- Email: [email protected]
A Mitochondrion in Entamoeba histolytica?
Comment
A Mitochondrion in Entamoeba histolytica? M. Müller Entamoeba histolytica, a protist parasite of the human intestine, is famous for its ‘simple’ cell structure1–4 and ‘primitive’, ‘bacterium-like’ metabolism4,5, and has been regarded as an ancestral eukaryote1,4. A prominent reason for this is that E. histolytica contains no structure with the morphological characteristics of a mitochondrion3 nor the enzymes of typical mitochondrial energy metabolism1,6. However, two papers on a newly recognized structure, published almost simultaneously, will dramatically change our views on E. histolytica. Tovar et al.7 and Mai et al.8 have reported the existence and properties of a small organelle, named, respectively, ‘mitosome’ or ‘crypton’, a discovery also discussed recently in Trends in Microbiology9. The organelle shares several biological characteristics with mitochondria and is likely to have descended from a common ancestor. The existence of such an organelle, a mitochondrial ‘remnant,’ has been predicted since the pioneering detection of two genes of putative mitochondrial origin, encoding the proteins chaperonin 60 (cpn60) and pyridine nucleotide (NAD/NADP) transhydrogenase10 in E. histolytica. Now that this putative organelle has been discovered, it opens up an important area of comparative cell biology and eukaryotic evolution to experimental research. The critical observation made by both groups was the localization of one of the products of these genes (cpn60) in a small structure in E. histolytica by confocal fluorescence microscopy and cell fractionation7,8. Most cells contained only one such organelle but a few cells, possibly those ready to divide, contained two, or occasionally three. Mai et al.8 also used antibodies reacting with proteins of other subcellular compartments (nucleus, endoplasmic reticulum, Golgi apparatus and cytosol), which showed distinctly different localizations in the cell. This supports the conclusion that the organelle seen is an entity sui generis. The fluorescent microscopic images (Fig. 5 in Ref. 7 and Fig. 7a and 7b in Ref. 8) do not permit the exact measurement of the size of the organelle, but do suggest a diameter of 1–2 mm. The size of an amoeba is approximately 20 mm; thus, the organelle cannot represent .0.1% of the total cell volume. The identification of these structures as mitochondrial ‘remnants’ is based on 368
two findings: First, the protein detected, cpn60, is known to be restricted to mitochondria, hydrogenosomes and chloroplasts11. Sequence analysis and phylogenetic reconstruction placed the E. histolytica cpn60 robustly in a clade together with its mitochondrial and hydrogenosomal homologs9,12, indicating an origin from a common ancestral gene. The most parsimonious explanation is that this ancestral gene was acquired through the endosymbiotic event that led to the establishment of the ancestor of these organelles12, although alternative scenarios cannot be entirely rejected. Second, the fate of E. histolytica cpn60 is identical to that of its mitochondrial and hydrogenosomal homologs7,8. The protein is encoded by nuclear genes and translation occurs on free ribosomes in the cytosol. The product is imported into the organelle post-translationally, a process in which a processed N-terminal targeting peptide plays a crucial role. When cpn60 was encoded by constructs in which the codons for the targeting peptide had been deleted, the protein did not enter the organelle but remained in the cytosol. If the targeting sequence was replaced by the targeting sequence of a mitochondrial protein of Trypanosoma cruzi, the protein was found in the small organelle in E. histolytica7. Cells of E. histolytica contain many membrane-bound vacuoles and vesicles of diverse sizes3. Thus, it should not come as a surprise that, without the use of specific markers, a unique small body had not been previously recognized. When the organelle is identified by immunoelectron microscopy, it should be