Phylogenetic analysis of the PC-PLD family. A, an unrooted tree of Arabidopsis, rice and selected non-plant PLDs, constructed by the neighbor-joining method on the basis of a MACAW-generated alignment (see Materials and Methods). C2-PLD and PXPH-PLD subfamilies are indicated by yellow and blue background, respectively. Rooting the tree with a bacterial PLD (not shown) revealed that the two subfamilies are monophyletic. B, more detailed phylogeny of C2-PLDs, based on a manually edited ClustalW alignment (tree constructed as above). Note that the topology corresponds to the previous tree, with the exception of the relationships within the cluster of PLDγ. Two major monophyletic subgroups indicated by different backgrounds appear to differ in the exon-intron organisation (as inferred from data from Arabidopsis, rice, cabbage and castor bean – only cDNA sequences are available for the remaining PLDs). The number of introns in the genes marked by asterisks (*) differs secondarily from the basic plans. AtPLDε, AtPLDζ and possibly OsPLDμ have independently acquired an additional exon (see Fig. 3), and OsPLDκ has lost 4 introns corresponding to the 3rd, 7th, 8th and 9th introns of beta, gamma, delta and nu PLDs. Numbers next to the nodes are percentages of bootstrap confidence levels calculated from 500 replicates. Trees with highly congruent topology were also obtained by a maximum parsimony method. Species abbreviations: At, Arabidopsis thaliana; Bo, Brassica oleracea; Ca, Candida albicans; Ce, Caenorhabditis elegans; Cp, Craterostigma plantagineum; Dm, Drosophila melanogaster, Gh, Gossypium hirsutum; Hs, Homo sapiens; Hv, Hordeum vulgare; Le, Lycopersicon esculentum; Nt, Nicotiana tabacum; Os, Oryza sativa; Sc, Saccharomyces cerevisiae; Rc, Ricinus communis; Vu, Vigna unguiculata; Zm, Zea mays. See Additional files for accession numbers and further sequence information.