高等植物病原相关蛋白

在过去的三十年中,人们对诱导系统性抗性——这一普遍存在于高等植物抗病过程中的现象——进行了深入研究。被真菌、细菌或病毒侵染后,植物表现出广泛的、长时间的系统性抗性。在这一过程中,植物细胞壁组成成分发生改变,表达各种病原相关蛋白(PR蛋白),并合成多种植物抗毒素。本文就主要的PR蛋白家族的结构和功能特性,PR蛋白的发现和分类,及PR蛋白的应用作一综述。     

植物病原相关蛋白研究进展

植物在受到真菌、细菌、病毒侵染或意外的伤害时体内会产生新的蛋白质,称作病原相关蛋白(pathogenesisrelatedprotein简称PR蛋白)。PR蛋白的产生与植物对病原物的抵御直接相关,本文就PR蛋白的诱导表达、调控和作用机理方面的最新研究进展作一综述。     

The chloroplast genome of Nicotiana sylvestris and Nicotiana tomentosiformis: complete sequencing confirms that the Nicotiana sylvestris progenitor is the maternal genome donor of Nicotiana tabacum

The tobacco cultivar Nicotiana tabacum is a natural amphidiploid that is thought to be derived from ancestors of Nicotiana sylvestris and Nicotiana tomentosiformis. To compare these chloroplast genomes, DNA was prepared from isolated chloroplasts from green leaves of N. sylvestris and N. tomentosiformis, and subjected to whole-genome shotgun sequencing. The N. sylvestris chloroplast genome comprises of 155,941 bp and shows identical gene organization with that of N. tabacum, except one ORF. Detailed comparison revealed only seven different sites between N. tabacum and N. sylvestris; three in introns, two in spacer regions and two in coding regions. The chloroplast DNA of N. tomentosiformis is 155,745 bp long and possesses also identical gene organization with that of N. tabacum, except four ORFs and one pseudogene. However, 1,194 sites differ between these two species. Compared with N. tabacum, the nucleotide substitution in the inverted repeat was much lower than that in the single-copy region. The present work confirms that the chloroplast genome from N. tabacum was derived from an ancestor of N. sylvestris, and suggests that the rate of nucleotide substitution of the chloroplast genomes from N. tabacum and N. sylvestris is very low. Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorized users.     

植物病程相关蛋白及其在烟草中的研究进展

植物在受到病原物侵染时,会产生一系列抗性反应,病程相关蛋白是其中参与抗病性的重要物质,能够被病原物诱导产生并在植物体内积累,对于诱导植物系统抗性,阻止病原物侵染具有重要作用。对植物病程相关蛋白的性质、诱导因素、分类和功能进行综述,并概述病程相关蛋白与烟草系统抗性的紧密联系以及烟草病程相关蛋白基因在增强系统抗性中的应用,为烟草抗病育种和病虫害防治提供了理论。     

The Extracellular Acidic and Basic Pathogenesis-Related Proteins of Soybean Induced by Viral Infection

Seventeen major host-encoded pathogenesis related (PR)-proteins have been found in intercellular fluids of necrotic virus-infected soybean leaves. None of them was present in fluids of healthy controls. By native and SDS-denaturing polyacrylamide gel electrophoresis, ten major acidic PR-proteins have been identified and classified on the basis of their molecular weight in three groups: group 1 included four proteins of 16–17 Kd; group 2, three proteins of 26 Kd; group 3, three proteins of 32 Kd. Seven PR-proteins were basic, and were classified in three groups: group 1 included three proteins of 16 Kd; group 2, one protein of 23 Kd; group 3, three proteins of 32 Kd. As found for tobacco and potato, soybean PR-protein patterns show high number of acidic and basic proteins.     

Plasmatubules in the pollen tubes of Nicotiana sylvestris

Ultrastructural studies of the pollen tubes of Nicotiana sylvestris grown in the pistil revealed an extensive development of plasmatubules formed by evaginations of the plasma membrane. The plasmatubules occurred as twisted tubular structures in the periplasmic space along the tube wall and, in cross section, exhibited circular profiles with an outer diameter of 28±4 nm. They were also seen in deep, pocket-like invaginations of the plasma membrane and in this case the profiles had an outer diameter of 34±8 nm. In the pocket-like invaginations they were partially branched and often closely packed to form groups with obvious patterns. The enlargement of the plasma-membrane area resulting from plasmatubules formed along the tube wall was about six-to tenfold. Pollen tubes grown in vitro exhibited poorly developed plasmatubules. It is suggested that the large extension of the plasma membrane could enhance the uptake of nutrients, and thus might be responsible for the comparatively fast growth of pollen tubes in the pistil. Moreover, it is also assumed that the turnover rate of the Golgi apparatus must be higher in pollen tubes growing in vivo than in vitro, in order to provide a sufficient amount of membrane for the formation of the plasma membrane with its tubular modifications.     