possible to pinpoint it on previously published electron micrographs. It is expected that, like mitochondria, the organelle will be enveloped by two membranes. It is not clear what else the organelle contains. A nuclear gene for a second ‘mitochondrial-type’ protein, NAD/NADP transhydrogenase, has also been found in E. histolytica9, although it remains to be established whether this protein gets imported into the organelle. A key metabolic enzyme, pyruvate : ferredoxin oxidoreductase, has been localized to, among other structures, small cytoplasmic bodies in E. histolytica13. However, the relationship of this structure to the
0169-4758/00/$ – see front matter © 2000 Elsevier Science Ltd. All rights reserved. PII: S0169-4758(00)01732-4
organelles discussed here remains to be elucidated. The small size of the organelle indicates that it is not a major site of metabolic flow, in contrast to mitochondria, hydrogenosomes and peroxisomes. Mitochondria usually occupy .10% of the cell volume, but can represent as much as one-third of it, eg. in hepatocytes, parabasalid hydrogenosomes are of the order of 10%. Peroxisomes with an active role in carbohydrate catabolism or anabolism (glycosomes in kinetoplastids, plant peroxisomes involved in photorespiration and plant glyoxysomes) also make a significant contribution to the total cell volume. Thus, one will have to look for functions that, while vital, are performed by enzymes in low copy number. When the genome of this organism is sequenced (an event which cannot be too far off), a clearer image of the composition of the organelle should emerge, although its role will be harder to determine. It is likely that identical organelles will be detected in all Entamoeba spp and related parasitic amoebae (Endolimax, and possibly Jodamoeba). However, Dientamoeba fragilis, found in the human colon, usually discussed together with other enteric amoebae, is an exception. Molecular data14 support earlier proposals that D. fragilis is a parabasalid, related to trichomonads15. The observation that it harbors morphologically identifiable hydrogenosomes substantiates this taxonomic position15. The findings discussed here support the idea that Entamoeba spp and their relatives descended from an ancestor that had experienced the ‘mitochondrial endosymbiotic event’, and their present ‘simple’ cytological makeup is actually a secondary characteristic. In essence, they appear to have evolved by functional and morphological losses. Of these, perhaps the most dramatic is the loss of the major metabolic functions of mitochondria and the retention of an, essentially cytosolic, extended glycolysis6,16. The reconstruction of this evolutionary history is hampered by the still unclarified position of Entamoeba spp among eukaryotes. They have no freeliving relatives with similarly simple morphology and energy metabolism, and molecular data on their position have been conflicting17. Based on diverse criteria, Cavalier-Smith18 suggested that Parasitology Today, vol. 16, no. 9, 2000
Comment their closest relatives are the slimemolds (eg. Dictyostelium and Physarum) and free-living ‘amitochondriate’ amoeboflagellates called pelobionts (eg. Mastigamoeba and the giant amoeba, Pelomyxa). He placed all these organisms in the newly established subphylum, the Conosa of the phylum Amoebozoa. Sophisticated analyses of small-subunit rRNA sequences support the relationship between entamoebids and pelobionts and do not contradict the relationship of these with the slimemolds17. A more detailed comparison of all these organisms is likely to shed light on the evolutionary history of entamoebids, which might just represent the most extreme examples of reductive evolution among those eukaryotes that did not establish an intracellular parasitic mode of life. Is the occurrence of homologous or analogous organelles possible in other organisms? Likely candidates are the diplomonads, with Giardia and Hexamita as the best-known parasitic representatives. These organisms do not contain mitochondria or hydrogenosomes as