Open AccessIdentification of potato nuclear proteins binding to the distal promoter region of the proteinase inhibitor II gene.
Author(s) -
José J. SánchezSerrano,
Hugo PeñaCortés,
Lothar Willmitzer,
Salomé Prat
Publication year - 1990
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.87.18.7205
Subject(s) - oligonucleotide , dna , nuclear protein , biology , biochemistry , microbiology and biotechnology , dna binding protein , gene , electrophoretic mobility shift assay , binding protein , binding site , guanine , nucleic acid sequence , nucleotide , gene expression , transcription factor
Potato nuclear proteins specifically bind to a DNA sequence at the most 5' distal region of the promoter of a potato proteinase inhibitor II gene. Binding studies using the electrophoretic mobility-shift assay showed the appearance of two protein-DNA complexes in the presence of both tuber and leaf nuclear protein extracts. Mechanical wounding of the leaves had no effect on the amount of specific protein-DNA complexes formed. DNase I protection analysis and binding to synthetic oligonucleotides identified the sequence 5'-GAGGGTATTTTCGTAA-3' as the target for the noncooperative binding of two potato nuclear proteins to the upstream element. Methylation interference experiments showed that guanine nucleotides separated by one turn of the DNA helix were in close contact with the proteins. The binding ability of a series of mutated synthetic oligonucleotides further defined the sequence requirements for protein binding, which appeared to contact one side of the DNA helix.