Sensitivity of rice to ultraviolet-B radiation

BACKGROUND: Depletion of the stratospheric ozone layer leads to an increase in ultraviolet-B (UVB: 280-320 nm) radiation reaching the earth's surface, and the enhanced solar UVB radiation predicted by atmospheric models will result in reduction of growth and yield of crops in the future. Over the last two decades, extensive studies of the physiological, biochemical and morphological effects of UVB in plants, as well as the mechanisms of UVB resistance, have been carried out. SCOPE: In this review, we describe recent research into the mechanisms of UVB resistance in higher plants, with an emphasis on rice (Oryza sativa), one of the world's most important staple food crops. Recent studies have brought to light the following remarkable findings. UV-absorbing compounds accumulating in the epidermal cell layers have traditionally been considered to function as UV filters, and to play an important role in countering the damaging effects of UVB radiation. Although these compounds are effective in reducing cyclobutane pyrimidine dimer (CPD) induction in plants exposed to a challenge exposure to UVB, certain levels of CPD are maintained constitutively in light conditions containing UVB, regardless of the quantity or presence of visible light. These findings imply that the systems for repairing DNA damage and scavenging reactive oxygen species (ROS) are essential for plants to grow in light conditions containing UVB. CONCLUSION: CPD photolyase activity is a crucial factor determining the differences in UVB sensitivity between rice cultivars. The substitution of one or two bases in the CPD photolyase gene can alter the activity of the enzyme, and the associated resistance of the plant to UVB radiation. These findings open up the possibility, in the near future, of increasing the resistance of rice to UVB radiation, by selective breeding or bioengineering of the genes encoding CPD photolyase.     

Increase in CPD photolyase activity functions effectively to prevent growth inhibition caused by UVB radiation

Rice cultivars vary widely in their sensitivity to ultraviolet B (UVB) and this has been correlated with cyclobutane pyrimidine dimer (CPD) photolyase mutations that alter the structure/function of this photorepair enzyme. Here, we tested whether CPD photolyase function determines the UVB sensitivity of rice (Oryza sativa) by generating transgenic rice plants bearing the CPD photolyase gene of the UV-resistant rice cultivar Sasanishiki in the sense orientation (S-B and S-C lines) or the antisense orientation (AS-D line). The S-B and S-C plants had 5.1- and 45.7-fold higher CPD photolyase activities than the wild-type, respectively, were significantly more resistant to UVB-induced growth damage, and maintained significantly lower CPD levels in their leaves during growth under elevated UVB radiation. Conversely, the AS-D plant had little photolyase activity, was severely damaged by elevated UVB radiation, and maintained higher CPD levels in its leaves during growth under UVB radiation. Notably, the S-C plant was not more resistant to UVB-induced growth inhibition than the S-B plant, even though it had much higher CPD photolyase activity. These results strongly indicate that UVB-induced CPDs are one of principal causes of UVB-induced growth inhibition in rice plants grown under supplementary UVB radiation, and that increasing CPD photolyase activity can significantly alleviate UVB-caused growth inhibition in rice. However, further protection from UVB-induced damage may require the genetic enhancement of other systems as well.     

Cyclobutane pyrimidine dimer (CPD) photolyase repairs ultraviolet-B-induced CPDs in rice chloroplast and mitochondrial DNA

Plants use sunlight as energy for photosynthesis; however, plant DNA is exposed to the harmful effects of ultraviolet‐B (UV‐B) radiation (280–320 nm) in the process. UV‐B radiation damages nuclear, chloroplast and mitochondrial DNA by the formation of cyclobutane pyrimidine dimers (CPDs), which are the primary UV‐B‐induced DNA lesions, and are a principal cause of UV‐B‐induced growth inhibition in plants. Repair of CPDs is therefore essential for plant survival while exposed to UV‐B‐containing sunlight. Nuclear repair of the UV‐B‐induced CPDs involves the photoreversal of CPDs, photoreactivation, which is mediated by CPD photolyase that monomerizes the CPDs in DNA by using the energy of near‐UV and visible light (300–500 nm). To date, the CPD repair processes in plant chloroplasts and mitochondria remain poorly understood. Here, we report the photoreactivation of CPDs in chloroplast and mitochondrial DNA in rice. Biochemical and subcellular localization analyses using rice strains with different levels of CPD photolyase activity and transgenic rice strains showed that full‐length CPD photolyase is encoded by a single gene, not a splice variant, and is expressed and targeted not only to nuclei but also to chloroplasts and mitochondria. The results indicate that rice may have evolved a CPD photolyase that functions in chloroplasts, mitochondria and nuclei, and that contains DNA to protect cells from the harmful effects of UV‐B radiation.     

