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Bradyrhizobium japonicum rhizobitoxine genes and putative enzyme functions: expression requires a translational frameshift.

ABSTRACT: Some strains of Bradyrhizobium japonicum produce rhizobitoxine, a phytotoxin that causes foliar chlorosis on susceptible host plants. We have previously obtained Tn5-induced rhizobitoxine null mutants of B. japonicum. DNA sequence analysis of the region surrounding two Tn5 insertions identifies two overlapping open reading frames. The first open reading frame (rtxA) predicts a 54-kDa protein for which the N-terminal 280 residues have sequence similarity to serine: pyruvate aminotransferase. The sequence homology to aminotransferase is consistent with the involvement of this gene in serinol production, a likely intermediate in rhizobitoxine biosynthesis. Previously, a mutant in this open reading frame was shown not to make serinol. The predicted amino acid sequence of the second open reading frame (rtxB) has similarity to yeast O-acetylhomoserine sulfhydrolase. This enzyme function is similar to that required for dihydrorhizobitoxine synthase. The DNA sequence shows that the rtxB open reading frame overlaps rtxA, suggesting that expression of rtxB requires a -1 translational frameshift. Protein expression experiments demonstrate production of an RtxAB fusion protein. The ability of the overlapping rtxA and rtxB sequences to promote a translational frameshift was confirmed in a heterologous expression system. In Escherichia coli, this frameshift appears to be unusually efficient, occurring at a frequency of 80-90%.

SUBMITTER: Ruan X

PROVIDER: S-EPMC46151 | biostudies-other | 1993 Apr

REPOSITORIES: biostudies-other

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Bradyrhizobium japonicum rhizobitoxine genes and putative enzyme functions: expression requires a translational frameshift.

Ruan X X Zhang C C Peters N K NK

Proceedings of the National Academy of Sciences of the United States of America 19930401 7

Some strains of Bradyrhizobium japonicum produce rhizobitoxine, a phytotoxin that causes foliar chlorosis on susceptible host plants. We have previously obtained Tn5-induced rhizobitoxine null mutants of B. japonicum. DNA sequence analysis of the region surrounding two Tn5 insertions identifies two overlapping open reading frames. The first open reading frame (rtxA) predicts a 54-kDa protein for which the N-terminal 280 residues have sequence similarity to serine: pyruvate aminotransferase. The ...[more]

PMID: 8464870

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Project description:Copper-limiting growth conditions were thought to cause an induction of genes possibly involved in copper uptake and sorting. This rationale in mind, we performed microarray analyses on B. japonicum cells grown in three variations of the BVM minimal medium. Variant 1 contained 2 M-NM-<M CuSO4 (copper excess). Variant 2 was prepared in HCl-treated glassware without any copper added (copper starvation). The residual copper concentration in this copper-starvation medium was analyzed by GF-AAS and determined to be 5 nM. Variant 3 (extreme copper limitation) was prepared like variant 2 but with the addition of 10 M-NM-<M BCS and 1 mM ascorbic acid where BCS chelates Cu(I) selectively, and ascorbic acid reduces any Cu(II) to Cu(I). Changes in the transcription profiles were recorded by the pairwise comparison of cells grown in variant 2 vs. 1, and variant 3 vs. 2. Only a small set of genes were differentially up- or down-regulated when copper-starved cells were compared with cells grown in copper excess. Most notably, five genes located adjacent to each other on the B. japonicum genome displayed an increased expression: bll4882 to bll4878. The five genes were named pcuA, pcuB, pcuC, pcuD, and pcuE (mnemonic of proteins for Cu trafficking). The genes with decreased expression are either of unknown function or M-bM-^@M-^S not surprisingly M-bM-^@M-^S play a role in copper resistance. Extreme copper limitation (variant 3 vs. 2) did not further enhance the expression of the five pcu genes. Instead, another cluster of adjacent genes was strongly up-regulated: bll0889 to bll0883, which code for unidentified transport functions. Incidentally, the list also includes the copper chaperone ScoI. Taken together, copper-limiting growth conditions have led to the de-repression of genes potentially involved in copper acquisition. Microarray-based transcriptome analysis of B. japonicum 110spc4 wild-type cells grown under normal, copper-limiting and copper excess conditions

Dataset Information

Bradyrhizobium japonicum rhizobitoxine genes and putative enzyme functions: expression requires a translational frameshift.

Publications

Bradyrhizobium japonicum rhizobitoxine genes and putative enzyme functions: expression requires a translational frameshift.

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