The Arabidopsis CORI3 promoter contains two cis-acting regulatory regions required for transcriptional activity in companion cells

Abstract

Companion cells are metabolically active and functionally specialized cells that behave as terminals for the transport of materials between phloem and the surrounding tissue. Although previous research has clarified the distinct function of companion cells, it is still largely unknown how plants establish and maintain the special identity of these cells. To shed further light on this issue, we carried out expressed sequence tag (EST) analysis. To minimize the difficulty of dissociating and gathering intact companion cells, vascular strings with an abundant content of companion cells were excised from the petioles of Brassica napus. By random sequencing with a string-specific cDNA library derived by suppression subtractive hybridization between the string itself and the petiole from which it had been removed, we identified 377 ESTs and assembled them into 247 EST groups. The most frequent EST was ExBdl-102 (15 of 377 ESTs), which showed the highest sequence similarity to the Arabidopsis CORI3 (CORONATINE INDUCED 3) gene. The CORI3 promoter:GUS showed predominant expression in the vascular tissue of Arabidopsis. Through transient expression assay using Brassica vasculature and gene-gun-mediated transient assay, we found two integrated cis-regulatory regions of the CORI3 promoter. This work has provided not only string-specific EST information and shown that two novel cis-regulatory regions sustain transcriptional activity in companion cells, but also a series of procedures for efficiently examining the transcriptional framework of companion cells by exploiting the histochemical advantage of B. napus as an experimental material.