Ultraviolet-B sensitivities in Japanese lowland rice cultivars: cyclobutane pyrimidine dimer photolyase activity and gene mutation

There is a cultivar difference in the response to ultraviolet-B(UVB: 280–320 nm) in rice (Oryza sativa L.). AmongJapanese lowland rice cultivars, Sasanishiki, a leading Japaneserice cultivar, is resistant to the damaging effects of UVB whileNorin 1, a close relative, is less resistant. We found previouslythat Norin 1 was deficient in cyclobutane pyrimidine dimer (CPD)photorepair ability and suggested that the UVB sensitivity inrice depends largely on CPD photorepair ability. In order toverify that suggestion, we examined the correlation betweenUVB sensitivity and CPD photolyase activity in 17 rice cultivarsof progenitors and relatives in breeding of UV-resistant Sasanishikiand UV-sensitive Norin 1. The amino acid at position 126 ofthe deduced amino acid sequence of CPD photolyase in cultivarsincluding such as Norin 1 was found to be arginine, the CPDphotolyase activities of which were lower. The amino acid atthat position in cultivars including such as Sasanishiki wasglutamine. Furthermore, cultivars more resistant to UVB werefound to exhibit higher photolyase activities than less resistantcultivars. These results emphasize that single amino acid alterationfrom glutamine to arginine leads to a deficit of CPD photolyaseactivity and that CPD photolyase activity is one of the mainfactors determining UVB sensitivity in rice. ¹ These authors contributed equally to the paper. ² Corresponding author: E-mail, kumagai{at}ige.tohoku.ac.jp; Fax,+81-22-217-5691.     

Assay of DNA Photolyase Activity in Spinach Leaves in Relation to Cell Compartmentation-Evidence for Lack of DNA Photolyase in Chloroplasts

Spinach cyclobutane pyrimidine dimer (CPD)-specific DNA photolyase was successfully detected in leaf extracts by an assay system for plant photolyase using an improved enzyme-linked immunosorbent assay (ELISA) which was newly introduced by novel horseradish peroxidase (HRP)-linked CPD specific monoclonal antibodies. The assay system includes two main steps: a photorepair reaction of CPD introduced in substrate DNA and measurement of CPD remained after the photorepair by the improved ELISA. When CPD- induced salmon sperm DNA was used as a substrate, high CPD-photolyase activities were observed in the enzyme fraction prepared from whole spinach leaf extracts, but not from chloroplast extracts. This strongly suggests that spinach CPD-specific photolyases are localized in cell compartments other than chloroplasts.     

Thermodynamics and kinetics for base pair opening in the DNA decamer duplexes containing cyclobutane pyrimidine dimer

The cyclobutane pyrimidine dimer (CPD) is one of the major classes of cytotoxic and carcinogenic DNA photoproducts induced by UV light. Hydrogen exchange rates of the imino protons were measured for various CPD-containing DNA duplexes to better understand the mechanism for CPD recognition by XPC-hHR23B. The results here revealed that double T·G mismatches in a CPD lesion significantly destabilized six consecutive base pairs compared to other DNA duplexes. This flexibility in a DNA duplex caused at the CPD lesions with double T·G mismatches might be the key factor for damage recognition by XPC-hHR23B.     

Cooler temperatures slow the repair of DNA damage in tadpoles exposed to ultraviolet radiation: Implications for amphibian declines at high altitude

Ultraviolet B radiation (UVBR) damages the DNA of exposed cells, causing dimers to form between adjacent pyrimidine nucleotides. These dimers block DNA replication, causing mutations and apoptosis. Most organisms utilize biochemical or biophysical DNA repair strategies to restore DNA structure; however, as with most biological reactions, these processes are likely to be thermally sensitive. Tadpoles exposed to elevated UVBR at low environmental temperatures have significantly higher rates of mortality and developmental deformities compared with tadpoles exposed to the same levels of UVBR at higher environmental temperatures. We hypothesized that low environmental temperatures impair the primary enzymatic (photolyase) DNA repair pathway in amphibians, leading to the accumulation of DNA damage. To test this hypothesis, we compared DNA repair rates and photolyase gene expression patterns in Limnodynastes peronii. Tadpoles were acutely exposed to UVBR for 1 hr at either 20 or 30°C, and we measured DNA damage and photolyase expression levels at intervals following this exposure. Temperature had a significant effect on the rate of DNA repair, with repair at 30°C occurring twice as fast as repair at 20°C. Photolyase gene expression (6‐4 PP and CPD) was significantly upregulated by UVBR exposure, with expression levels increasing within 6 hr of UVBR exposure. CPD expression levels were not significantly affected by temperature, but 6‐4 PP expression was significantly higher in tadpoles in the 30°C treatment within 12 hr of UVBR exposure. These data support the hypothesis that DNA repair rates are thermally sensitive in tadpoles and may explain why enigmatic amphibian declines are higher in montane regions where UVBR levels are naturally elevated and environmental temperatures are lower.     

