We describe a gene, SCB1 (Seed Coat BURP-domain protein 1), that is expressed specifically within the soybean (Glycine max [L.] Merrill) seed coat early in its development. Northern blot analysis and mRNA in situ hybridization revealed novel patterns of gene expression during seed development. SCB1 mRNA accumulated first within the developing thick-walled parenchyma cells of the inner integument and later in the thick- and thin-walled parenchyma cells of the outer integument. This occurred prior to the period of seed coat maturation and seed filling and before either of the layers started to degrade. SCB1 may therefore play a role in the differentiation of the seed coat parenchyma cells. In addition, the protein product appears to be located within cell walls. The SCB1 gene codes for a new member of a class of modular proteins that possess a carboxy-terminal BURP domain and a variety of different repeated sequences. The sequence of the genomic clone revealed the insertion of a Tgm transposable element in the upstream promoter region but it is not certain whether it contributes to the tissue-specific pattern of SCB1 expression.

A seed coat-specific gene, SCS1 (Seed Coat Subtilisin 1), from soybean, Glycine max [L.] Merill, has been identified and studied. The gene belongs to a small family of genes with sequence similarity to the subtilisins, which are serine proteases. Northern blot analysis showed that SCS1 RNA accumulates to maximal levels in seed coats at 12 days post anthesis, preceding the final stages of seed coat differentiation. The SCS1 RNA was not found in other tissues including embryos, seed pods, flowers, stems, roots or leaves. In-situ hybridization studies confirmed the temporal pattern of expression observed by Northern blot analysis and further revealed a restricted pattern of RNA accumulation in thick-walled parenchyma cells of the seed coats. These cells are important in the apoplastic translocation of nutrients en route to the embryo from the vascular tissues. The tissue-specific subtilisin-like gene may be required for regulating the differentiation of the thick-walled parenchyma cells.

The soybean Ep gene encodes an anionic peroxidase enzyme that accumulates in large amounts in seed coat tissues. We have isolated a second peroxidase gene, Prx2, that is also highly expressed in developing seed coat tissues. Sequence analysis of Prx2 cDNA indicates that this transcript encodes a cationic peroxidase isozyme that is far removed from Ep in peroxidase phylogeny. To determine the expression patterns for these two peroxidases in developing seeds, the abundance and localization of the Ep and Prx2 transcripts were compared by in situ hybridization. Results show the expression of Ep begins in a small number of cells flanking the vascular bundle in the seed coat, spreads to encircle the seed, and then migrates to the hourglass cells as they develop. Expression of Prx2 occurs throughout development in all cell layers of the seed coat, and is also evident in the pericarp and embryo. Nonetheless, the Ep-encoded enzyme accounts for virtually all of the peroxidase activity detected in mature seed coats. The Prx2 enzyme is either insoluble in a catalytically inactive form, or is subject to degradation during seed maturation.