Manolo Gouy Laboratoire de Biométrie & Biologie Evolutive - CNRS / Univ. Lyon 1 January 2009

1. Recent discoveries about the molecular evolution of the three domains of life (Bacteria, Archaea, Eukaryota) Manolo Gouy Laboratoire de Biométrie & Biologie Evolutive - CNRS / Univ. Lyon 1 January 2009

2. Today the 3 domains of life LUCA First cell Origin(s) of life Third age : Post-luca cellular world LUCA: last universal common ancestor Second age : Pre-luca cellular world First age : Pre-cellular world

3. Discovery in 1977 of the three domains of life

4. SAB : similarity score between fragments of 2 rRNA molecules. SAB scores are high within each of the 3 groups and low between groups.

6. Ancestral duplication Ancestral duplication Ancestral duplication Search of the root of the universal phylogeny E: eucaryotes; B: bacteria; A: archea LUCA: Last Universal Common Ancestor

8. Current hypotheses about the origin of the three domains The most widely accepted one, based of analyses of a few pre-LUCA gene duplicates. Iconoclastic hypothesis proposed by some authors: the prokaryotic state (simple) is seen as resulting from a simplification rather than as ancestral. 4) The eukaryotic cell is seen as resulting from an archae + bacterium fusion. Several scenarios have been proposed.

9. Evolutionary history of the mitochondrial endosymbiosis • what was the donor organism ? • was the endosymbiosis unique or repeated ? • when did it occur ? what eukaryotic lineages received it ?

10. The unique endosymbiotic origin of mitochondria from an ancestral -proteobacterium -proteobacteria mitochondria Concatenation of amino acid sequences of respiratory chain proteins apocytochromeb(Cob) and cytochrome oxidase subunits 1 to 3 (Cox1-3).

11. Conservation of gene order between mitochondria and -proteobacteria

12. The gene-richest mitochondrial genome known as of today.

13. Evolutionary history of the mitochondrial endosymbiosis • what was the donor organism ? • was the endosymbiosis unique or repeated ? • when did it occur ? what eukaryotic lineages received it ?

14. Phylogenetic analysis of small subunit ribosomal RNA Mitochondrial symbiosis? Amitochondrial eucaryotes: « Archezoa » Cavalier-Smith & Chao (1996) J Mol Evol 43:551

15. Eucaryotes from before the birth of mitochondria Tom Cavalier-Smith (1987) Nature 326:332 “It is a widespread fallacy that mitochondria are found in all eukaryotic cells.” “It is not the mitochondria, but the nucleus, endomembrane system and cytoskeleton that are the true hallmarks of the eukaryote cell.” “The idea that some protozoa are the living relics of the earliest phase of eukaryote cell evolution and diverged from our ancestors before the symbiotic origin of mitochondria is given strong support by DNA sequence studies.”

16. spore polar tube Microsporidia sporoplasm (nucleus + cytoplasm) spore of Nosema algerae (Undeen 1997) • > 1000 species • unicellular eukaryotes of very small size • obligate intracellular parasites • amitochondriate, aperoxysomal • evolutionary origin subject of much debate

17. Phylogenetic analysis of b-tubulin Edlind et al. (1996) Mol. Phyl. Evol. 5:359.

18. Phylogenetic analysis of RNA polymerase II large subunit Hirt et al. (1999) Proc.Natl.Acad.Sci. USA 96:580

19. Is it possible to reconcile ribosomal RNAs, tubulins and RNA polymerases ? MICROSPORIDIA ? Amitochondrial eucaryotes

20. The Long Branch Attraction artifact[ Felsenstein (1978) Syst Zool 27:401 ] Philippe et al. (2000) Proc. Royal Soc. Lond. B 267:1213.

21. Distance-based analysis of 42 LSU rRNA sequences from microsporidia and other eukaryotes. Distances were corrected for site-to-site rate variation. So this analysis uses a more realistic model of molecular evolution. Van de Peer et al. (2000) Gene 246:1

22. Conclusion at this stage: • The nice correspondence, for microsporidia, between • absence of mitochondria • and • early origin among eucaryotes • does not hold anymore.

23. Diversity of (anaerobe) ‘amitochondrial’ protists • no detectable‘mitochondrial’ organelle (microsporidia, diplomonads, Entamoeba,…) • hydrogenosomes: genomeless organelles producing ATP and H2 (some ciliates, anaerobe fungi, Parabasalia (ex: Trichomonas)) from Roger & Silberman (2002) Nature 418:827.

24. Gene shuffling during evolution of mitochondria Ancestral -proteobacterial endosymbiont transfer to nucleus loss stay A gene of mitochondrial evolutionary origin can be carried by the nuclear genome of a eucaryote.

