Dehydrin genes and their expression in recalcitrant oak (<Emphasis Type="Italic">Quercus robur</Emphasis>) embryos

In this work, three dehydrin genes, QrDhn1, QrDhn2, QrDhn3, were isolated from recalcitrant oak (Quercus robur). Their expression pattern was analyzed in both zygotic and somatic embryos as well as in vegetative tissues exposed to different kinds of abiotic stresses including desiccation, osmotic stress, and chilling. The QrDhn1 gene encoding for Y_nSK_n type dehydrin was expressed during later stages of zygotic embryo development but in somatic embryos only when exposed to osmotic or desiccation stress. In contrast, the other two oak dehydrin genes encoding for putative K_n type dehydrins were expressed only in somatic embryos (both not-treated and osmotically stressed) and leaves of oak seedlings exposed to desiccation. Behavior of these genes suggests that different dehydrins are involved in processes of seed maturation and response to altered osmotic (water status) conditions in somatic embryos. Revealing further members of dehydrin gene family in recalcitrant oak might contribute to clarify non-orthodox seed behavior as well as identify mechanisms contributing to desiccation tolerance in plants.     

Isolation of a dehydrin cDNA from orange and grapefruit citrus fruit that is specifically induced by the combination of heat followed by chilling temperatures

Dehydrins (DHNs; late embryogenesis abundant D-11) are a family of plant proteins induced in response to environmental stresses such as water stress, salinity and freezing or which occur during the late stages of embryogenesis. Previously, it was reported that citrus contains a small gene family encoding a unique class of dehydrins that differs from most other plant dehydrins in various respects, such as having an unusual K-segment similar to that of gymnosperms. In the present study, we identified by cDNA differential display analysis a 'Navel' orange 202-bp polymerase chain reaction (PCR) fragment, which encoded the typical plant angiosperm-type K-segment consensus sequence, and of which the expression was down-regulated by exposure to low oxygen levels. The full-length cDNA sequence of the orange DHN, designated csDHN (for Citrus sinensis DHN), was further isolated by 5'-and 3'-RACE; it had a total length of 933 bp and encoded a predicted polypeptide of 235 amino acids. In addition, the same 202-bp 'Navel' dehydrin PCR fragment was used to screen a 'Star Ruby' grapefruit flavedo cDNA library, and its full-length grapefruit homologue, designated cpDHN (for C. paradisi DHN) was isolated and found to have a total length of 1024 bp and to encode a predicted polypeptide of 234 amino acids. The defined orange and grapefruit DHN proteins were completely identical in the 196 amino acids of their N-terminus but differed in their C-terminus region. Overall, the csDHN and cpDHN proteins share 84% identity and contain the conserved dehydrin serine cluster (S-segment) and a putative nuclear localization signal, but csDHN has one conserved dehydrin K-segment consensus sequence, whereas cpDHN contains two dehydrin K-segments. Both csDHN and cpDHN represent single copy genes, in 'Navel' orange and 'Star Ruby' grapefruit genomes, respectively. We found that the cpDHN gene was consistently expressed in the fruit peel tissue at harvest, but that its message levels dramatically decreased during storage at either ambient or low temperatures. However, a pre-storage hot water treatment, given to enhance fruit-chilling tolerance, increased cpDHN mRNA levels during the first 3 weeks of cold storage at 2 degrees C, and enabled the message levels to be retained for up to a further 8 weeks of cold storage at 2 degrees C. The hot water treatment by itself had no inductive effect on cpDHN gene expression when the fruits were held at non-chilling temperatures. Other stresses applied to the fruit, such as wounding, UV irradiation, water stress, low oxygen and exposure to the stress hormone ethylene decreased DHN mRNA levels, whereas abscisic acid had no effect at all.     

