Golgi/plastid‐type manganese superoxide dismutase involved in heat‐stress tolerance during grain filling of rice

Superoxide dismutase (SOD) is widely assumed to play a role in the detoxification of reactive oxygen species caused by environmental stresses. We found a characteristic expression of manganese SOD 1 (MSD1) in a heat‐stress‐tolerant cultivar of rice (Oryza sativa). The deduced amino acid sequence contains a signal sequence and an N‐glycosylation site. Confocal imaging analysis of rice and onion cells transiently expressing MSD1‐YFP showed MSD1‐YFP in the Golgi apparatus and plastids, indicating that MSD1 is a unique Golgi/plastid‐type SOD. To evaluate the involvement of MSD1 in heat‐stress tolerance, we generated transgenic rice plants with either constitutive high expression or suppression of MSD1. The grain quality of rice with constitutive high expression of MSD1 grown at 33/28 °C, 12/12 h, was significantly better than that of the wild type. In contrast, MSD1‐knock‐down rice was markedly susceptible to heat stress. Quantitative shotgun proteomic analysis indicated that the overexpression of MSD1 up‐regulated reactive oxygen scavenging, chaperone and quality control systems in rice grains under heat stress. We propose that the Golgi/plastid MSD1 plays an important role in adaptation to heat stress.     

水稻发芽期和幼苗前期耐碱性的鉴定方法研究

通过采用4种不同浓度的Na2CO3模拟碱胁迫,以水稻发芽期发芽势和发芽率的相对碱害率以及幼苗前期的根数、根长和苗高的相对碱害率为鉴定评价指标,研究水稻发芽期和幼苗前期耐碱性鉴定的碱胁迫条件。结果表明,水稻发芽势对碱胁迫的反应较发芽率更为敏感;根长对碱胁迫的反应较苗高和根数更为敏感;幼苗前期对碱胁迫的反应较发芽期更为敏感。在0．15％或0．20％Na2CO3胁迫下水稻发芽势和发芽率的相对碱害率平均值、方差和变异幅度较大。且其分布均匀合理,可作为水稻发芽期耐碱性鉴定的碱胁迫条件;而在0．10％或0．15％Na2CO3胁迫下,幼苗前期根数、根长和苗高的相对碱害率以及综合相对碱害率的平均值、方差和变异幅度较大,且分布相对均匀合理,可作为水稻幼苗前期耐碱性鉴定的碱胁迫条件。在碱胁迫下,水稻发芽势、发芽率、根数、根长和苗高的表型值分别与它们自身的相对碱害率呈极显著的负相关;发芽率的相对碱害率与根数、根长和苗高的相对碱害率呈显著的正相关;发芽期的综合相对碱害率与幼苗前期的综合相对碱害率呈显著的正相关;因此,以水稻发芽期的耐碱性可间接判断水稻幼苗前期的耐碱性。     

The use of comparative quantitative proteomics analysis in rice grain-filling in determining response to moderate soil drying stress

Moderate soil drying (MSD) stress at the grain filling stage can improve grain filling efficiently and thus increase grain yield. To elucidate the molecular response of grain filling to MSD stress, a labeling LC-based quantitative proteomics approach using tandem mass tags was applied to determine the changes in leaf and grain protein abundance level at 15 days after flowering. A total of 2109 leaf proteins and 3220 grain proteins were detected, and 251 leaf proteins and 220 grain proteins were differentially expressed under MSD stress. Based on MapMan ontology, differentially expressed proteins in leaf and grain were categorized within 22 and 18 functional categories, respectively. The patterns observed were interesting in that in some categories such as photosynthesis-related protein in leaf and cell division related proteins in grain showed higher expression abundant under MSD stress, which facilities increasing the source supply and sink size. In other categories, such as carbohydrate metabolism and mitochondrial electron transport, surprisingly showed a completely different expression pattern between leaf and grain under MSD stress, which led to faster and better remobilization of carbon from leaf to grain. Additionally, the complicated functional network including the small GTP-binding proteins, calmodulin, and 14-3-3 proteins play an important role in regulation carbon remobilization mediated by the stressful signals from soil after rice plants were treated with MSD at grain-filling stage. The findings provide theoretical evidence for better quality control and scientific improvement of rice in practice.     