水稻病程相关蛋白质在逆境胁迫下的表达研究

植物病程相关(PR)基因一般在病原物侵染过程中受诱导发生转录上调．目前有证据提示植物PR基因在非生物逆境胁迫下也发生转录变化,但其蛋白质的表达变化情况还鲜有报道．为了解水稻PR蛋白质在逆境胁迫下的表达特征,本文采用免疫印迹技术(Western blotting,WB)调查了8个PR蛋白质在冷、热、旱、淹和盐等5种胁迫下的表达谱．结果表明：在冷胁迫下PR8表达上调,在热胁迫下PR1a、PR3、PR5和PR16表达下调;在旱胁迫下PR1a、PR2和PR8表达上调,而PR5 和PR16表达下调,在淹胁迫下PR1、PR2和PR15表达上调,PR1a、PR3、PR5和PR8表达下调;在盐胁迫下PR2和PR3表达上调,而PR1a、PR5、PR8和PR16表达下调．另外,对这些PR 基因的上游启动子区进行分析,发现存在与胁迫响应相关的调控元件,其中脱落酸反应元件(ABRE)、TC-rich repeats和HSE的出现频率较高．这些蛋白质表达数据进一步佐证了PR蛋白在逆境胁迫反应中发挥着重要且不尽相同的作用．     

Going Viral with Fluorescent Proteins

Many longstanding questions about dynamics of virus-cell interactions can be answered by combining fluorescence imaging techniques with fluorescent protein (FP) tagging strategies. Successfully creating a FP fusion with a cellular or viral protein of interest first requires selecting the appropriate FP. However, while viral architecture and cellular localization often dictate the suitability of a FP, a FP''s chemical and physical properties must also be considered. Here, we discuss the challenges of and offer suggestions for identifying the optimal FPs for studying the cell biology of viruses.     

Genome-Wide Association Study Identifies Loci Associated with Resistance to Viral Nervous Necrosis Disease in Asian Seabass

Viral nervous necrosis disease (VNN), caused by nervous necrosis virus (NNV), is one major threat to mariculture. Identifying loci and understanding the mechanisms associated with resistance to VNN are important in selective breeding programs. We performed a genome-wide association study (GWAS) using genotyping-by-sequencing (GBS) to study the genomic architecture of resistance to NNV infection in Asian seabass. We genotyped 986 individuals from 43 families produced by 15 founders with 44498 bi-allelic genetic variants using GBS. The GWAS identified three genome-wide significant loci on chromosomes 16, 19, and 20, respectively, and six suggestive loci on chromosomes 1, 8, 14, 15, 21, and 24, respectively, associated with resistance to NNV infection measured as binary and quantitative traits. Using the 500 most significant markers in combination with a training population of 800 samples could reach a genomic prediction accuracy of 0.7. Candidate genes significantly associated with resistance to NNV, including lysine-specific demethylase 2A, beta-defensin 1, and cystatin-B, which play important roles in immune responses against virus infection, were identified. Almost all the candidate genes were differentially expressed in different tissues against NNV infection. The significant genetic variants can be used in genomic selection and help understand the mechanism of resistance to VNN. Future studies should use populations of large effective size and whole genome resequencing to identify more useful genetic variants.     

The Nucleocytoplasmic Transport of Viral Proteins

Molecules can enter the nucleus by passive diffusion or active transport mechanisms, depending on their size. Small molecules up to size of 50-60 kDa or less than 10 nm in diameter can diffuse passively through the nuclear pore complex (NPC), while most proteins are transported by energy driven transport mechanisms. Active transport of viral proteins is mediated by nuclear localization signals (NLS), which were first identified in Simian Virus 40 large T antigen and had subsequently been identified in a lar...     

Amplification of repetitive DNA from Nicotiana plumbaginifolia in asymmetric somatic hybrids between Nicotiana sylvestris and Nicotiana plumbaginifolia