Postharvest heat and conditioning treatments activate different molecular responses and reduce chilling injuries in grapefruit

A combination of hot water (a rinse at 62 degrees C for 20 s) and conditioning (pre-storage at 16 degrees C for 7 d) treatments synergistically reduced chilling injury development in grapefruit (Citrus paradisi, cv. "Star Ruby") during cold storage at 2 degrees C, suggesting that the treatments may activate different chilling tolerance responses. To study the molecular mechanisms involved, chilling- and conditioning-responsive genes were isolated by polymerase chain reaction (PCR) cDNA subtraction, cDNA libraries were constructed from hot water- and conditioning-treated fruit, and cDNA sequencing was used to identify putative stress-responsive and chilling tolerance genes. PCR cDNA subtraction revealed the identification of 17 chilling-responsive and heat- and conditioning-induced genes, and the expression patterns of 11 additional stress-related genes, antioxidant defensive genes, and genes encoding enzymes involved in membrane lipid modifications were characterized. It was found that hot water and conditioning treatments had little effect on gene expression by themselves, but rather had a priming effect, and enabled the fruit to activate their defence responses after subsequent exposure to chilling. RNA gel blot hybridizations revealed that the expression patterns of eight genes, including HSP19-I, HSP19-II, dehydrin, universal stress protein (USP), EIN2, 1,3;4-beta-D-glucanase, and superoxide dismutase (SOD), were specifically regulated by the heat treatment, and four genes, including fatty acid desaturase2 (FAD2) and lipid transfer protein (LTP), were specifically regulated by the conditioning treatment. Furthermore, four more genes were identified, including a translation initiation factor (SUI1), a chaperonin, and alcohol dehydrogenase (ADH), that were commonly regulated by both heat and conditioning treatments. According to these data, it is suggested that pre-storage heat and conditioning treatments may enhance fruit chilling tolerance by activating different molecular mechanisms. The hot water treatment activates mainly the expression of various stress-related genes, whereas the conditioning treatment activates mainly the expression of lipid membrane modification enzymes.     

Aluminium-induced drought and oxidative stress in barley roots

The aim of the present study was to examine the relation between Al accumulation in root tissues, root growth inhibition, root water content, cell viability and expression of oxidative and drought stress-related genes in barley roots growing on the filter paper. Al-induced root growth inhibition correlated with Al uptake and cell death. Water content of Al-treated root represented only half of the control one. The expression of the dehydrin gene dhn4, which is a marker for drought stress in plant tissues, was strongly induced during Al stress. Al treatment also induced expression of oxidative stress-related genes such as glutathione peroxidase (gpx), pathogen-related peroxidase (prx8), glutathione reductase (gr) and dehydroascorbate reductase (dhar). The present results suggest correlation between Al uptake, Al-induced drought stress, oxidative stress, cell death and root growth inhibition.     

Molecular cloning of the anther-preferential nonspecific lipid transfer protein cDNA in hot pepper by mRNA differential display

To better understand the molecular control of anther development, an anther-preferential mRNA was isolated from hot pepper (Capsicum annuum) using mRNA differentially display. Using the displayed fragment as a probe, a full-length cDNA named CaLTP was isolated. A nucleotide sequence analysis of CaLTP revealed that the clone contains an open reading frame of 123 amino acids, which exhibits a 60-23% identity with nonspecific lipid transfer proteins (nsLTP). Northern and RT-PCR analysis of the clone confirmed that CaLTP mRNA was predominant to anther tissues. The basal expression level in the leaves was slightly induced only by abscisic acid (ABA) treatment. Southern analysis reveals that CaLTP is present as a single-copy gene in hot pepper genome. We hypothesize that CaLTP might have an important role in protecting the reproductive tissues from environmental stresses.     