organelles of core metabolism, although a gene with mitochondrial ancestry encoding cpn60 has been detected in G. lamblia12. The free-living pelobionts also have small intracellular bodies that could be similar in nature to those found in E. histolytica19. The study of only a few unicellular eukaryotes revealed an unexpected structural and metabolic diversity compatible with the eukaryotic phenotype. Among these are the ‘amitochondriate’ protists, which showed that this phenotype is not linked obligatorily to the presence of a mitochondrion performing oxidative phosphorylation6,16. In the past two decades, the origin of such ‘amitochondriate’ organisms has become a hotly debated topic of evolutionary studies16,20,21. The view that ‘amitochondriate’ protists are relics of the ancestral, premitochondriate eukaryotic cell is now less popular. Mounting evidence indicates that all the ‘amitochondriate’ eukaryotes studied to date arose from ancestors that experienced the same endosymbiotic event that led to the establishment of the ancestral mitochondrion16,20. Perhaps the most studied examples are the parabasalids, primarily the trichomonads. Their characteristic metabolic organelle, the hydrogenosome, is generally considered a derivative of the ancestral mitochondrial symbiont22,23. Hydrogenosomes, possibly differing in their composition, have also been detected in other protists and anaerobic fungi24. The findings of Parasitology Today, vol. 16, no. 9, 2000
Tovar et al.7 and Mai et al.8 have revealed another organelle that probably arose from the same source. Future studies need to include an increasing number of unicellular eukaryotes in order to uncover the varied endpoints of mitochondrial diversification and reconstruct the evolutionary events leading to them25. Acknowledgements Original research in the laboratory of MM is supported by National Institutes of Health Grant AI11942 and National Science Foundation Grant MBC 9615659. References 1 Reeves, R.E. (1984) Metabolism of Entamoeba histolytica Schaudinn, 1903. Adv. Parasitol. 23, 105–142 2 McLaughlin, J. and Aley, S. (1985) The biochemistry and functional morphology of Entamoeba. J. Protozool. 32, 221–240 3 Martinez-Palomo, A. (1982) The Biology of Entamoeba histolytica, Research Studies Press 4 Bakker-Grunwald, T. and Wöstmann, C. (1993) Entamoeba histolytica as a model for the primitive eukaryotic cell. Parasitol. Today 9, 27–31 5 Samuelson, J. (1999) Why metronidazole is active against bacteria and parasites. Antimicrob. Agents Chemother. 43, 1533–1541 6 Müller, M. (1998) Enzymes and compartmentalization of core energy metabolism of anaerobic protists – a special case in eukaryotic evolution? in Evolutionary relationships among protozoa (Coombs, G.H. et al., eds), pp 109–131, Kluwer Scientific 7 Tovar, J. et al. (1999) The mitosome, a novel organelle related to mitochondria in the amitochondrial parasite, Entamoeba histolytica. Mol. Microbiol. 32, 1013–1021 8 Mai, Z. et al. (1999) Hsp60 is targeted to a cryptic mitochondrion-derived organelle (‘crypton’) in the microaerophilic protozoan parasite Entamoeba histolytica. Mol. Cell. Biol. 19, 2198–2205 9 Boore, J.L. and Fuerstenberg, S.I. (1999) Entamoeba histolytica: a derived, mitochondriate eukaryote? Trends Microbiol. 7, 426–428 10 Clark, C.G. and Roger, A.J. (1995) Direct evidence for secondary loss of mitochondria in Entamoeba histolytica. Proc. Natl. Acad. Sci. U. S. A. 92, 6518–6521 11 Gupta, R.S. (1995) Evolution of the chaperonin families (Hsp60, Hsp10 and Tcp-1) of proteins
