain-containing genes were conserved in five gramineous crops. Under the stress of anaerobic germination and seedling submergence treatment, 33 DUF26 domain-containing genes were differentially expressed in varying degrees. Further correlation analysis with the expression of known submergence tolerance genes showed that these DUF26 domain-containing genes may jointly regulate the submergence tolerance process with these known submergence tolerance genes in DXWR.     

Proteomic Analysis of the Response of Liangyoupeijiu （Super High-Yield Hybrid Rice） Seedlings to Cold Stress

Liangyoupeijiu is a super high-yield hybrid rice. Despite its advantages with respect to yield and grain quality, it is sensitive to cold, which keeps it from being widely cultivated. We subjected Liangyoupeijiu seedlings to 4 ℃ cold treatment, then extracted the leaf proteins. After 2-D gel electrophoresis separation and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis, a series of differentially displayed proteins were identified. Some metabolism-associated proteins were found among the downregulated proteins, such as carbamoyl phosphate synthetase, transketolase 1, dihydrolipoamide dehydrogenase and glyceraldehyde 3-phosphate dehydrogenase. The upregulated proteins included both stress-resistance proteins such as nucleoside diphosphate kinase I and proteins that are negative for rice growth, such as FtsH-like protein, plastid fusion and/or translocation factor （Pftf） and actin. Our results indicate that cold may inhibit Liangyoupeijiu growth through decreasing metabolic activity and damaging cell structure.