来源于疣粒野生稻的白叶枯病新抗源的鉴定

疣粒野生稻对白叶枯病高抗甚至免疫。粳稻品种8411与疣粒野生稻体细胞杂交获得了2个高抗所有栽培稻白叶枯病抗性基因均不能抵御的国际广致病菌系P6的新种质SH5和SH76,遗传分析鉴定出新种质含有1个抗P6小种的显性基因。本研究以高感白叶枯病水稻品种IR24及携带不同抗性基因的16个材料为参照,对分蘖期、孕穗期的SH5和SH76分别接种11个白叶枯病小种,抗性分析表明SH5和SH76的抗谱广,与IRBB21(Xa21)抗谱一致,与IRBB5(xa5)、IRBB7(Xa7)和Asominori(Xa17)的较相近。用Xa21的分子标记pTA248和XA21检测,确定SH5和SH76不携带Xa21基因,前期研究结果证实新种质中不含xa5和Xa7;与Asominori的杂交试验表明其抗性基因与Xa17基因不等位。这些结果表明SH5和SH76中存在1个抗P6小种的新基因。     

培矮64S/爪哇稻F1农艺性状表现和对照优势分析

本文报道了利用原始爪哇稻资源与光温敏雄性不育系培矮64S配制的27个籼爪交组合在长沙的农艺性状和杂种优势表现.从总体上来看,籼爪交组合与对照相比在每穗实粒数和理论产量上无显著差异,在其它性状上均有极显著的差异;籼爪交组合在秆高、穗长、每穗总粒数、每穗实粒数和千粒重方面有正向对照优势,在播始历期、有效分蘖数、结实率、理论产量和实际产量上存在负向对照优势.从个体上来看,籼爪交组合理论产量对照优势＞40%的比例为11.1%.实际产量对照优势＞40%的机率为3.7%,说明爪哇稻资源在籼爪交杂种优势利用中具有利用价值.本文还对爪哇稻资源在籼爪杂种优势利用中的一些问题进行了讨论.     

Production of a highly asymmetric somatic hybrid between rice and Zizania latifolia (Griseb): evidence for inter-genomic exchange

 A highly asymmetric and fertile somatic hybrid plant was obtained via protoplast fusion in an intergenric combination. Gamma-ray-irradiated Zizania latifolia (Griseb). Turcz. ex Stapf mesophyll protoplasts were electrofused with idoacetamide-inactivated rice protoplasts derived from a 2-month-old suspension cell culture. Two of the six putative hybrid calli regenerated plants. Cytological observation showed that the somatic chromosome numbers of both plants were the same as the rice parent (2n=24). Nevertheless, the hybrid nature and inter-genomic exchange events of one of the plants, i.e. SH6 (SH for somatic hybrid), were confirmed by Southern analysis using both total genomic DNA and moderate-copy, Z. latifolia-abundant DNA sequences as probes; in both cases, parental specific and/or new intergenomic recombinant hybridization fragments were detected. In both plant and seed morphology, the hybrid (SH6) was distinct from its rice parental cultivar, as well as from the wild donor species, Z. latifolia. Received: 15 August 1998 / Accepted: 30 September 1998     

水稻白叶枯病候选抗性基因SHNLR的RGAs克隆及分析

旨在从含有疣粒野生稻抗白叶枯病基因的新种质SH5、SH76基因组中克隆抗病基因。利用RGAs法得到1个NBS-LRR类同源基因,暂命名为SHNLR(登录号为JF934724)。结果表明,SHNLR的开放阅读框长度为3 105 bp,编码1 034个氨基酸,含有CC、NB-ARC与LRR结构域,具备CC-NBS-LRR类植物抗病基因的结构特征。BLASTn和BLASTp比对显示SHNLR是单拷贝基因,未发现同源性较高且功能已知的基因,仅NBS保守域序列与番茄Prf基因的相似度最高。对SHNLR基因电子定位,发现其位于水稻第11号染色体的长臂末端,但与11号染色体上已定位或克隆的8个白叶枯病抗性基因具有不同序列或处于不同的位置。半定量RT-PCR分析表明,SHNLR在抗病新种质叶片中的表达明显受到白叶枯病菌Zhe173的诱导。因此推测SHNLR可能是1个与抗白叶枯病相关的R基因。     

Hydrophobicity of stored (15, 35 °C), or dry-heated (120 °C) rice flour and deteriorated breadmaking properties baked with these treated rice flour/fresh gluten flour

Rice flour was stored at 15 °C/9 months, at 35 °C/14 days, or dry-heated at 120 °C/20 min. The breadmaking properties baked with this rice flour/fresh gluten flour deteriorated. In addition, the rice flour was mixed with oil in water vigorously, and oil-binding ability was measured. Every rice flour subjected to storage or dry-heated at 120 °C showed higher hydrophobicity, owing to changes in proteins. Then, proteins in the stored rice flour were excluded with NaOH solution, and bread baked with the deproteinized rice flour showed the same breadmaking properties as unstored rice flour/fresh gluten flour. The viscoelasticity of wheat glutenin fraction decreased after the addition of dry-heated rice flour in a mixograph profile. DDD staining increased Lab in color meter, which suggested an increase in SH groups in rice protein. The increase in SH groups caused a reduction in wheat gluten protein resulting in a deterioration of rice bread quality.     

