Nucleotide sequence and putative regulatory elements of gene 2 of the soybean (Glycine max) chalcone synthase multigene family.

Abstract

CHS is the first committed enzyme of flavonoid biosynthesis catalyzing the condensation of one molecule of pcoumaroyl-CoA with three molecules of malonyl-CoA to give rise to naringenin chalcone (Hahlbrook and Scheel, 1989). This chalcone is the central intermediate in the biosynthesis of compounds that provide much of the coloring in nature, are involved in various physiological processes including protection against UV-B radiation and microbial infection, and play an important role in nodulation in leguminous plants (Lamb et al., 1989). We have identified seven members of the chs multigene family in the genome of Glycine max L. Merr. cv Williams and in this communication we present the nucleotide sequence and some of the characteristics of a gene initially designated as gene 2 by Wingender et al. (1989). Unlike genes 1, 3, and 4 (Akada et al., 1991, and refs. therein), which exist as a gene cluster spanning about a 10kb region of the soybean genome, gene 2 seems to exist independently. However, gene 2 is still very similar to genes 1, 3, and 4 in that exon I has 178 bp and exon II has 989 bp of protein coding sequences, constituting an open reading frame of 1167 bp encoding a polypeptide of 388 amino adds. The intron (137 bp) for gene 2 is slightly larger than for genes 1, 3, and 4 (121-122 bp), but the nucleotide sequence in the protein-coding region of gene 2 is quite similar to that of genes 1, 3, and 4 (Table I). In spite of sequence diversity in the 5' nontranslated region, the consensus sequence for a TATA box in this gene exists in the same location as in genes 1, 3, and 4 (Table l). The basic integrity of gene 2 suggests that this gene is not a pseudogene but quite possibly a functional gene. Earlier work on CHS isoforms (Grab et al., 1985; Welle and Griesebach, 1987) suggested that at least six different members of the chs gene family are expressed in soybean suspensionculture cells. Gene 2 may well be one of them, because our computer-calculated isoelectric point for a protein of the amino acid sequence derived from the gene 2 sequence is pH 6.41, which is in good agreement with the observed isoelectric point of about pH 6.4 for a sugar-induced CHS from suspension-culture cells (Welle and Griesebach, 1987). Consensus sequences for some regulatory elements including a sugarresponsive element are found in the 5' flanking region of gene 2 (Table I, Fig. 1).