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and the origin of eukaroytic cells. Mol. Microbiol. 15, 1–11 Roger, A.J. et al. (1998) A mitochondrial-like chaperonin 60 gene in Giardia lamblia: evidence that diplomonads once harbored an endosymbiont related to the progenitor of mitochondria. Proc. Natl. Acad. Sci. U. S. A. 95, 229–234 Rodriguez, M.A. et al. (1998) The pyruvate : ferredoxin oxidoreductase enzyme is localized in the plasma membrane and in a cytoplasmic structure in Entamoeba. Microb. Pathogen. 25, 1–10 Silberman, J.D. et al. (1996) Dientamoeba fragilis shares a recent common evolutionary history with the trichomonads. Mol. Biochem. Parasitol. 76, 311–314 Camp, R.R. et al. (1974) Study of Dientamoeba fragilis Jepps & Dobell. I. Electromicroscopic observations of the binucleate stages. II. Taxonomic position and revision of the genus. J. Protozool. 21, 69–82 Martin, W.F. and Müller, M. (1998) The hydrogen hypothesis of the first eukaryote. Nature 392, 37–41 Silberman, J.D. et al. (1999) Phylogeny of the genera Entamoeba and Endolimax as deduced from small-subunit ribosomal RNA sequences. Mol. Biol. Evol. 16, 1740–1751 Cavalier-Smith, T. (1998) A revised sixkingdom system of life. Biol. Rev. 73, 203–266 Simpson, A.G.B. et al. (1997) The organisation of Mastigamoeba schizophrenia n.sp.: more evidence of ultrastructural idiosyncrasy and simplicity of pelobiont protists. Eur. J. Protistol. 33, 87–98 Roger, A.J. (1999) Reconstructing early events in eukaryotic evolution. Am. Nat. 154 (suppl.), S146–S163 Embley, T.M. and Hirt, R.P. (1998) Early branching eukaryotes? Curr. Opin. Gen. Develop. 8, 624–629 Müller, M. (1997) Evolutionary origin of hydrogenosomes. Parasitol. Today 13, 166–167 Plümper, E. et al. (1998) Implications of protein import on the origin of hydrogenosomes. Protist 149, 303–311 Hackstein, J.H.P. et al. (1999) Hydrogenosomes: eukaryotic adaptations to anaerobic environments. Trends Microbiol. 7, 441–447 Embley, T.M. and Martin, W. (1998) A hydrogen-producing mitochondrion. Nature 396, 517–518
Miklós Müller is at the Rockfeller University, 1230 York Avenue, New York, NY 10021, USA. Tel: +1 212 327 8153, Fax: +1 212 327 7974, e-mail: mmuller@rockvax. rockefeller.edu
Organizing a Meeting? Are you organizing a meeting that is of interest to parasitologists? If so, please supply Parasitology Today with the details (dates, title, location) and a contact name, and we will include your meeting in our Diary page. Please make sure we receive the details at least 8–10 weeks prior to the meeting dates. Send details to: [email protected] 369
A Mitochondrion in Entamoeba histolytica? M. Müller Entamoeba histolytica, a protist parasite of the human intestine, is famous for its ‘simple’ cell structure1–4 and ‘primitive’, ‘bacterium-like’ metabolism4,5, and has been regarded as an ancestral eukaryote1,4. A prominent reason for this is that E. histolytica contains no structure with the morphological characteristics of a mitochondrion3 nor the enzymes of typical mitochondrial energy metabolism1,6. However, two papers on a newly recognized structure, published almost simultaneously, will dramatically change our views on E. histolytica. Tovar et al.7 and Mai et al.8 have reported the existence and properties of a small organelle, named, respectively, ‘mitosome’ or ‘crypton’, a discovery also discussed recently in Trends in Microbiology9. The organelle shares several biological characteristics with mitochondria and is likely to have descended from a common ancestor. The existence of such an organelle, a mitochondrial ‘remnant,’ has been predicted since the pioneering detection of two genes of putative mitochondrial origin, encoding the proteins chaperonin 60 (cpn60) and pyridine nucleotide (NAD/NADP) transhydrogenase10 in E. histolytica. Now that this putative organelle has been discovered, it opens up an important area of comparative cell biology and eukaryotic evolution to experimental research. The critical observation made by both groups was the localization of one of the products of these genes (cpn60) in a small structure in E. histolytica by confocal fluorescence microscopy and cell fractionation7,8. Most cells contained only one such organelle but a few cells, possibly those ready to divide, contained two, or occasionally three. Mai et al.8 also used antibodies reacting with proteins of other subcellular compartments (nucleus, endoplasmic reticulum, Golgi apparatus and cytosol), which showed distinctly different localizations in the cell. This supports the conclusion that the organelle seen is an entity sui generis. The fluorescent microscopic images (Fig. 5 in Ref. 7 and Fig. 7a and 7b in Ref. 8) do not permit the exact measurement of the size of the organelle, but do suggest a diameter of 1–2 mm. The size of an amoeba is approximately 20 mm; thus, the organelle cannot represent .0.1% of the total cell volume. The identification of these structures as mitochondrial ‘remnants’ is based on 368
