Biochemical and Structural Insights into the Mechanism of DNA Recognition by Arabidopsis ETHYLENE INSENSITIVE3.
Gaseous hormone ethylene regulates numerous stress responses and developmental adaptations in plants by controlling gene expression via transcription factors ETHYLENE INSENSITIVE3 (EIN3) and EIN3-Like1 (EIL1). However, our knowledge regarding to the accurate definition of DNA-binding domains (DBDs) within EIN3 and also the mechanism of specific DNA recognition by EIN3 is limited. Here, we identify EIN3 82-352 and 174-306 as the optimal and core DBDs, respectively. Results from systematic biochemical analyses reveal that both the number of EIN3-binding sites (EBSs) and the spacing length between two EBSs affect the binding affinity of EIN3; accordingly, a new DNA probe which has higher affinity with EIN3 than ERF1 is also designed. Furthermore, we show that palindromic repeat sequences in ERF1 promoter are not necessary for EIN3 binding. Finally, we provide, to our knowledge, the first crystal structure of EIN3 core DBD, which contains amino acid residues essential for DNA binding and signaling. Collectively, these data suggest the detailed mechanism of DNA recognition by EIN3 and provide an in-depth view at molecular level for the transcriptional regulation mediated by EIN3.
State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, and Peking-Tsinghua Center for Life Sciences, Peking University, Beijing, China.
Jinghui
J
Song
State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, and Peking-Tsinghua Center for Life Sciences, Peking University, Beijing, China.
Chenxu
C
Zhu
State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, and Peking-Tsinghua Center for Life Sciences, Peking University, Beijing, China.
Xing
X
Zhang
State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, and Peking-Tsinghua Center for Life Sciences, Peking University, Beijing, China.
Xing
X
Wen
State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, and Peking-Tsinghua Center for Life Sciences, Peking University, Beijing, China.
Lulu
L
Liu
State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, and Peking-Tsinghua Center for Life Sciences, Peking University, Beijing, China.
Jinying
J
Peng
State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, and Peking-Tsinghua Center for Life Sciences, Peking University, Beijing, China.
Hongwei
H
Guo
State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, and Peking-Tsinghua Center for Life Sciences, Peking University, Beijing, China; Department of Chemical Biology and Synthetic and Functional Biomolecules Center, College of Chemistry and Molecular Engineering, Peking University, Beijing, China.
Chengqi
C
Yi
D017360:Arabidopsis / Q000737:chemistry / Q000235:genetics*; D029681:Arabidopsis Proteins / Q000737:chemistry* / Q000235:genetics; D001665:Binding Sites; D018360:Crystallography, X-Ray; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics; D024202:Electrophoretic Mobility Shift Assay; D005030:Ethylenes / Q000378:metabolism; D018506:Gene Expression Regulation, Plant; D055029:Inverted Repeat Sequences; D009687:Nuclear Proteins / Q000737:chemistry* / Q000235:genetics; D010454:Peptide Termination Factors / Q000737:chemistry / Q000235:genetics; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D015398:Signal Transduction; D014157:Transcription Factors / Q000737:chemistry* / Q000235:genetics
D029681:Arabidopsis Proteins; D004268:DNA-Binding Proteins; C494589:EIL1 protein, Arabidopsis; C106987:EIN3 protein, Arabidopsis; D005030:Ethylenes; D009687:Nuclear Proteins; D010454:Peptide Termination Factors; D014157:Transcription Factors; C494978:eukaryotic release factor 1, Arabidopsis; C036216:ethylene