Summary Asymmetric somatic hybrids were obtained between a chlorophyll-deficient mutant of Nicotiana sylvestris (V42) and a nitrate-reductase (NR)-deficient line of N. plumbaginifolia (cnx20 or Nia26), using each of the parents alternately as the irradiated donor. Irradiation doses applied ranged from 10 to 1,000 Gy of gamma-rays. Hybrid selection was based on complementation of NR deficiency with wild-type NR genes. To aid in the analysis of somatic hybrids, species-specific repetitive DNA sequences from N. plumbaginifolia (NPR9 and NPR18) were cloned. NPR18 is a dispersed repetitive sequence occupying about 0.4% of the N. plumbaginifolia genome. In turn, NPR9, which is part of a highly repetitive DNA sequence, occupies approximately 3% of the genome. The species-specific plant DNA repeats, together with cytological analysis data, were used to assess the relative amount of the N. plumbaginifolia genome in the somatic hybrids. In fusion experiments using irradiated N. plumbaginifolia, an increase in irradiation dose prior to fusion led to a decrease in N. plumbaginifolia nuclear DNA content per hybrid genome. For some hybrid lines, an increase in the quantity of repetitive sequences was detected. Thus, hybrid lines 1NV/21, 100NV/7, 100NV/ 9, and 100NV/10 (where N. plumbaginifolia was the irradiated donor) were characterized by amplification of NPR9. In the reverse combination (where N. sylvestris was the irradiated donor), an increase in the copy number of NPR18 was determined for hybrid clones 1VC/2, 1VC/3, 100VC/2 and oct100/7. Possible reasons for the amplification of the repeated sequences are discussed.     

Induction of embryos in pollen cultures of Nicotiana sylvestris

Induction of embryos in pollen cultures was possible when young buds from plants induced to flower at 15°C were given a cold treatment at 10°C for 15 days. Following selective isolation of embryogenic pollen the cultures raised were given cold treatment at 10°C for 12 or more days. In such cultures the embryos appeared at a frequency of 1.2% of the cultured pollen on mineral-sucrose medium at pH 6.8.     

Characteristics of Pathogenesis-Related Proteins Induced in Phaseolus vulgaris cv. Pinto Following Viral Infection

Ten acidic and eight basic pathogenesis-related (PR) proteins were induced in primary leaves of Phaseolus vulgaris cv. Pinto following infection with southern bean mosaic virus (SBMV), a virus that induces circular, brown necrotic local lesions. These PR proteins included: four, 17 kDa, serologically related, acidic proteins of unknown functions; two chitinases, one acidic (29 kDa) and one basic (32 kDa) possessing antifungal activities; and four (21 kDa, 28 kDa, 29 kDa, 36 kDa) serologically related, acidic glucanases. The production of PR proteins was minimal during the period of active SBMV multiplication; greatest increase of PR proteins occurred after SBMV had reached a maximum level. Several of these PR proteins, including the 29 kDa acidic chitinase and two acidic glucanases (21 kDa and 36 kDa) accumulated in the intercellular space fluid. Similar PR proteins were synthesized in Pinto bean primary leaves infects with any of three other viruses that formed lesions of divergent phenotypes. However, the proportion of some of the PR proteins in the various lesion-types differed considerably. These results indicate that PR proteins play no active role in viral localization or inactivation and that their production is a metabolic adjustment to endogenous stress.     

Microheterogeneity of PSI-E Subunit of Photosystem I in Nicotiana sylvestris

Microheterogeneity of a photosystem I (PSI) subunit encodedby a nuclear gene psaE was examined in Nicotiana sylvestris,with the aid of cDNA cloning, peptide mapping analysis and proteinsequencing. The psaE product of this plant has four isoformswhose mobilities in PAGE are slightly different from each other.We isolated two types of psaE cDNAs from a N. sylvestris cDNAlibrary, and designated the corresponding genes as psaEa andpsaEb, respectively. The psaEa and psaEb genes are 77% homologousat DNA level, and their translation products share 80.4% homologyfor the precursor proteins and 89.1% for the mature forms. Comparativeanalysis of the four isoproteins and the putative products ofthe two psaE genes revealed that two isoproteins out of fourare derived from psaEa gene, and the difference between thesetwo isoproteins lies in the respective presence or absence ofN-terminal alanine. Likewise, the other two proteins are derivedfrom psaEb with similar N-terminal heterogeneity. These resultsindicate that multi-gene organization and heterogeneous N-terminalformation at post-translational level are two possible causesfor PSI subunit polymorphism in isogenic plant lines. (Received October 8, 1993; Accepted November 30, 1993)     

Metabolism of polyamines in NaCl-resistant cell lines from Nicotiana sylvestris

The polyamine titers in three cell lines of Nicotiana sylvestris were compared: Type 1, rapidly adapting to NaCl; Type 2, constantly resistant to NaCl; Type 3, a saltsensitive wild strain. During short-term cultivation in MS medium in the presence of 170 mM NaCl (1 passage, 14 d) the changes in polyamine titer in cell suspensions of type 1 (in a slightly adapted state) and non-adapted wild strain (type 3) showed a considerable increase in spermidine and spermine and a decrease in putrescine. After prolonged adaptation to NaCl (20 passages) the putrescine content in the cells of type 1 and of type 2 was increased at the expense of the polyamines. This suggests that the pattern of polyamine titer varies under short- and long-term adaptation to NaCl. The inverse ratio between growth processes and changes in polyamine and proline level indicates that polyamines fulfil primarily a protective and osmorepulatory function in plant cells under NaCl stress.