The molecular basis of white pericarps in African domesticated rice: novel mutations at the Rc gene

Repeated phenotypic evolution can occur at both the inter- and intraspecific level and is especially prominent in domesticated plants, where artificial selection has favoured the same traits in many different species and varieties. The question of whether repeated evolution reflects changes at the same or different genes in each lineage can now be addressed using the domestication and improvement genes that have been identified in a variety of crops. Here, we document the genetic basis of nonpigmented ('white') pericarps in domesticated African rice (Oryza glaberrima) and compare it with the known genetic basis of the same trait in domesticated Asian rice (Oryza sativa). In some cases, white pericarps in African rice are apparently caused by unique mutations at the Rc gene, which also controls pericarp colour variation in Asian rice. In one case, white pericarps appear to reflect changes at a different gene or potentially a cis-regulatory region.     

水稻长穗颈基因eui紧密连锁SSR标记获得

02428h是从半矮秆材料02428体细胞培养后代中发现的隐性高秆突变体, 其株高性状由1对长穗颈基因eui和1对半矮秆基因sd-1共同控制。以02428h与半矮秆材料南京11杂交的F₂为作图群体, 利用Gramene公布的SSR标记和根据NCBI中的BAC序列自己新开发的SSR标记, 将eui基因定位在第5染色体上的RM3673和RM0012之间, 两侧遗传距离分别为0.3 cM和1.0 cM, 为该基因的分子标记辅助选择奠定了基础。     

家蝇介导的水稻基因逃逸风险评估(英文)

【目的】对转基因作物进行生态风险评估是大面积种植前的一个必要步骤,水稻Oryza sativa访花昆虫有上百种,包括家蝇Musca domestica。本研究旨在明确访花昆虫家蝇介导转基因水稻外源基因逃逸的风险。【方法】2010年,我们使用转基因水稻B1, B6和G8-7作为父本(花粉供体),用同源非转基因水稻Jiazao 935和Wuyunjing 7作为母本（花粉受体）,并用家蝇作为授粉昆虫,在浙江大学华家池和长兴试验基地开展了田间种植试验,对收割的后代水稻种子进行室内种植培养,对种苗用潮霉素B和草甘膦处理进行转基因杂交种检测,对存活植株进行潮霉素和草甘膦抗性基因PCR检测,测试家蝇介导的转基因水稻外源基因逃逸频率。【结果】对浙江两个测试基地3个转基因水稻品种共计超过216 500粒后代水稻种子进行的检测及结果表明,在毗邻区域杂交种少,家蝇授粉区和无家蝇授粉区转基因水稻外源基因向非转基因水稻逃逸频率均较低(0~0.64%)。【结论】家蝇介导的转基因水稻外源基因逃逸频率较低,家蝇没有增加转基因水稻外源基因逃逸的风险。     

A paralog of the MtN3/saliva family recessively confers race-specific resistance to Xanthomonas oryzae in rice

Approximately one third of the identified 34 rice major disease resistance (R) genes conferring race-specific resistance to different strains of Xanthomonas oryzae pv. oryzae (Xoo), which causes rice bacterial blight disease, are recessive genes. However, only two of the recessive resistance genes have been characterized thus far. Here we report the characterization of another recessive resistance gene, xa25, for Xoo resistance. The xa25, localized in the centromeric region of chromosome 12, mediates race-specific resistance to Xoo strain PXO339 at both seedling and adult stages by inhibiting Xoo growth. It encodes a protein of the MtN3/saliva family, which is prevalent in eukaryotes, including mammals. Transformation of the dominant Xa25 into a resistant rice line carrying the recessive xa25 abolished its resistance to PXO339. The encoding proteins of recessive xa25 and its dominant allele Xa25 have eight amino acid differences. The expression of dominant Xa25 but not recessive xa25 was rapidly induced by PXO339 but not other Xoo strain infections. The nature of xa25-encoding protein and its expression pattern in comparison with its susceptible allele in rice-Xoo interaction indicate that the mechanism of xa25-mediated resistance appears to be different from that conferred by most of the characterized R proteins.     

