Characterization of the chalcone synthase multigene family and the temporal pattern of chalcone synthasemRNA expression in developing soybean seedlings

Colleen Beth Jonsson, Purdue University

Abstract

The structure, organization, and organ-regulated expression of the chalcone synthase (CHS) gene family was examined in soybean (Glycine max (L.) Merr). This gene family has at least six members and two other genes that hybridize at low stringency. Four genomic clones containing six of the CHS genes were isolated. One CHS gene was fully sequenced and a second CHS gene was sequenced in areas flanking the coding region. The open reading frame for one genomic clone, GmaCHS300A, was identified by sequence comparison with the CHS gene from Phaseolus vulgarus (Ryder et al., 1987). The coding region of 1167 bp was split by one intron and has 80% similarity at the nucleotide level with the P. vulgaris CHS cDNA. Comparisons of the deduced amino acid sequence of several dicot CHS genes with the CHS sequence deduced from soybean showed amino acid identities ranging from 82% (Arabidopsis thaliana) to 88% (P. vulgaris). The 5$\sp\prime$ upstream regions of two of the soybean CHS genes shared promoter motifs that were conserved in similar positions in other plant CHS genes and pathogen-defense related genes. The relative levels of CHS mRNA were found to be more abundant in roots than in hypocotyls or cotyledons of young soybean seedlings. No detectable hybridization was observed in seeds and leaves. Immature embryos, flower buds and flowers with petals were also examined in field grown soybeans. Amongst these tissues, only flowers with petals had CHS RNA, but at a very low level. Phenyalanine ammonia-lyase RNA was expressed in the same organs as CHS. CHS showed a temporally regulated pattern of activity in roots, hypocotyls, and cotyledons in young seedlings during each day over the several days examined. CHS enzyme activity, however, did not reflect this pattern of RNA expression. Instead, hypocotyls and root CHS RNA levels increased each day, but were fairly constant throughout a given day. Increases or decreases in activity usually occurred between midnight and 6 A.M. Finally, the levels of CHS RNA in soybean cv. Forrest leaves increased in response to Pseudomonas syringae pv. glycinae Race 4 with and without the avrA gene. The response of the leaves to the pathogen differed in the amount and the time of maximal accumulation of CHS mRNA, as has been seen in previous experiments (Dhawale et al., 1989).

Degree

Ph.D.

Advisors

Herrmann, Purdue University.

Subject Area

Biochemistry|Molecular biology

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