25. Homo-mt 100 Mus-mt 100 Caenorhabditis-mt 0.2 subst./site Candida albicans-mt 100 85 Candida maltosa-mt 100 Saccharomyces-mt 99 Schizosaccharomyces-mt 38 Encephalitozoon 86 33 Arabidopsis-mt Zygomonas Rickettsia 37 84 37 Synechocystis Escherichia Haemophilus 100 Rhodobacter capsulatus 100 Rhodobacter sphaeroides 97 Rhizobium 55 100 Klebsiella 100 Enterobacter Azospirillum 100 39 Azotobacter chroococcum 100 Azotobacter vinelandii 100 99 48 Anabaena azollae Anabaena sp 100 Helicobacter 48 59 Thermotoga Lactobacillus 69 Archaeoglobus Aquifex Deinococcus Bacillus 75 Methanobacterium 100 100 Pyrococcus 100 100 Aeropyrum Discovery in the genome of microsporidion Encephalitozoon cuniculi of several genes of mitochondrial evolutionary origin Example: IscS gene Mycoplasma sp. Mycoplasma genitalium

26. Identification of 6 putative proteins that are closer to their mitochondrial or bacterial than to their eukaryotic homologues. These E. cuniculi proteins are very probably of mitochondrial evolutionary origin. • Yeast homologues of these 6 E. cuniculi proteins : •  ATM1 (mitochondrial transporter ) •  ISU1 et ISU2 •  NFS1 (IscS cysteine desulfurase) •  SSQ1 (Heat Shock Protein 70 homologue) •  YAH1 •  PDB1 (pyruvate dehydrogenase E1 component b subunit) 5 of them are involved in the assembly of Fe-S clusters, co-factors of several mitochondrial and cytoplasmic enzymes. Katinka et al. (2001) Nature 414:450.

27. The microsporidian mitosome predicted by genome analysis: evolutionarily, it derives from a mitochondrion Vivarès et al. (2002) Current Opinion in Microbiology 5:499.

28. Detection of double- membraned organelles by anti-HSP70 antibodies

29. The mitosome of the amitochondrial parasite Entamoeba histolytica Phylogeny of CPN60 mitosome Identification by cellular mapping of the CPN60 protein in Entamoeba histolytica Tovar et al. (1999) Mol. Microbiol. 32:1013. Clark & Roger (1995) PNAS 92:6518.

30. The mitosome of the amitochondrial parasite Giardia intestinalis Identification in Giardia of genes coding for mitochondrially targeted proteins in other eukaryotes : Cpn60, Hsp70, IscS (cysteine desulfurase) One example: IscS Tachezy et al. (2001) Mol. Biol. Evol. 18:1919.

31. double membrane Immunofluorescence mapping of IscS and IscU in Giardia trophozoites.

32. The missing link between mitochondrion and hydrogenosome Discovery [Akhmanova et al. (1998) Nature 396:527] and partial sequencing of the hydrogenosomal genome of the ciliate Nyctothermus ovalis 14,027 bp fragment of the hydrogenosomal genome • Several putative proteins of the hydrogenosomal genome group with their mitochondrial homologues of aerobic ciliate . • Identification of several nuclear genes coding for components of the mitochondrial proteome (pyruvate dehydrogenase, complex II).

33. Phylogenetic analyses of 2 hydrogenosomal genome genes nad7 (s.u. 49 kDa complex I) 12S (SSU) rRNA Boxma et al. (2005) Nature 434:74.

34. Death of the concept of primitively amitochondrial eucaryotes Mitochondrial symbiosis Amitochondrial eukaryotes « Archezoa »

35. Eukaryotic distribution of mitochondrial-derived organelles Roger & Silberman (2002) Nature 418:827.

36. Evolutionary history of the chloroplastic endosymbiosis • what was the donor organism ? • was the endosymbiosis unique or repeated ? • what eukaryotic lineages received it ?

38. Demonstration of the unique origin of primary photosynthetic eukaryotes 50 plastid proteins 143 nuclear proteins

39. Model of primary chloroplastic endosymbiosis

40. Secondary chloroplastic endosymbiosis

41. Secondary chloroplastic endosymbiosis in the cryptophyte Guillardia theta

42. ? Euglenozoa Dinoflagellates Heterokonts Cryptophytes Apicomplexa Chlorarachniophytes Secondary endosymbioses Haptophytes Primary endosymbiosis Rhodophytes Glaucophytes Green plants

43. Secondary endosymbioses Euglenozoa Dinoflagellates Heterokonts Cryptophytes Euglenozoa Apicomplexa Chlorarachniophytes Secondary plastid replacement (Lepidodinium) Kryptoperidinium Chlorarachniophytes Haptophytes Dinophysis Tertiary endosymbioses Rhodophytes Glaucophytes Green plants Dinoflagellates Dinoflagellates Karenia

44. State of the art about phylogenetic knowledge at the scale of each domain of the tree of life • Much debate for bacterial and archeal domains. Is the concept of phylogenetic tree adequate ? • Eukaryotic domain phylogeny : after much confusion, some structure begins to emerge.

45. Bacterial domain phylogeny. In the « classical » vision, a natural division in phyla exists.

46. Archaeal domain phylogeny. Recent discovery of a new phylum.

47. Standard model of the tree of life naturaldivision in phyla or kingdoms

48. Alternative model: horizontal gene transfers between prokaryotes are so frequent that the notion of natural phyla does not apply.

49. Eukaryotic domain phylogeny. Emerging consensus for the identification of five super- phyla. Relationships between them remain very uncertain.

50. Models of the origin of the eukaryotic cell • the fusion hypotheses • how to test these hypotheses