RAV1, a novel DNA-binding protein, binds to bipartite recognition sequence through two distinct DNA-binding domains uniquely found in higher plants.
We have cloned and characterized two novel DNA binding proteins designated RAV1 and RAV2 from Arabidopsis thaliana. RAV1 and RAV2 contain two distinct amino acid sequence domains found only in higher plant species. The N-terminal regions of RAV1 and RAV2 are homologous to the AP2 DNA-binding domain present in a family of transcription factors represented by the Arabidopsis APETALA2 and tobacco EREBP proteins, while the C-terminal region exhibits homology to the highly conserved C-terminal domain, designated B3, of VP1/ABI3 transcription factors. Binding site selection assays using a recombinant glutathione S-transferase fusion protein have revealed that RAV1 binds specifically to bipartite recognition sequences composed of two unrelated motifs, 5'-CAACA-3' and 5'-CACCTG-3', separated by various spacings in two different relative orientations. Analyses using various deletion derivatives of the RAV1 fusion protein show that the AP2 and B3-like domains of RAV1 bind autonomously to the CAACA and CACCTG motifs, respectively, and together achieve a high affinity and specificity of binding. From these results, we suggest that the AP2 and B3-like domains of RAV1 are connected by a highly flexible structure enabling the two domains to bind to the CAACA and CACCTG motifs in various spacings and orientations.
Kagaya
Y
Y
Center for Molecular Biology and Genetics and Department of Bioresources, School of Bioresources, Mie University, 1515 Kamihama-cho, Tsu 514-8507, Japan.
Ohmiya
K
K
Hattori
T
T
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000235:genetics*; D029681:Arabidopsis Proteins; D001665:Binding Sites; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D019684:Magnoliopsida / Q000235:genetics; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid; D015534:Trans-Activators; D050656:Transcription Factor AP-2; D014157:Transcription Factors / Q000235:genetics
C091601:ABI3 protein, Arabidopsis; D029681:Arabidopsis Proteins; D004268:DNA-Binding Proteins; D010940:Plant Proteins; C117695:RAV1 protein, Arabidopsis; C117696:RAV2 protein, Arabidopsis; D015534:Trans-Activators; D050656:Transcription Factor AP-2; D014157:Transcription Factors