Repression of the defense gene PR-10a by the single-stranded DNA binding protein SEBF.
The potato pathogenesis-related gene PR-10a is transcriptionally activated in response to pathogen infection or elicitor treatment. Characterization of the cis-acting elements of the PR-10a promoter revealed the presence of a silencing element between residues -52 and -27 that contributes to transcriptional regulation. In this study, we have isolated a silencing element binding factor (SEBF) from potato tuber nuclei that binds to the coding strand of the silencing element in a sequence-specific manner. The consensus binding site of SEBF, PyTGTCNC, is present in a number of PR genes and shows striking similarity to the auxin response element. Mutational analysis of the PR-10a promoter revealed an inverse correlation between the in vitro binding of SEBF and the expression of PR-10a. SEBF was purified to homogeneity from potato tubers, and sequencing of the N terminus of the protein led to the isolation of a cDNA clone. Sequence analysis revealed that SEBF is homologous with chloroplast RNA binding proteins that possess consensus sequence-type RNA binding domains characteristic of heterogeneous nuclear ribonucleoproteins (hnRNPs). Overexpression of SEBF in protoplasts repressed the activity of a PR-10a reporter construct in a silencing element-dependent manner, confirming the role of SEBF as a transcriptional repressor.
Boyle
B
B
Department of Biochemistry, Université de Montréal, Montréal, Québec, Canada H3C 3J7.
Brisson
N
N
D000595:Amino Acid Sequence; D001483:Base Sequence; D002467:Cell Nucleus / Q000378:metabolism / Q000502:physiology; D002736:Chloroplasts / Q000378:metabolism*; D003001:Cloning, Molecular; D017931:DNA Primers; D004277:DNA, Single-Stranded / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D018506:Gene Expression Regulation, Plant; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics*; D018515:Plant Leaves / Q000502:physiology; D010940:Plant Proteins; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism; D016415:Sequence Alignment; D011198:Solanum tuberosum / Q000235:genetics*; D014158:Transcription, Genetic
D017931:DNA Primers; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; C416516:PBF-2 protein, Solanum tuberosum; D010940:Plant Proteins; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D012097:Repressor Proteins