Legends Supplemental Information
Supplemental Figure 1. DNA sequences of all rice HD-Zip I, II and III genes with translations of the ORFs. Regions of the ORFs resulting from different exons are indicated with colours.
Supplemental Figure 2.Strict consensus tree of the 23 maximum parsimony trees derived from HD-Zip I, II and III sequences from rice, Arabidopsis and C. plantagineum as aligned in Supplemental Figure 3. The tree was constructed using a heuristic search with TBR branch swapping in the PAUP program (Swofford, 2002). Family III was used as an outgroup. Bootstrap support from 500 replicates is indicated above the branches and values above 50 are shown. Family I is subdivided in clades α, β1, β2, , , , 1, 2 and (Henriksson et al., 2005, this manuscript). After each gene the intron subfamily is indicated with a box (family I), circle (family II) and triangle (family III) in different grey tones or patterns. Below: Subfamily division based on exon-intron organization. The exon-intron pattern is divided into seven subfamilies for family I (IA, IB, IC, ID, IE, IF and IG), and both family II (IIA, IIB, IIC and IID) and III (IIIA, IIIB, IIIC and IIID) have four subfamilies. Open and filled boxes represent coding and conserved domains, respectively. The HD, Zip and START domains are indicated in light grey, dark grey and black, respectively. The position of the introns is shown with black arrowheads. The white arrowhead and star (*) refers to an alternative transcript occurring with Oshox6 in subfamily IA.
Supplemental Figure 3.Multiple sequence alignment of HD-Zip family I, II and III proteins from Arabidopsis, rice and C. plantagineum. Amino acid sequences of each protein were aligned using Clustal W (Thompson et al., 1994) and revised manually. Gaps (-) were introduced for maximum alignment. Identical sequences are outlined in dark grey and similar sequences in light grey. The HD and ZIP domains are indicated in white and grey boxes respectively above the alignment. The three α-helixes in the HD are represented with black bars above the alignment. Below the alignment the first residue of each heptad of the leucine zipper is marked by the letter a, with leucine residues at position a or d.
Supplemental Figure 4. Intron-exon organization of rice and Arabidopsis HD-Zip genes and classification in different subfamilies. Every subfamily is headed by a generic sketch of the exon-intron organization (intron indicated by a black arrowhead). The genes are represented by dark grey boxes (exons) and dashes (introns). The conserved HD, Zip and START domains are coloured with green, red and yellow, respectively. In subfamily IB, Oshox6, has an alternative transcript (white arrowhead and star). (A) family I, (B) family II, and (C) family III. The scale bar is 200bp.
Supplemental Figure 5. Northern blot analysis of rice HD-Zip genes Oshox4, Oshox8, Oshox19 and Oshox22 in drought-stressed japonica rice (Oryza sativa L. ssp. japonica) varieties IRAT 112, Cabacu, Nipponbare and Taipei 309. RNA was isolated at day 0, 1 and 2 after starting of the drought-stress treatment by withholding water from two-week old seedlings. The northern blots contained 10 μg of total RNA in each lane and were probed with radioactive labelled probes of the indicated HD-Zip genes. Equal loading was verified with ethidium bromide staining.