Studies on DNA-binding selectivity of WRKY transcription factors lend structural clues into WRKY-domain function.
WRKY transcription factors have been shown to play a major role in regulating, both positively and negatively, the plant defense transcriptome. Nearly all studied WRKY factors appear to have a stereotypic binding preference to one DNA element termed the W-box. How specificity for certain promoters is accomplished therefore remains completely unknown. In this study, we tested five distinct Arabidopsis WRKY transcription factor subfamily members for their DNA binding selectivity towards variants of the W-box embedded in neighboring DNA sequences. These studies revealed for the first time differences in their binding site preferences, which are partly dependent on additional adjacent DNA sequences outside of the TTGACY-core motif. A consensus WRKY binding site derived from these studies was used for in silico analysis to identify potential target genes within the Arabidopsis genome. Furthermore, we show that even subtle amino acid substitutions within the DNA binding region of AtWRKY11 strongly impinge on its binding activity. Additionally, all five factors were found localized exclusively to the plant cell nucleus and to be capable of trans-activating expression of a reporter gene construct in vivo.
Department of Plant Microbe Interactions, Max Planck Institute for Plant Breeding Research, Carl-von-Linné-Weg 10, Koln, Germany.
Ingo
I
Ciolkowski
Dierk
D
Wanke
Rainer P
RP
Birkenbihl
Imre E
IE
Somssich
D000595:Amino Acid Sequence; D029681:Arabidopsis Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002467:Cell Nucleus / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D029681:Arabidopsis Proteins; D004268:DNA-Binding Proteins; D014157:Transcription Factors; C517320:WRKY11 protein, Arabidopsis; C446285:WRKY6 protein, Arabidopsis