Characterization and expression of a heptaubiquitin gene from tomato.
Ubiquitin is highly conserved 76 amino acid protein involved, among other functions, in the selective degradation of proteins in the cell. From a tomato (Lycopersicon esculentum Mill. cv. Craigella) genomic library, we have isolated a clone encoding a polyubiquitin gene, designated ubq1-1 comprising seven repeats of ubiquitin and two C-terminal extension amino acids. The ubq1-1 gene contains an intron of 1128bp immediately upstream of the translation start codon. DNA sequence comparison revealed that the 5' and 3' non-coding regions of the tomato ubq1-1 gene are nearly identical to the sequence of a polyubiquitin cDNA clone isolated from potato (Garbarino et al., 1992; Plant Mol. Biol. 20, 235-244). The ubq1-1 gene is expressed in leaves to rather low levels in tomato, and the abundance of ubq1-1 transcripts is increased under heat shock conditions. For functional analyses, a chimeric gene construct containing the intron and 1.6kb of ubq1-1 sequence 5' to the intron fused to the gus reporter gene was introduced into the tobacco genome. In leaves of transgenic tobacco plants, reporter gene expression was generally lower from the ubq1-1 promoter than from the cauliflower mosaic virus 35S RNA promoter. In addition, the tomato ubq1-1 promoter was not found to respond to heat shock in transgenic tobacco plants. Histochemical analysis of the plants demonstrated localization of gus reporter gene activity in the vascular systems of the leaves and the roots. Deletion of the intron from the reporter gene construct markedly reduced reporter gene expression in transformed tobacco plants, thus suggesting that the intron may influence transcript levels deriving from the ubq1-1 promoter.
Rollfinke
I K
IK
Botanisches Institut der Ludwig-Maxmilians Universität, Menzinger Str. 67, D-80638, München, Germany.
Silber
M V
MV
Pfitzner
U M
UM
D001483:Base Sequence; D001704:Biopolymers / Q000235:genetics*; D018744:DNA, Plant / Q000235:genetics / Q000302:isolation & purification; D015870:Gene Expression / Q000235:genetics; D017343:Genes, Plant / Q000235:genetics*; D017930:Genes, Reporter / Q000235:genetics; D018745:Genome, Plant; D005966:Glucuronidase / Q000235:genetics; D006651:Histocytochemistry; D007438:Introns / Q000235:genetics; D018551:Solanum lycopersicum / Q000737:chemistry / Q000235:genetics*; D008969:Molecular Sequence Data; D030821:Plants, Genetically Modified / Q000235:genetics; D010947:Plants, Toxic; D025821:Polyubiquitin; D011401:Promoter Regions, Genetic / Q000235:genetics / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000235:genetics; D017422:Sequence Analysis, DNA; D012689:Sequence Homology, Nucleic Acid; D014026:Nicotiana / Q000737:chemistry / Q000201:enzymology / Q000235:genetics; D014170:Transformation, Genetic; D014452:Ubiquitins / Q000235:genetics*
D001704:Biopolymers; D018744:DNA, Plant; D011993:Recombinant Fusion Proteins; D014452:Ubiquitins; D025821:Polyubiquitin; D005966:Glucuronidase