A novel oxidative stress-inducible peroxidase promoter from sweetpotato: molecular cloning and characterization in transgenic tobacco plants and cultured cells.
A strong oxidative stress-inducible peroxidase (POD) promoter was cloned from sweetpotato (Ipomoea batatas) and characterized in transgenic tobacco plants and cultured cells in terms of environmental stress. A POD genomic clone (referred to as SWPA2) consisted of 1824 bp of sequence upstream of the translation start site, two introns (743 bp and 97 bp), and a 1073 bp coding region. SWPA2 had previously been found to encode an anionic POD which was highly expressed in response to oxidative stress. The SWPA2 promoter contained several cis-element sequences implicated in oxidative stress such as GCN-4, AP-1, HSTF, SP-1 reported in animal cells and a plant specific G-box. Employing a transient expression assay in tobacco protoplasts, with five different 5'-deletion mutants of the SWPA2 promoter fused to the beta-glucuronidase (GUS) reporter gene, the 1314 bp mutant deletion mutant showed about 30 times higher GUS expression than the CaMV 35S promoter. The expression of GUS activity in transgenic tobacco plants under the control of the -1314 SWPA2 promoter was strongly induced in response to environmental stresses including hydrogen peroxide, wounding and UV treatment. Furthermore, GUS activity in suspension cultures of transgenic cells derived from transgenic tobacco leaves containing the -1314 bp SWPA2 promoter-GUS fusion was strongly expressed after 15 days of subculture compared to other deletion mutants. We anticipate that the -1314 bp SWPA2 promoter will be biotechnologically useful for the development of transgenic plants with enhanced tolerance to environmental stress and particularly transgenic cell lines engineered to produce key pharmaceutical proteins.
Kim
Kee-Yeun
KY
Laboratory of Environmental Biotechnology, Korea Research Institute of Bioscience and Biotechnology, Oun-dong 52, Yusong-gu, Daejeon, 305-333 Korea.
Kwon
Suk-Yoon
SY
Lee
Haeng-Soon
HS
Hur
Yunkang
Y
Bang
Jae-Wook
JW
Kwak
Sang-Soo
SS
D002460:Cell Line; D003001:Cloning, Molecular; D018744:DNA, Plant / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D005091:Exons; D015971:Gene Expression Regulation, Enzymologic / Q000528:radiation effects; D018506:Gene Expression Regulation, Plant / Q000528:radiation effects; D005966:Glucuronidase / Q000235:genetics / Q000378:metabolism; D007438:Introns; D027741:Ipomoea / Q000201:enzymology / Q000235:genetics*; D008969:Molecular Sequence Data; D009154:Mutation; D018384:Oxidative Stress / Q000502:physiology; D010544:Peroxidases / Q000235:genetics*; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics; D017422:Sequence Analysis, DNA; D017384:Sequence Deletion; D013314:Stress, Mechanical; D014026:Nicotiana / Q000166:cytology / Q000235:genetics; D014466:Ultraviolet Rays
D018744:DNA, Plant; D011993:Recombinant Fusion Proteins; D010544:Peroxidases; C120494:anionic peroxidase; D005966:Glucuronidase