The WRKY transcription factor superfamily: its origin in eukaryotes and expansion in plants

Abstract

Background: WRKY proteins are newly identified transcription factors involved in many plant processes including plant responses to biotic and abiotic stresses. To date, genes encoding WRKY proteins have been identified only from plants. Comprehensive search for WRKY genes in non-plant organisms and phylogenetic analysis would provide invaluable information about the origin and expansion of the WRKY family.

Results: We searched all publicly available sequence data for WRKY genes. A single copy of the WRKY gene encoding two WRKY domains was identified from Giardia lamblia, a primitive eukaryote, Dictyostelium discoideum, a slime mold closely related to the lineage of animals and fungi, and the green alga Chlamydomonas reinhardtii, an early branching of plants. This ancestral WRKY gene seems to have duplicated many times during the evolution of plants, resulting in a large family in evolutionarily advanced flowering plants. In rice, the WRKY gene family consists of over 100 members. Analyses suggest that the C-terminal domain of the two-WRKY-domain encoding gene appears to be the ancestor of the single-WRKY-domain encoding genes, and that the WRKY domains may be phylogenetically classified into five groups. We propose a model to explain the WRKY family's origin in eukaryotes and expansion in plants.

Conclusions: WRKY genes seem to have originated in early eukaryotes and greatly expanded in plants. The elucidation of the evolution and duplicative expansion of the WRKY genes should provide valuable information on their functions.

Figure 1

Unrooted phylogenetic tree of the WRKY domains. The tree was reconstructed from the amino acid sequences using the neighbor-joining program from Phylip 3.57. Clades of WRKY domains are labelled according to the classifications of AtWRKY domains by Eulgem et al [3] who proposed three groups and five subgroups in Group 2 (a, b, c, d and e). We suggest classifying WRKY domains into five groups modified from the old system. While Groups 1 and 3 are unchanged, the original subgroup 2_c is promoted to Group 2_c. Subgroups 2_a and 2_b, and subgroups 2_d and 2_e are combined to form two new groups, 2_a + 2_b, and 2_d + 2_e, respectively (see text for details). WRKY domains from G. lamblia are represented by thick and dark-green branches; the slime mold, thick and cyan; the green alga, thick and magenta; Arabidopsis, thin and blue; and rice, thin and red.

Figure 2

Phylogram of Group 3 WRKY domains from Arabidopsis (AtWRKY) and rice (OsWRKY). The amino acid sequences were analysed with the neighbor-joining and parsimony algorithms implemented in PHYLIP 3.57. Bootstrap values ≥ 50% are indicated above the nodes for distance analysis. The C-terminal domains, AtWRKY1C, was used as the outgroup. OsWRKY proteins with the variant WRKYGEK are marked by *.

Figure 3

Model of the origin and duplications of WRKY gene family. The phylogenetic tree of eukaryotes using the archaea as the outgroup is modified from Baldauf and Doolittle [43] and Kenrick and Crane [50]. The solid lines correspond to branches where WRKY homologues are identified, while the thickness of the line represents the relative size of WRKY family for the branch, from the thinnest for one copy in Giardia, the slime mold and the green alga to the thickest for over 100 copies in rice. The broken lines represent branches where WRKY genes are not present or have not been identified. The WRKY gene is symbolized by the box for the WRKY domain and the lines for sequences around the domain. The text in the box indicates the group the WRKY domain belongs to (1, Group 1; 1N and 1C, N- and C-terminal domains of Group 1 proteins; a + b: Group 2_a + 2_b; c: Group 2_c; d + e, Group 2_d + 2_e; 3: Group 3). The major gene duplications and diversifications are shown above the branch. The number shown below the branch is the divergence time (million years ago) of its children branches. The branch length is not scaled to the evolutionary distance.