ANALYSIS OF CIS-REGULATORY ELEMENTS INVOLVED IN THE ACTIVATION OF A MEMBER OF CHALCONE SYNTHASE GENE FAMILY (PSCHS1) IN PEA

Cis-regulatory elements involved in the activation of the plant defens e-related gene encoding chalcone synthase 1 (PsChs1) in pea (Pisum sat ivum L.) were examined by transient transfection, gel mobility shift a ssay and in vitro DNase I-footprinting analysis. Transient transfectio n assay revealed that a 61 bp DNA fragment spanning from -242 to -182 of PsChs1 was required for the maximal promoter activity and possibly involved in the enhancement of elicitor-mediated activation. Nuclear i solate from elicitor-treated pea epicotyl tissues contained some facto r(s) that specifically bound to this DNA fragment to form a complex wi th low mobility (LMC, low mobility complex) in gel mobility shift assa y. DNase I-footprinting analysis of LMC revealed that among three prot ected regions detected in a 61 bp DNA fragment, two regions contained identical AT-rich sequence, TAAAATACT. Site directed mutation in eithe r or both identical sequences, T<(AA)under bar>AATACT to T<(GG)under b ar>AATACT, resulted in the reduction or loss in the ability to form LM C. Detailed analysis of 61 bp DNA fragment demonstrated that the regio n from -242 to -226 containing promoter-distal TAAAATACT motif was imp erative for the maximal elicitor-mediated activation of PsChs1.