Supplementary Information for New organelles illuminate the origins of Trichomonas hydrogenosomes and Giardia mitosomes Leger, Kolisko, Kamikawa, et al. Supplementary Table 1. Glycine cleavage system protein distribution in metabolically reduced MROs. Supplementary Figure 1. PhyloBayes phylogeny of eukaryotes based on 159 proteins. Supplementary Figure 2. RAxML phylogeny of eukaryotes based on 159 proteins. Supplementary Figure 3. IQ-TREE phylogeny of eukaryotes based on 159 proteins. Supplementary Figure 4. Single-gene phylogeny of Glycine cleavage system P protein P1 domain. Supplementary Figure 5. Single-gene phylogeny of Glycine cleavage system P protein P2 domain. Supplementary Figure 6. Single-gene phylogeny of Glycine cleavage system T protein. Supplementary Figure 7. Single-gene phylogeny of Glycine cleavage system H protein. Supplementary Figure 8. Single-gene phylogeny of GCS L protein. Supplementary Figure 9. Single-gene phylogeny of HydE. Supplementary Figure 10. Single-gene phylogeny of HydF. Supplementary Figure 11. Single-gene phylogeny of HydG. Supplementary Figure 12. Single-gene phylogeny of [FeFe]-hydrogenase, HydA. Supplementary Figure 13. Single-gene phylogeny of SCS alpha subunit. Supplementary Figure 14. Single-gene phylogeny of SCS beta subunit. Supplementary Figure 15. Single-gene phylogeny of ASCT 1B proteins. Supplementary Figure 16. Single-gene phylogeny of ASCT 1C proteins. Supplementary Figure 17. Single-gene phylogeny of ACS1. Supplementary Figure 18. Single-gene phylogeny of ACS2. Supplementary Figure 19. Single-gene phylogeny of eukaryotic Cardiolipin synthase CLS_cap. Supplementary Figure 20. Single-gene phylogeny of aminoadipate-semialdehyde dehydrogenase. Supplementary Table 1 - Glycine cleavage system protein distribution in metabolically reduced MROs GCS 1 2 Survey Life styles Organisms MROs H T P L Discoba Stygiella incarcerata H Transcriptome Free-living + + + + Sawyeria marylandensis H Transcriptome Free-living + + + + Fungi Encephalitozoon cuniculi M Genome Parasitic - - - - Piromyces sp. H Transcriptome Commensal - - - - Pygsuia biforma H Transcriptome Free-living + + + + Amoebozoa Entamoeba histolytica M Genome Parasitic - - - - Mastigamoeba balamuthi H Transcriptome Free-living + + + + Rhizaria Mikrocytos mackini M Transcriptome Parasitic - - - - Alveolata Cryptosporidium parvum M Genome Parasitic - - - - Stramenopiles Cantina marsupialis H Transcriptome Free-living + + + + 1 Organisms with MROs lacking genomes, with highly reduced metabolic functions 2 hydrogenosome (H), and mitosome (M) This table is based on Stairs et al. (2015) with additional, recently published data from Leger et al. (2016) and Noguchi et al. (2015) Trimastix marina Paratrimastix pyriformis Monocercomonoides sp. Excavata Tritrichomonas foetus Trichomonas vaginalis Pentatrichomonas hominis Carpediemonas membranifera Ergobibamus cyprinoides Aduncisulcus paluster Chilomastix cuspidata Chilomastix caulleri Kipferlia bialata Dysnectes brevis Giardia intestinalis 0.94 Trepomonas sp. Spironucleus vortens Spironucleus salmonicida Spironucleus barkhanus Malawimonas jakobiformis Malawimonas californiana Collodictyon triciliatum Acanthamoeba castellanii Mastigamoeba balamuthi Physarum polycephalum Amorphea Dictyostelium discoideum Thecamonas trahens Nuclearia simplex Spizellomyces punctatus Blastocladiella emersonii Rhizopus oryzae Ustiago maydis Neurospora crassa Sphaeroforma arctica Capsaspora owczarzaki Salpingoeca rosetta Monosiga brevicolis Homo sapiens 0.81 Drosophila melanogaster Trichoplax adhaerens Nematostella vectensis 0.54 Amphmedon queenlandica Stygiella incarcerata Andalucia godoyi Jakoba bahamensis Seculamonas ecuadorensis Excavata Jakoba libera Reclinomonas americana Histiona aroides Tsukubamonas globosa Naegleria gruberi Stachyamoeba lipophora Sawyeria marylandensis Peranema trichophorum Euglena gracilis Diplonema papillatum Trypanosoma brucei Phytomonas serpens Leishmania major Glaucocystis nostochinearum Telonema subtilis Cyanidioschyzon merolae Porphyridium cruentum Diaphoretickes Calliarthron tuberculosum Gracilaria changii 0.63 Chondrus crispus Guillardia theta Pavlova lutheri Prymnesium parvum Isochrysis galbana Emiliania huxleyi Oryza sativa Arabidopsis thaliana Micromonas sp. 0.62 Polytomella parva Chlamydomonas reinhardtii 0.62 Reticulomyxa filosa Cercomonas longicauda Bigelowiella natans Blastocystis hominis Phytophthora ramorum Thalassiosira pseudonana Phaeodactylum tricornatum Oxytricha trifallax Tetrahymena thermophila Paramecium tetraurelia Perkinsus marinus Oxyrrhis marina Karenia brevis Alexandrium tamarense Cryptosporidium parvum Toxoplasma gondii Sarcocystis neurona Theileria annulata Plasmodium falciparum 0.4 Supplementary figure 1 PhyloBayes phylogeny of eukaryotes based