超高产杂交水稻皮秒分辨荧光特性

采用时间分辨荧光光谱测试系统,研究了超高产杂交水稻(Oryza sativa L.)两优培九(P9)和对照汕优63(SH 63)类囊体膜的荧光光谱特性和时间特性.以脉宽为120 ps,重复率为4MHz,波长为514 nm的Ar+激光分别激发P9和SH 63水稻类囊体膜荧光.通过对其超快荧光的时间特性和光谱特性比较研究发现:无论是在灌浆期还是在扬花期,P9水稻类囊体膜中光系统I激发能传递的速度比光系统Ⅱ的快;P9和SH 63两种水稻类囊体膜在灌浆期的激发能传递速度都比扬花期的快;两种水稻类囊体膜的光谱特性还给出,在从扬花期到灌浆期这一生长发育过程中,SH 63水稻类囊体膜的色素成分和结合状态发生了变化,而P9却没有出现这种变化.     

Development and characterization of a simple perfused oral microcosm

AIMS: To validate perfused, inline, filter-based fermentation systems (multiple Sorbarod devices, MSD) for their ability to maintain stable oral bacterial communities. MSD enable replicate (n=5) microcosm biofilms (BF) to be established and sampled, together with their perfusates (PA, cells in eluted medium). METHODS AND RESULTS: Fresh saliva from human volunteers was used to inoculate MSD, incubated in an anaerobic cabinet and perfused with artificial saliva at 7 ml h(-1). BF within Sorbarod filters and cells eluted in the PA were analysed at 24-h intervals by differential bacteriological culture and checkerboard DNA-DNA hybridization (CKB, 40 oral species). Dynamic stability was apparent after 2-3 days within both BF and PA as evidenced by culture, CKB data and pH measurements. BF harboured large numbers of anaerobic species and facultative anaerobes [ca 10-11 log10 colony-forming units (CFU)/filter] comprising considerable numbers of streptococci and Gram-negative species. PA contained ca 9-10 log(10) CFU ml(-1) suggesting an apparent mean growth rate of 0.1 h(-1) for the BF, as a whole corresponding to a mean generation time of 10 h. CKB analysis revealed considerable bacterial diversity within the respective MSD. Inter-individual variations in the relative species abundance of inocula was broadly reproduced in the MSD (BF and PA), although considerable variation was apparent between triplicate models established using saliva from one saliva donor or from three individual donors. The dominance of Gram-negative species, indicated by culture was supported by CKB analysis (major species, Prevotella melaninogenica and Fusobacterium nucleatum). CONCLUSIONS: Data obtained from the various analytical approaches showed a high degree of congruence. The MSD enables the maintenance of complex, stable salivary microcosms and represents a simple, reproducible tool for modelling individual oral bacterial ecosystems. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrates the utility of the MSD for studying the micro-ecology of the oral cavity.     

Force-Regulated Refolding of the Mechanosensory Domain in the Platelet Glycoprotein Ib-IX Complex

In platelets, the glycoprotein (GP) Ib-IX receptor complex senses blood shear flow and transmits the mechanical signals into platelets. Recently, we have discovered a juxtamembrane mechanosensory domain (MSD) within the GPIbα subunit of GPIb-IX. Mechanical unfolding of the MSD activates GPIb-IX signaling into platelets, leading to their activation and clearance. Using optical tweezer-based single-molecule force measurement, we herein report a systematic biomechanical characterization of the MSD in its native, full-length receptor complex and a recombinant, unglycosylated MSD in isolation. The native MSD unfolds at a resting rate of 9 × 10^?3 s^?1. Upon exposure to pulling forces, MSD unfolding accelerates exponentially over a force scale of 2.0 pN. Importantly, the unfolded MSD can refold with or without applied forces. The unstressed refolding rate of MSD is ～17 s^?1 and slows exponentially over a force scale of 3.7 pN. Our measurements confirm that the MSD is relatively unstable, with a folding free energy of 7.5 k_BT. Because MSD refolding may turn off GPIb-IX’s mechanosensory signals, our results provide a mechanism for the requirement of a continuous pulling force of >15 pN to fully activate GPIb-IX.     

