A system for tissue-specific copper-controllable gene expression in transgenic plants: nodule-specific antisense of aspartate aminotransferase-P2.

Abstract

A vector system, based on copper controllable gene expression, has been developed to give control over place as well as time of expression of an introduced gene. This system consists of two elements: (1) the yeast ace1 gene encoding a metallo-regulatory transcription factor, ACE1, under control of either an organ-specific or a constitutive promoter; and (2) a gene of interest under control of a chimaeric promoter consisting of the 46 bp TATA fragment of the CaMV 35S RNA promoter linked to four repeats of the ACE1 binding site. The functioning of the system in an organ-specific manner was tested in nodulated Lotus corniculatus plants which consisted of non-transformed shoots plus transformed hairy root tissue 'wild-type tops/transgenic roots'. After addition of copper ions to the plant nutrient solution, beta-glucuronidase (GUS) expression was visualized either specifically in nodules or in both roots and nodules when the ace1 gene was placed under control of the nod45 promoter or the CaMV 35S RNA promoter, respectively. The nodule-specific system was used to express antisense constructs of aspartate aminotransferase-P2 in transgenic Lotus corniculatus plants. When expression was induced by the addition of copper ions to the plant nutrient solution aspartate aminotransferase-P2 activity declined dramatically, and a decrease of up to 90% was observed in nodule asparagine concentration.