| The enzyme 1-aminocyclopropane-1-carboxylate deaminase (ACCD) converts ACC, the precursor of the plant hormone ethylene, to alpha-ketobutyrate and ammonia. A soluble recombinant ACCD from Pseudomonas stutzeri UW4 of molecular weight 41 kDa has been cloned, expressed, and purified.; PCR with degenerate primers (DegACC5' and DegACC3 ') was used to rapidly identify and isolate 1-aminocyclopropane-1-carboxylate (ACC) deaminase genes from eighteen different soil bacteria including both gram-positive and gram-negative species. The technique proved sensitive enough to distinguish between organisms containing the ACC deaminase gene and organisms without the gene. In every instance, when approximately 750 bp band was obtained after PCR it contained the ACC deaminase gene. Upon analysis of the ACC deaminase amino-acid sequences it was observed that the ACC deaminase proteins are largely conserved within a bacterial genus. In addition when comparing the bacterial 16s rDNA sequence phylogenetic tree with the ACC deaminase phylogenetic tree, it was observed that there is very little similarity. This provides some evidence for lateral gene transfer of the ACC deaminase gene and its regulatory Lrp protein.; The technique of RNA arbitrarily primed PCR (RAP-PCR) was used to study changes in gene expression, over time, in canola roots treated with the 1-aminocyclopropane-1-carboxylate deaminase-containing plant growth-promoting bacterium P. stutzeri UW4, and compared with a mutant of P. stutzeri UW4 in which the ACC deaminase structural gene acdS was replaced by homologous recombination with a tetracycline resistance gene. Genes up or down-regulated over a three day study, in the plant growth-promoting bacteria-treated canola plants were isolated, cloned and sequenced; all appeared to have high homology with Arabidopsis thaliana genes. These results indicate that roots isolated from canola seeds treated with the ACC deaminase-producing P. stutzeri UW4, up-regulate genes involved in cell division and proliferation but down-regulate stress genes. This data is in agreement with a model in which ACC deaminase-containing plant growth promoting bacteria reduce plant stress and induce root elongation and proliferation in plants, largely by lowering ethylene levels. (Abstract shortened by UMI.)... |