菜豆重金属胁迫响应基因:cDNA克隆及其表达分析

通过差别筛选HgCl2胁迫下的菜豆叶片cDNA库,分离出7组不同的cDNA克隆（Phaseolusvulgarisstress-relatedprotein,PvSR1～7）。cDNA序列和同源性分析结果表明：PvSR1编码富含脯氨酸细胞壁蛋白（PRP）,PvSR2和PvSR7编码新的HgCl2胁迫相关蛋白,PvSR3编码脱水蛋白（dehydrin）,PvSR4编码病原相关（PR）蛋白,PvSB5编码polyubiqui－tin,PvSR6编码DuaJ－like蛋白。HgCl2胁迫可强烈地请导PvSR2和PR蛋白基因的表达,并能提高PRP,、dehydrinlike和polyubiq－uitin基因的转录水平。这些蛋白质共同作用可能对维持细胞的正常代谢和抵抗重金属胁迫方面有重要作用。     

辣椒CaNAC36转录因子耐盐性分析

【目的】辣椒是中国种植面积最大的蔬菜作物,随着土地盐碱化问题的日趋严重,加强辣椒耐盐机制研究对促进产业可持续发展具有重要意义。因而,急需加快辣椒耐盐相关关键基因的功能研究。【方法】研究组前期挖掘到与辣椒耐盐性相关的转录因子CaNAC36,在此基础上,以耐盐辣椒PI201224和敏盐辣椒PI438643为供试品种,克隆获得CaNAC36全长gDNA和cDNA序列,通过荧光定量分析CaNAC36及可能的互作基因在盐胁迫条件下不同组织部位的表达情况,并进一步结合生物信息学分析探究CaNAC36及其互作基因之间存在的潜在关系。【结果】结果表明,CaNAC36序列在耐盐和敏盐材料中DNA和cDNA同源性分别为99.86%和100%;荧光定量的结果表明,CaNAC36在耐盐材料根和茎组织中表现为诱导上调表达,在敏盐材料根和叶中表现为诱导下调表达;对可能与CaNAC36存在互作关系的48个基因的注释信息进行分析后,发现跨膜蛋白、转运蛋白、水孔蛋白、氯离子通道蛋白、解毒蛋白等14个基因可能与CaNAC36存在功能互作。进一步分析发现,在PI201224和PI438643盐胁迫处理不同时间点、不同组织中,5个相关基因（Capana08g002748、Capana00g004514、Capana09g000275、Capana07g001450、Capana02g001031）的表达呈现显著差异。同时发现,CaNAC36及5个关联基因启动子域含有大量的逆境相关顺式作用元件。【结论】结合基因克隆、基因表达水平分析以及生物信息学分析,表明CaNAC36是辣椒响应盐胁迫的重要转录因子,并可能与其他基因相互作用以提高植株的耐盐性,可为深度研究辣椒耐盐性以及选育耐盐品种提供数据支撑。     

The CaPRP1 gene encoding a putative proline-rich glycoprotein is highly expressed in rapidly elongating early roots and leaves in hot pepper (Capsicum annuum L. cv. Pukang)

Most of the proline-rich cell wall glycoprotein genes isolated from higher plants are preferentially expressed in the transmitting tissues of the flower organ. In conducting expressed sequence tag (EST) analysis, which was prepared from 5-day-old early roots of hot pepper (Capsicum annuum L. cv. Pukang), we identified a cDNA clone, pCaPRP1, encoding a putative cell wall proline-rich glycoprotein. CaPRP1 (Mr=28 kDa, pI=9.98) was most closely related to Nicotiana alata NaPRP4 (71%), while most distantly related to soybean PvPRP (37%). The predicted primary structure of CaPRP1 contains a putative N-terminal signal peptide, six repeats of the Lys-Pro-Pro tripeptide, four repeats of a five-amino acid sequence [Pro-(Ser/The)-Pro-Pro-Pro] and one potential N-glycosylation site (Asn-Asn-Ser). In contrast to most proline-rich cell wall glycoprotein genes, CaPRP1 was highly expressed in rapidly elongating very early roots and young leaves as well as developing flower tissues. Although the physiological function of CaPRP1 is not yet clear, there are several possibilities for its role in cell expansion and elongation during early development of hot pepper plants.