A cis-acting element and a trans-acting factor involved in the wound-induced expression of a horseradish peroxidase gene.
The mechanisms that control the wound-induced expression of the prxC2 gene for horseradish peroxidase (HRP) have been investigated. Analysis of the regulatory properties of 5'-deleted promoters showed that a positive element involved in the response to wounding was located between -307 and -99 bp from the site of initiation of translation. In in vitro binding assays of tobacco nuclear proteins and DNA fragments of prxC2 promoter, the binding site was the Box 1 from -296 to -283 containing the CACGTG motif. To identify the functional role of Box 1, the prxC2 promoter that has been digested from the 5' end to -289 with a disrupted Box 1 was fused to a reporter gene for beta-glucuronidase (GUS). No induction of GUS activity was observed in transgenic tobacco plants with the prxC2 (-289)/GUS construct. These data indicated that the expression of prxC2 in response to wounding required the Box 1 sequence from -296 to -283. Furthermore, a tobacco cDNA expression library was screened and a cDNA clone for a protein, designated TFHP-1, that bound specifically to the Box 1 sequence was identified. The putative TFHP-1 protein contains a basic region and leucine zipper (bZip) motif and a helix-loop-helix (HLH) motif. The mRNA for TFHP-1 was abundant in roots and stems, and it was not induced by wounding in leaves. In tobacco protoplasts, antisense TFHP-1 suppressed the expression of prxC2(-529)/GUS.
Kawaoka
A
A
Department of Biotechnology, Faculty of Engineering, Osaka University, Suita, Japan.
Kawamoto
T
T
Sekine
M
M
Yoshida
K
K
Takano
M
M
Shinmyo
A
A
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D015342:DNA Probes; D004268:DNA-Binding Proteins / Q000096:biosynthesis*; D004330:Drosophila; D004926:Escherichia coli; D015870:Gene Expression; D015971:Gene Expression Regulation, Enzymologic; D015723:Gene Library; D005966:Glucuronidase / Q000032:analysis / Q000096:biosynthesis; D018257:Helix-Loop-Helix Motifs; D006735:Horseradish Peroxidase / Q000096:biosynthesis* / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D018521:Plant Physiological Phenomena; D010940:Plant Proteins; D010944:Plants / Q000201:enzymology / Q000235:genetics; D030821:Plants, Genetically Modified; D010947:Plants, Toxic; D011401:Promoter Regions, Genetic; D017433:Protein Structure, Secondary; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D015183:Restriction Mapping; D017386:Sequence Homology, Amino Acid; D014026:Nicotiana / Q000235:genetics; D015534:Trans-Activators / Q000096:biosynthesis*; D014947:Wounds and Injuries
D015342:DNA Probes; D004268:DNA-Binding Proteins; D010940:Plant Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; D006735:Horseradish Peroxidase; C089537:TFHP-1 protein, Nicotiana tabacum; D005966:Glucuronidase