two findings: First, the protein detected, cpn60, is known to be restricted to mitochondria, hydrogenosomes and chloroplasts11. Sequence analysis and phylogenetic reconstruction placed the E. histolytica cpn60 robustly in a clade together with its mitochondrial and hydrogenosomal homologs9,12, indicating an origin from a common ancestral gene. The most parsimonious explanation is that this ancestral gene was acquired through the endosymbiotic event that led to the establishment of the ancestor of these organelles12, although alternative scenarios cannot be entirely rejected. Second, the fate of E. histolytica cpn60 is identical to that of its mitochondrial and hydrogenosomal homologs7,8. The protein is encoded by nuclear genes and translation occurs on free ribosomes in the cytosol. The product is imported into the organelle post-translationally, a process in which a processed N-terminal targeting peptide plays a crucial role. When cpn60 was encoded by constructs in which the codons for the targeting peptide had been deleted, the protein did not enter the organelle but remained in the cytosol. If the targeting sequence was replaced by the targeting sequence of a mitochondrial protein of Trypanosoma cruzi, the protein was found in the small organelle in E. histolytica7. Cells of E. histolytica contain many membrane-bound vacuoles and vesicles of diverse sizes3. Thus, it should not come as a surprise that, without the use of specific markers, a unique small body had not been previously recognized. When the organelle is identified by immunoelectron microscopy, it should be possible to pinpoint it on previously published electron micrographs. It is expected that, like mitochondria, the organelle will be enveloped by two membranes. It is not clear what else the organelle contains. A nuclear gene for a second ‘mitochondrial-type’ protein, NAD/NADP transhydrogenase, has also been found in E. histolytica9, although it remains to be established whether this protein gets imported into the organelle. A key metabolic enzyme, pyruvate : ferredoxin oxidoreductase, has been localized to, among other structures, small cytoplasmic bodies in E. histolytica13. However, the relationship of this structure to the
0169-4758/00/$ – see front matter © 2000 Elsevier Science Ltd. All rights reserved. PII: S0169-4758(00)01732-4
organelles discussed here remains to be elucidated. The small size of the organelle indicates that it is not a major site of metabolic flow, in contrast to mitochondria, hydrogenosomes and peroxisomes. Mitochondria usually occupy .10% of the cell volume, but can represent as much as one-third of it, eg. in hepatocytes, parabasalid hydrogenosomes are of the order of 10%. Peroxisomes with an active role in carbohydrate catabolism or anabolism (glycosomes in kinetoplastids, plant peroxisomes involved in photorespiration and plant glyoxysomes) also make a significant contribution to the total cell volume. Thus, one will have to look for functions that, while vital, are performed by enzymes in low copy number. When the genome of this organism is sequenced (an event which cannot be too far off), a clearer image of the composition of the organelle should emerge, although its role will be harder to determine. It is likely that identical organelles will be detected in all Entamoeba spp and related parasitic amoebae (Endolimax, and possibly Jodamoeba). However, Dientamoeba fragilis, found in the human colon, usually discussed together with other enteric amoebae, is an exception. Molecular data14 support earlier proposals that D. fragilis is a parabasalid, related to trichomonads15. The observation that it harbors