A two-dimensional proteome reference map of Herbaspirillum seropedicae proteins

Herbaspirillum seropedicae is an endophytic diazotroph associated with economically important crops such as rice, sugarcane, and wheat. Here, we present a 2-D reference map for H. seropedicae. Using MALDI-TOF-MS we identified 205 spots representing 173 different proteins with a calculated average of 1.18 proteins/gene. Seventeen hypothetical or conserved hypothetical ORFs were shown to code for true gene products. These data will support the genome annotation process and provide a basis on which to undertake comparative proteomic studies.     

Sequencing and functional analysis of the nifENXorf1orf2 gene cluster of Herbaspirillum seropedicae

A 5.1-kb DNA fragment from the nifHDK region of H. seropedicae was isolated and sequenced. Sequence analysis showed the presence of nifENXorf1orf2 but nifTY were not present. No nif or consensus promoter was identified. Furthermore, orf1 expression occurred only under nitrogen-fixing conditions and no promoter activity was detected between nifK and nifE, suggesting that these genes are expressed from the upstream nifH promoter and are parts of a unique nif operon. Mutagenesis studies indicate that nifN was essential for nitrogenase activity whereas nifXorf1orf2 were not. High homology between the C-terminal region of the NifX and NifB proteins from H. seropedicae was observed. Since the NifX and NifY proteins are important for FeMo cofactor (FeMoco) synthesis, we propose that alternative proteins with similar activities exist in H. seropedicae.     

Iron depletion affects nitrogenase activity and expression of nifH and nifA genes in Herbaspirillum seropedicae

Herbaspirillum seropedicae Z67 is a nitrogen-fixing bacterium able to colonize the rhizosphere and the interior of several plants. As iron is a key element for nitrogen fixation, we examined the response of this microorganism to iron deficiency under nitrogen fixing conditions. We identified a H. seropedicae exbD gene that was induced in response to iron limitation and is involved in iron homeostasis. We found that an exbD mutant grown in iron-chelated medium is unable to fix nitrogen. Moreover, we provide evidence that expression of the nifH and nifA genes is iron dependent in a H. seropedicae genetic background.     

Production of indole-3-acetic acid and gibberellins A1 and A3 by Acetobacter diazotrophicus and Herbaspirillum seropedicae in chemically-defined culture media

The characterization by capillary gas chromatography-mass spectrometry of the plant hormones indole-3-acetic acid and the gibberellins GA₁ and GA₃ from chemically-defined cultures of Acetobacter diazotrophicus and Herbaspirillum seropedicae is reported. Both bacteria are endophytic in gramineae species where they promote growth and yield. Quantification was also done by selected ion monitoring with [17,17-²H₂]-Gibberellin A₁, [17,17-²H₂]-Gibberellin A₃ and [¹³C₆]-indole-3-acetic acid as internal standards. The results presented show the importance of studying phytohormonal production when the interrelationships between plants and microorganisms are analyzed and may help explain the beneficial effects of endophytic bacteria to the host plant, as has been demonstrated previously for Azospirillum spp.     

盐胁迫下粪产碱菌在水稻根表的定殖和异源固氮基因的表达

异源nif LacZ融合基因在粪产碱菌A15 6 1中的表达活性随盐浓度增加而升高 ,然后逐渐降低 ,nifH LacZ融合基因可以正常表达的盐浓度在 0 .1%～0 .5 %之间 ,盐浓度为 0 .0 5 %时活性最高。A15 6 1在盐浓度为 0 .0 6 %时趋化能力最强 ,随着盐浓度的提高逐渐下降 ,当盐浓度为 3 .0 %时完全丧失趋化能力。一定的盐浓度 (0 .5 % )对固氮粪产碱菌的根表定殖有促进作用 ,该条件下根表定殖的菌体数远大于对照。 3种nif LacZ融合基因在根内的表达部位有显著差异。nifH的表达部位主要分布于根的皮层薄壁组织细胞间隙 ,在条件适宜 (无铵和微量氧 )的部位或某些特殊位置如侧根伸出部位高水平表达。盐胁迫下水稻 耐盐粪产碱菌A15 6 1的联合固氮效率明显高于A15 6 1纯培养物     

Use of viral replicons for the expression of genes in plants

Autonomously replicating virus-based vectors have been investigated as a means of introducing heterologous genes into plants. This approach has a number of potential advantages over stable genetic transformation, particularly in terms of speed and levels of expression that can be obtained. Several groups of plant viruses, with genomes consisting of both DNA and RNA, have been investigated as possible gene vectors. In the case of DNA viruses, it has generally been possible to identify nonessential regions of the genome that can be replaced by foreign sequences. However, there appear to be limitations on the size of insert which can be tolerated. In the case of RNA viruses, replacement of viral sequences usually has a drastic effect on the viability. However, in several cases it has proved possible to substantially increase the size of the viral genome by the direct insertion of additional sequences while still retaining the ability of the viruses to multiply and spread in plants. These RNA virus-based systems appear to have the greatest potential as gene vectors.