Abstract
Genes encoding enzymes of the tetrapyrrole biosynthetic pathway were searched within Euglena gracilis EST databases and 454 genome reads and their 5' end regions were sequenced when not available. Phylogenetic analyses and protein localization predictions support the hypothesis concerning the presence of two separated tetrapyrrole pathways in E. gracilis. One of these pathways resembles the heme synthesis in primarily heterotrophic eukaryotes and was presumably present in the host cell prior to secondary endosymbiosis with a green alga. The second pathway is similar to the plastid-localized tetrapyrrole syntheses in plants and photosynthetic algae. It appears to be localized to the secondary plastid, presumably derived from an algal endosymbiont and probably serves only for the production of plastidial heme and chlorophyll. Thus, E. gracilis represents an evolutionary intermediate in a metabolic transformation of a primary heterotroph to a photoautotroph through secondary endosymbiosis. We propose here that the tetrapyrrole pathway serves as a highly informative marker for the evolution of plastids and plays a crucial role in the loss of plastids.