morphologically identifiable hydrogenosomes substantiates this taxonomic position15. The findings discussed here support the idea that Entamoeba spp and their relatives descended from an ancestor that had experienced the ‘mitochondrial endosymbiotic event’, and their present ‘simple’ cytological makeup is actually a secondary characteristic. In essence, they appear to have evolved by functional and morphological losses. Of these, perhaps the most dramatic is the loss of the major metabolic functions of mitochondria and the retention of an, essentially cytosolic, extended glycolysis6,16. The reconstruction of this evolutionary history is hampered by the still unclarified position of Entamoeba spp among eukaryotes. They have no freeliving relatives with similarly simple morphology and energy metabolism, and molecular data on their position have been conflicting17. Based on diverse criteria, Cavalier-Smith18 suggested that Parasitology Today, vol. 16, no. 9, 2000
Comment their closest relatives are the slimemolds (eg. Dictyostelium and Physarum) and free-living ‘amitochondriate’ amoeboflagellates called pelobionts (eg. Mastigamoeba and the giant amoeba, Pelomyxa). He placed all these organisms in the newly established subphylum, the Conosa of the phylum Amoebozoa. Sophisticated analyses of small-subunit rRNA sequences support the relationship between entamoebids and pelobionts and do not contradict the relationship of these with the slimemolds17. A more detailed comparison of all these organisms is likely to shed light on the evolutionary history of entamoebids, which might just represent the most extreme examples of reductive evolution among those eukaryotes that did not establish an intracellular parasitic mode of life. Is the occurrence of homologous or analogous organelles possible in other organisms? Likely candidates are the diplomonads, with Giardia and Hexamita as the best-known parasitic representatives. These organisms do not contain mitochondria or hydrogenosomes as organelles of core metabolism, although a gene with mitochondrial ancestry encoding cpn60 has been detected in G. lamblia12. The free-living pelobionts also have small intracellular bodies that could be similar in nature to those found in E. histolytica19. The study of only a few unicellular eukaryotes revealed an unexpected structural and metabolic diversity compatible with the eukaryotic phenotype. Among these are the ‘amitochondriate’ protists, which showed that this phenotype is not linked obligatorily to the presence of a mitochondrion performing oxidative phosphorylation6,16. In the past two decades, the origin of such ‘amitochondriate’ organisms has become a hotly debated topic of evolutionary studies16,20,21. The view that ‘amitochondriate’ protists are relics of the ancestral, premitochondriate eukaryotic cell is now less popular. Mounting evidence indicates that all the ‘amitochondriate’ eukaryotes studied to date arose from ancestors that experienced the same endosymbiotic event that led to the establishment of the ancestral mitochondrion16,20. Perhaps the most studied examples are the parabasalids, primarily the trichomonads. Their characteristic metabolic organelle, the hydrogenosome, is generally considered a derivative of the ancestral mitochondrial symbiont22,23. Hydrogenosomes, possibly differing in their composition, have also been detected in other protists and anaerobic fungi24. The findings of Parasitology Today, vol. 16, no. 9, 2000