Effects of silkworm hemolymph on cell viability and hCTLA4Ig production in transgenic rice cell suspension cultures

Silkworm hemolymph (SH), prepared from fifth-instar larvae of Bombyx mori and heat-treated at 60 degrees C for 30 min, was used to improve cell viability and the production of human cytotoxic T-lymphocyte antigen 4-immunoglobulin (hCTLA4Ig) in transgenic Oryza sativa L. cell suspension cultures. Even though SH could not elevate cell viability at the concentrations up to 3% (v/v), addition of 0.3% (v/v) SH to a culture medium enhanced the production of hCTLA4Ig by 36.8% over an SH-free medium. Moreover, the production period of hCTLA4Ig could be shortened in a 0.3% (v/v) SHadded medium compared with that in an SH-free culture. As a result, addition of 0.3% (v/v) SH improved the productivity of hCTLA4Ig significantly in transgenic rice cell cultures.     

Characterisation of grids of point detectors in maximum skin dose measurement in fluoroscopically-guided interventional procedures

PurposePoint detectors are frequently used to measure patient's maximum skin dose (MSD) in fluoroscopically-guided interventional procedures (IP). However, their performance and ability to detect the actual MSD are rarely evaluated. The present study investigates the sampling uncertainty associated with the use of grids of point detectors to measure MSD in IP.MethodChemoembolisation of the liver (CE), percutaneous coronary intervention (PCI) and neuroembolisation (NE) procedures were studied. Spatial dose distributions were measured with XR-RV3 Gafchromic^® films for 176 procedures. These distributions were used to simulate measurements performed using grids of detectors such as thermoluminescence detectors, with detector spacing from 1.4 up to 10 cm.ResultsThe sampling uncertainty was the highest in PCI and NE procedures. With 40 detectors covering the film area (36 cm × 44 cm), the maximum dose would be on average 86% and 63% of the MSD measured with Gafchromic^® films in CE and PCI procedures, respectively. In NE procedures, with 27 detectors covering the film area (14 cm × 35 cm), the maximum dose measured would be on average 82% of the MSD obtained with the Gafchromic^® films.ConclusionThermoluminescence detectors show good energy and dose response in clinical beam qualities. However the poor spatial resolution of such point-like dosimeters may far outweigh their good dosimetric properties. The uncertainty from the sampling procedure should be estimated when point detectors are used in IP because it may lead to strong underestimation of the MSD.     

Genetic analysis and molecular mapping of a nuclear recessive male sterility gene, ms91(t), in rice.

Mutations that result in plant male sterility provide means not only to probe reproductive development but also to facilitate commercial heterosis application and hybrid seed production. In this study, we report a novel male sterility gene, ms91(t), in a spontaneous mutant line (SH38) from a Chinese rice cultivar (Oryza sativa subsp. japonica 'Jijing14'). The sterility of SH38 was studied by examining its progenies derived from crosses with 6 japonica cultivars. Corresponding F2 populations were obtained by selfing each of the 6 F1s and a backcross population was produced by crossing SH38 to the F1 of SH38 x C18. Our results revealed that SH38 has normal agronomic traits but produces no pollen grains. Segregations of male-sterile and male-fertile progenies in the F2 and backcross populations fit well with ratios of 3:1 and 1:1, respectively, indicating that ms91(t) is a single recessive gene. Amplified fragment length polymorphism (AFLP) analysis of SH38 and Jijing14 plants showed the presence of a unique band in SH38. Simple sequence repeat (SSR) analysis of the bulked and individual progenies of the F2 population of SH38 x C18 showed linkage of ms91(t) with the SSR marker RM5853 on chromosome 1. Subsequently, ms91(t) was fine-mapped to the interval between markers RM7075 (3.75 cM) and RM5638 (3.57 cM). Our results would facilitate the isolation of ms91(t) and male sterility in heterosis application.     

