Fig. 4. The interaction between DRN and class III HD-ZIP family proteins involves the AP2 and PAS-like domains, respectively. (A)A comparison of the C-terminal regions of the Arabidopsis class III HD-ZIP proteins. Accession numbers for the proteins are as follows: CNA, AAW88440; AtHB8, CAD29660; REV AAF42938; PHV CAD29544; PHB, NP_181018. Homology is compared over the region from PHV amino acid 654 to the C-terminus. The MEKHLA domain, homology to the PAS domain from http://smart.embl-heidelberg.de and the -helices within the PHV PAS domain are marked. (B) Full-length DRN is precipitated by C-terminal regions of all class III HD-ZIP members, but not by DRNL. HA-tagged class III HD-ZIP proteins or DRNL were used for precipitation of DRN. The grid shows IP and Co-IP lanes and which antibodies were used. The position of the DRN protein is marked with an arrowhead. Asterisks represent the IgG heavy or light chains. The control lanes show IPs in the absence of either or both in vitro transcribed/translated products, confirming the identity of the lower IgG band. (C) Full-length PHV cannot co-precipitate full-length STM-GFP. The first and third lanes contain in vitro translated proteins before precipitation. PHV-HA was used for precipitation. (D) Full-length DRN can co-precipitate the 71 amino acid PHV_PAS domain (first and second lanes) and, reciprocally, PHV_PAS can co-precipitate full-length DRNL (third and fourth lanes). (E) An alignment of the AP2 domains of DRN and DRNL. Identical amino acids are shaded and the position of the unique cysteine and serine residues within the -helix are in bold. (F) The DRN AP2 domain (IP) can precipitate PHVs (Co-IP) (first and third lanes) and the DRNL AP2 domain can precipitate PHVs (second and fourth lanes). (G) 3D crystal structure of the AP2 domain of the Arabidopsis ERF1 protein (Allen et al., 1998; Marchler-Bauer et al., 2005) binding a DNA helix shown on the right, viewed from above into the helix. Cysteine/serine residues of the DRN/DRNL proteins are highlighted in yellow.