不同化感潜力水稻(Oryza sativa L.)苗期的氮素养分效率及相关基因的表达

为了阐明养分水平引起水稻(Oryza sativa L.)化感抑草潜力变化的生理生态机制,研究了不同N素营养处理下,不同化感潜力水稻苗期对N素营养逆境的响应特性及N素养分效率的差异,并运用实时荧光定量RT-PCR技术(FQ-PCR)检测与N素代谢和次生代谢关键酶的基因表达.结果表明:弱化感水稻品种Lemont对N素营养胁迫较敏感,强化感水稻品种PI312777对资源波动的适应性较强,N素养分效率较高.FQ-PCR分析结果显示,在低N条件下Lemont中的亚硝酸还原酶基因(nir),谷氨酰胺合成酶基因(gs)相对表达量均有不同幅度的下调, PI312777分别下调了1.2倍和1.4倍,而Lemont分别下调了3.0倍和1.8倍, Lemont下调的幅度分别是PI312777的2.5倍和1.3倍,但对于苯丙氨酸解氨酶基因(pal)与3-羟基-3甲基戊二酰辅酶A还原酶基因(hmgr)而言, PI312777叶组织中的pal和hmgr均上调表达,与对照相比上调了6.0倍和1.6倍,而Lemont中对应的基因均下调表达,分别下调了1.3倍和6.8倍,佐证了上述差异的分子生态学特性.     

A rice gene encoding glycosyl hydrolase plays contrasting roles in immunity depending on the type of pathogens

Because pathogens use diverse infection strategies, plants cannot use one-size-fits-all defence and modulate defence responses based on the nature of pathogens and pathogenicity mechanism. Here, we report that a rice glycoside hydrolase (GH) plays contrasting roles in defence depending on whether a pathogen is hemibiotrophic or necrotrophic. The Arabidopsis thaliana MORE1 (M agnaporthe o ryzae re sistance 1) gene, encoding a member of the GH10 family, is needed for resistance against M. oryzae and Alternaria brassicicola, a fungal pathogen infecting A. thaliana as a necrotroph. Among 13 rice genes homologous to MORE1, 11 genes were induced during the biotrophic or necrotrophic stage of infection by M. oryzae. CRISPR/Cas9-assisted disruption of one of them (OsMORE1a) enhanced resistance against hemibiotrophic pathogens M. oryzae and Xanthomonas oryzae pv. oryzae but increased susceptibility to Cochliobolus miyabeanus, a necrotrophic fungus, suggesting that OsMORE1a acts as a double-edged sword depending on the mode of infection (hemibiotrophic vs. necrotrophic). We characterized molecular and cellular changes caused by the loss of MORE1 and OsMORE1a to understand how these genes participate in modulating defence responses. Although the underlying mechanism of action remains unknown, both genes appear to affect the expression of many defence-related genes. Expression patterns of the GH10 family genes in A. thaliana and rice suggest that other members also participate in pathogen defence.     

Genetic and physical mapping of Pi37(t), a new gene conferring resistance to rice blast in the famous cultivar St. No. 1

The famous rice cultivar (cv.), St. No. 1, confers complete resistance to many isolates collected from the South China region. To effectively utilize the resistance, a linkage assay using microsatellite markers (SSR) was performed in the three F₂ populations derived from crosses between the donor cv. St. No. 1 and each of the three susceptible cvs. C101PKT, CO39 and AS20-1, which segregated into 3R:1S (resistant/susceptible) ratio, respectively. A total of 180 SSR markers selected from each chromosome equally were screened. The result showed that the two markers RM128 and RM486 located on chromosome 1 were linked to the resistance gene in the respective populations above. This result is not consistent with those previously reported, in which a well-known resistance gene Pif in the St. No. 1 is located on chromosome 11. To confirm this result, additional four SSR markers, which located in the region lanked by RM128 and RM486, were tested. The results showed that markers RM543 and RM319 were closer to, and RM302 and RM212 completely co-segregated with the resistance locus detected in the present study. These results indicated that another resistance gene involved in the St. No. 1, which is located on chromosome 1, and therefore tentatively designated as Pi37(t). To narrow down genomic region of the Pi37(t) locus, eight markers were newly developed in the target region through bioinformatics analysis (BIA) using the publicly available sequences. The linkage analysis with these markers showed that the Pi37(t) locus was mapped to a ≈ 0.8 centimorgans (cM) interval flanked by RM543 and FPSM1, where a total of seven markers co-segregated with it. To physically map the locus, the Pi37(t)-linked markers were landed on the reference sequence of cv. Nipponbare through BIA. A contig map corresponding to the locus was constructed based on the reference sequence aligned by the Pi37(t)-linked markers. Consequently, the Pi37(t) locus was defined to 374 kb interval flanking markers RM543 and FPSM1, where only four candidate genes with the resistance gene conserved structure (NBS-LRR) were further identified to a DNA fragment of 60 kb in length by BIA.