Regulation of catalase gene expression in soybean

Catalase is one of the important enzymes in the cellular active-oxygen scavenging system of aerobic organisms. In plants, catalase is located in peroxisomes where it converts hydrogen peroxide generated during 6-oxidation of fatty acids and photorespiration to oxygen and water. This study attempts to serve as an initial step in understanding the regulation of catalase gene expression in plants.

Multiple isoforms of catalase are present in several plant species such as maize, cotton and tobacco. Expression of these isoforms is under developmental and environmental control. In pea and soybean, a single catalase isoform has been detected in all tissues and developmental stages tested. Genomic DNA blot analysis suggested that there is a single catalase gene in pea genome, while soybean appears to have two catalase loci. A pea cDNA that includes the entire coding sequence for a catalase subunit was isolated and characterized. The deduced amino acid sequence proved to be highly similar to several other plant catalase sequences. Using the pea cDNA as probe, a soybean genomic clone that includes catalase coding sequences and 1.8 kb of upstream sequence was isolated. Sequence analysis indicated that the soybean catalase gene contains six introns, and the deduced amino acid sequence has a high percentage of identity with the pea catalase subunit. In soybean seedlings, very low levels of catalase activity were detected in germinated seeds one day after imbibition (DAI), but catalase activity increased rapidly, reaching a maximum by 4 DAI. Catalase mRNA was detectable by 1 DAI and increased gradually to reach maximum levels within 4 DAI. In dark grown seedlings, catalase mRNA was maintained at relatively high levels through 6 DAI . Exposure of seedlings to light after 3 DAI caused a more rapid decline in catalase mRNA levels when compared to dark grown seedlings. This pattern of differential accumulation in light/dark conditions was not observed in the catalase activity levels which were determined during the same developmental period. The upstream region of the soybean catalase gene was fused to the GUS (P-glucuronidase) reporter gene, and a series of 5' deletions were generated using Exonuclease III. Six of these deletion constructs were used to transform tobacco plants. Fluorometric GUS analysis indicated that GUS expression levels were much higher in imbibed seeds than in mature leaves, suggesting that this catalase promoter is seed-specific. Histochemical GUS staining of transgenic plant tissues showed that GUS is expressed at a high level in the cotyledons at early stages of seedling development, but the expression is maintained primarily in the vascular tissues, specifically in the phloem of the mature plants.