on 159 proteins. The analysis was performed on 94 taxa and 39,089 sites, under the CAT-GTR model incorporating among-site rate variation approximated by a discrete gamma distribution with four categories (CAT-GTR + Γ model). Bayesian posterior probabilities (BPPs) are shown on nodes when BPPs are smaller than 1.0. Andalucia godoyi 99 Stygiella incarcerata Jakoba bahamensis Seculamonas ecuadorensis Jakoba libera 96 Histiona aroides 83 Reclinomonas americana Tsukubamonas globosa Naegleria gruberi 84 Stachyamoeba lipophora 98 Sawyeria marylandensis Peranema trichophorum 98 Euglena gracilis Diplonema papillatum Trypanosoma brucei Leishmania major Phytomonas serpens Trimastix marina Monocercomonoides sp. Paratrimastix pyriformis Excavata Tritrichomonas foetus Trichomonas vaginalis Pentatrichomonas hominis Carpediemonas membranifera Ergobibamus cyprinoides Aduncisulcus paluster 88 Chilomastix caulleri Chilomastix cuspidata Kipferlia bialata Dysnectes brevis Giardia intestinalis 98 Spironucleus vortens Trepomonas sp. Spironucleus barkhanus Spironucleus salmonicida Malawimonas jakobiformis 87 Malawimonas californiana Collodictyon triciliatum Acanthamoeba castellanii 98 98 Mastigamoeba balamuthi Physarum polycephalum Amorphea Dictyostelium discoideum Thecamonas trahens 87 Nuclearia simplex 84 Blastocladiella emersonii Spizellomyces punctatus Rhizopus oryzae Neurospora crassa Ustiago maydis Sphaeroforma arctica Capsaspora owczarzaki Monosiga brevicolis Salpingoeca rosetta Amphmedon queenlandica 47 Nematostella vectensis Trichoplax adhaerens 41 Homo sapiens Drosophila melanogaster Toxoplasma gondii Sarcocystis neurona Plasmodium falciparum Theileria annulata Diaphoretickes Cryptosporidium parvum Perkinsus marinus Oxyrrhis marina Karenia brevis 98 Alexandrium tamarense Oxytricha trifallax 94 Paramecium tetraurelia Tetrahymena thermophila Blastocystis hominis Phytophthora ramorum Phaeodactylum tricornatum Thalassiosira pseudonana 49 Reticulomyxa filosa Bigelowiella natans Cercomonas longicauda Oryza sativa Arabidopsis thaliana 73 Micromonas sp. 51 Chlamydomonas reinhardtii 33 Polytomella parva Glaucocystis nostochinearum Guillardia theta Pavlova lutheri 53 Prymnesium parvum Emiliania huxleyi 84 Isochrysis galbana Telonema subtilis 78 Chondrus crispus Gracilaria changii 36 Calliarthron tuberculosum Porphyridium cruentum Cyanidioschyzon merolae 0.2 Supplementary figure 2 RAxML phylogeny of eukaryotes based on 159 proteins. The analysis was performed on 94 taxa and 39,089 sites, under the LG model incorporating empirical amino acid frequencies and among-site rate variation approximated by a discrete gamma distribution with four categories (LG + Γ + F model). ML bootstrap values are shown on nodes when ML bootstrap values are smaller than 100. Andalucia godoyi Stygiella incarcerata Jakoba bahamensis Seculamonas ecuadorensis Jakoba libera 99 Histiona aroides 97 Reclinomonas americana Tsukubamonas globosa Naegleria gruberi Sawyeria marylandensis 99 Stachyamoeba lipophora Peranema trichophorum 72 Euglena gracilis Diplonema papillatum Leishmania major Phytomonas serpens Trypanosoma brucei Trimastix marina Paratrimastix pyriformis Monocercomonoides sp. Excavata Tritrichomonas foetus Trichomonas vaginalis Pentatrichomonas hominis Carpediemonas membranifera Ergobibamus cyprinoides Aduncisulcus paluster 99 Chilomastix cuspidata Chilomastix caulleri Kipferlia bialata Dysnectes brevis Giardia intestinalis Spironucleus barkhanus Spironucleus salmonicida Trepomonas sp. Spironucleus vortens Malawimonas jakobiformis 94 Malawimonas californiana Collodictyon triciliatum Nuclearia simplex Blastocladiella emersonii Spizellomyces punctatus Amorphea Neurospora crassa Ustiago maydis Rhizopus oryzae 99 Monosiga brevicolis Salpingoeca rosetta 87 Trichoplax adhaerens 71 Nematostella vectensis Drosophila melanogaster Homo sapiens Amphmedon queenlandica Capsaspora owczarzaki 99 Sphaeroforma arctica Thecamonas trahens Acanthamoeba castellanii Physarum polycephalum Dictyostelium discoideum Mastigamoeba balamuthi Toxoplasma gondii Sarcocystis neurona Plasmodium falciparum Theileria annulata Diaphoretickes Cryptosporidium parvum 99 Oxyrrhis marina Alexandrium tamarense Karenia brevis Perkinsus marinus Tetrahymena thermophila 96 Paramecium tetraurelia Oxytricha trifallax Thalassiosira pseudonana Phaeodactylum tricornatum Phytophthora ramorum Blastocystis hominis Bigelowiella natans Cercomonas longicauda Reticulomyxa filosa Arabidopsis thaliana Oryza sativa Chlamydomonas reinhardtii Polytomella parva 87 51 Micromonas sp. Prymnesium parvum 34 Isochrysis galbana Emiliania huxleyi 51 Pavlova lutheri Guillardia theta Glaucocystis nostochinearum Telonema subtilis 99 Calliarthron tuberculosum 55 Gracilaria changii 96 Chondrus crispus Porphyridium cruentum Cyanidioschyzon