Tovar et al.7 and Mai et al.8 have revealed another organelle that probably arose from the same source. Future studies need to include an increasing number of unicellular eukaryotes in order to uncover the varied endpoints of mitochondrial diversification and reconstruct the evolutionary events leading to them25. Acknowledgements Original research in the laboratory of MM is supported by National Institutes of Health Grant AI11942 and National Science Foundation Grant MBC 9615659. References 1 Reeves, R.E. (1984) Metabolism of Entamoeba histolytica Schaudinn, 1903. Adv. Parasitol. 23, 105–142 2 McLaughlin, J. and Aley, S. (1985) The biochemistry and functional morphology of Entamoeba. J. Protozool. 32, 221–240 3 Martinez-Palomo, A. (1982) The Biology of Entamoeba histolytica, Research Studies Press 4 Bakker-Grunwald, T. and Wöstmann, C. (1993) Entamoeba histolytica as a model for the primitive eukaryotic cell. Parasitol. Today 9, 27–31 5 Samuelson, J. (1999) Why metronidazole is active against bacteria and parasites. Antimicrob. Agents Chemother. 43, 1533–1541 6 Müller, M. (1998) Enzymes and compartmentalization of core energy metabolism of anaerobic protists – a special case in eukaryotic evolution? in Evolutionary relationships among protozoa (Coombs, G.H. et al., eds), pp 109–131, Kluwer Scientific 7 Tovar, J. et al. (1999) The mitosome, a novel organelle related to mitochondria in the amitochondrial parasite, Entamoeba histolytica. Mol. Microbiol. 32, 1013–1021 8 Mai, Z. et al. (1999) Hsp60 is targeted to a cryptic mitochondrion-derived organelle (‘crypton’) in the microaerophilic protozoan parasite Entamoeba histolytica. Mol. Cell. Biol. 19, 2198–2205 9 Boore, J.L. and Fuerstenberg, S.I. (1999) Entamoeba histolytica: a derived, mitochondriate eukaryote? Trends Microbiol. 7, 426–428 10 Clark, C.G. and Roger, A.J. (1995) Direct evidence for secondary loss of mitochondria in Entamoeba histolytica. Proc. Natl. Acad. Sci. U. S. A. 92, 6518–6521 11 Gupta, R.S. (1995) Evolution of the chaperonin families (Hsp60, Hsp10 and Tcp-1) of proteins
12
13
14
15
16 17
18 19
20 21 22 23 24 25
and the origin of eukaroytic cells. Mol. Microbiol. 15, 1–11 Roger, A.J. et al. (1998) A mitochondrial-like chaperonin 60 gene in Giardia lamblia: evidence that diplomonads once harbored an endosymbiont related to the progenitor of mitochondria. Proc. Natl. Acad. Sci. U. S. A. 95, 229–234 Rodriguez, M.A. et al. (1998) The pyruvate : ferredoxin oxidoreductase enzyme is localized in the plasma membrane and in a cytoplasmic structure in Entamoeba. Microb. Pathogen. 25, 1–10 Silberman, J.D. et al. (1996) Dientamoeba fragilis shares a recent common evolutionary history with the trichomonads. Mol. Biochem. Parasitol. 76, 311–314 Camp, R.R. et al. (1974) Study of Dientamoeba fragilis Jepps & Dobell. I. Electromicroscopic observations of the binucleate stages. II. Taxonomic position and revision of the genus. J. Protozool. 21, 69–82 Martin, W.F. and Müller, M. (1998) The hydrogen hypothesis of the first eukaryote. Nature 392, 37–41 Silberman, J.D. et al. (1999) Phylogeny of the genera Entamoeba and Endolimax as deduced from small-subunit ribosomal RNA sequences. Mol. Biol. Evol. 16, 1740–1751 Cavalier-Smith, T. (1998) A revised sixkingdom system of life. Biol. Rev. 73, 203–266 Simpson, A.G.B. et al. (1997) The organisation of Mastigamoeba schizophrenia n.sp.: more evidence of ultrastructural idiosyncrasy and simplicity of pelobiont protists. Eur. J. Protistol. 33, 87–98 Roger, A.J. (1999) Reconstructing early events in eukaryotic evolution. Am. Nat. 154 (suppl.), S146–S163 Embley, T.M. and Hirt, R.P. (1998) Early branching eukaryotes? Curr. Opin. Gen. Develop. 8, 624–629 Müller, M. (1997) Evolutionary origin of hydrogenosomes. Parasitol. Today 13, 166–167 Plümper, E. et al. (1998) Implications of protein import on the origin of hydrogenosomes. Protist 149, 303–311 Hackstein, J.H.P. et al. (1999) Hydrogenosomes: eukaryotic adaptations to anaerobic environments. Trends Microbiol. 7, 441–447 Embley, T.M. and Martin, W. (1998) A hydrogen-producing mitochondrion. Nature 396, 517–518
Miklós Müller is at the Rockfeller University, 1230 York Avenue, New York, NY 10021, USA. Tel: +1 212 327 8153, Fax: +1 212 327 7974, e-mail: mmuller@rockvax. rockefeller.edu
Organizing a Meeting? Are you organizing a meeting that is of interest to parasitologists? If so, please supply Parasitology Today with the details (dates, title, location) and a contact name, and we will include your meeting in our Diary page. Please make sure we receive the details at least 8–10 weeks prior to the meeting dates. Send details to: [email protected] 369