Abstract
Bradyrhizobium japonicum is a nitrogen-fixing symbiont of soybean. In previous studies, transcriptomic profiling of B. japonicum USDA110, grown under various environmental conditions, revealed the highly induced gene aceA, encoding isocitrate lyase (ICL). The ICL catalyzes the conversion of isocitrate to succinate and glyoxylate in the glyoxylate bypass of the TCA cycle. Here, we evaluated the functional role of B. japonicum ICL under desiccation-induced stress conditions. We purified AceA (molecular mass = 65 kDa) from B. japonicum USDA110, using a His-tag and Ni-NTA column approach, and confirmed its ICL enzyme activity. The aceA mutant showed higher sensitivity to desiccation stress (27% relative humidity (RH)), compared to the wild type. ICL activity of the wild type strain increased approximately 2.5-fold upon exposure to 27% RH for 24 h. The aceA mutant also showed an increased susceptibility to salt stress. Gene expression analysis of aceA using qRT-PCR revealed a 148-fold induction by desiccation, while other genes involved in the glyoxylate pathway were not differentially expressed in this condition. Transcriptome analyses revealed that stress-related genes, such as chaperones, were upregulated in the wild-type under desiccating conditions, even though fold induction was not dramatic (ca. 1.5–2.5-fold).
Highlights
Bradyrhizobium japonicum is a soil bacterium that has the ability to form nitrogen-fixing root nodules on soybean (Glycine max L.)
AceA is of particular interest because it was highly induced under both stress conditions and the encoded protein isocitrate lyase (ICL) is a key enzyme in the glyoxylate pathway, a bypass of the tricarboxylic acid (TCA) cycle
Given the facts that the B. japonicum aceA is induced by desiccation and oxidative stresses, and that the genes encoding ICL in other microorganisms are involved in stress tolerance and virulence, it is plausible that aceA may play a key role in the survival of B. japonicum under various environmental stresses, and perhaps protection of the bacterium from plant defense mechanisms, such as an oxidative burst, during infection
Summary
Bradyrhizobium japonicum is a soil bacterium that has the ability to form nitrogen-fixing root nodules on soybean (Glycine max L.). AceA (blr2455) is of particular interest because it was highly induced under both stress conditions and the encoded protein isocitrate lyase (ICL) is a key enzyme in the glyoxylate pathway, a bypass of the tricarboxylic acid (TCA) cycle. Given the facts that the B. japonicum aceA is induced by desiccation and oxidative stresses, and that the genes encoding ICL in other microorganisms are involved in stress tolerance and virulence, it is. Plausible that aceA may play a key role in the survival of B. japonicum under various environmental stresses, and perhaps protection of the bacterium from plant defense mechanisms, such as an oxidative burst, during infection. We performed genetic, transcriptomic, physiological, and phenotyping experiments to determine how aceA responds to various stress conditions and how mutations of this gene affect survival and nodulation capacity of B. japonicum. Through transcriptomic comparisons between the wild type and the aceA mutant, we determined whether ICL altered expression of other genes in this bacterium
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