Transpositional activation of <Emphasis Type="Italic">mPing</Emphasis> in an asymmetric nuclear somatic cell hybrid of rice and <Emphasis Type="Italic">Zizania latifolia</Emphasis> was accompanied by massive element loss

We have reported previously that the most active miniature inverted terminal repeat transposable element (MITE) of rice, mPing, was transpositionally mobilized in several rice recombinant inbred lines (RILs) derived from an introgressive hybridization between rice and wild rice (Zizania latifolia Griseb.). To further study the phenomenon of hybridization-induced mPing activity, we undertook the present study to investigate the element’s behavior in a highly asymmetric somatic nuclear hybrid (SH6) of rice and Z. latifolia, which is similar in genomic composition to that of the RILs, though probably contains more introgressed alien chromatins from the donor species than the RILs. We found that mPing, together with its transposase-donor, Pong, underwent rampant transpositional activation in the somatic hybrid (SH6). Because possible effects of protoplast isolation and cell culture can be ruled out, we attribute the transpositional activation of mPing and Pong in SH6 to the process of asymmetric somatic hybridization, namely, one-step introgression of multiple chromatin segments of the donor species Z. latifolia into the recipient rice genome. A salient feature of mPing transposition in the somatic hybrid is that the element’s activation was accompanied by massive loss of its original copies, i.e., abortive transpositions, which was not observed in previously reported cases of mPing activity. These data not only corroborated our earlier finding that wide hybridization and introgression may trigger transpositional activation of otherwise quiescent transposable elements, but also suggest that transpositional mobilization of a MITE like mPing can be accompanied by dramatic reduction of its original copy numbers under certain conditions, thus provide novel insights into the dynamics of MITEs in the course of genome evolution.     

Heavy genetic load associated with the subspecific differentiation of japonica rice (Oryza sativa ssp. japonica L.)

Genetic load in the genome of the model species, rice, was genetically dissected by mapping quantitative trait loci (QTLs) affecting the radiosensitivity of 226 recombinant inbred lines (RILs) to gamma-ray- and spaceflight-induced radiation. The parents and RILs varied considerably in their radiosensitivity to gamma-ray irradiation. A total of 28 QTLs affecting the two index traits, seedling height (SH) and seed fertility (SF), of radiosensitivity were identified. The japonica parent, Lemont, was much more sensitive to gamma-ray irradiation than the indica parent, Teqing, and its alleles at almost all QTLs were associated with increased radiosensitivity, suggesting a much higher genetic load in the japonica genome of rice. Six QTLs (QSh2a, QSh2b, QSh5a, QSh7, QSf3b, and QSf10b) were located in the genomic regions particularly sensitive to radiation and thus might represent possible 'mutation hot spots' in the japonica genome. Detailed characterization of these genomic regions may shed light on the evolution and subspecific differentiation of rice.     

NCAM polypeptides in heart development: association with Z discs of forms that contain the muscle-specific domain

Previous studies of neural cell adhesion molecule (NCAM) cDNAs have revealed an alternatively spliced set of small exons (12A, 12B, 12C, and 12D) that encode a region in the extracellular portion of the molecule known as the muscle-specific domain (MSD). The entire MSD region can be expressed in skeletal muscle, heart, and skin; only exons 12A and 12D have been found in brain. These studies did not reveal which NCAM polypeptides contain the MSD region or the immunohistochemical distribution of these NCAM molecules. To address these questions, we prepared antibodies against the oligopeptides encoded by exons 12A and 12B and by exons 12C and 12D, and we used these antibodies to study the forms of NCAM containing the MSD region expressed during embryonic chicken heart development. These antibodies recognize certain forms of NCAM found in the heart, but they do not recognize brain NCAM. In the heart, each of the splice variants of NCAM (large cytoplasmic domain, small cytoplasmic domain, and small surface domain) that differ in their mode of attachment to the plasma membrane or in the size of their cytoplasmic domain is expressed in a form that contains and in a form that lacks the MSD region. No microheterogeneity is observed in the size of NCAM molecules containing the MSD region, even at the level of cyanogen bromide fragments, suggesting that exons 12A-D are expressed as a single unit. Depending on the site and the stage of development, the percent of NCAM molecules containing the MSD region can vary from nearly 0 to 100%. In general, this percentage increases during development. In immunohistochemical studies of hearts from stage 18 embryos, forms of NCAM containing the MSD region colocalized with Z discs. No other adhesion molecules were found in this distribution at this early stage of development. Studies on isolated cells in vitro demonstrate that the colocalization with Z discs of NCAM molecules containing the MSD region does not depend on cell-cell contact, and they raise the possibility that this form of NCAM is involved in cell-extracellular matrix interactions. The association of NCAM molecules containing the MSD region with Z discs suggests that this form of NCAM is involved in early myofibrillogenesis.