Mitosomes in Entamoeba histolytica contain a sulfate activation pathway

Abstract

Hydrogenosomes and mitosomes are mitochondrion-related organelles in anaerobic/microaerophilic eukaryotes with highly reduced and divergent functions. The full diversity of their content and function, however, has not been fully determined. To understand the central role of mitosomes in Entamoeba histolytica, a parasitic protozoon that causes amoebic dysentery and liver abscesses, we examined the proteomic profile of purified mitosomes. Using 2 discontinuous Percoll gradient centrifugation and MS analysis, we identified 95 putative mitosomal proteins. Immunofluorescence assay showed that 3 proteins involved in sulfate activation, ATP sulfurylase, APS kinase, and inorganic pyrophosphatase, as well as sodium/sulfate symporter, involved in sulfate uptake, were compartmentalized to mitosomes. We have also provided biochemical evidence that activated sulfate derivatives, adenosine-5-phosphosulfate and 3-phosphoadenosine-5-phosphosulfate, were produced in mitosomes. Phylogenetic analysis showed that the aforementioned proteins and chaperones have distinct origins, suggesting the mosaic character of mitosomes in E. histolytica consisting of proteins derived from ␣-proteobacterial, ␦-proteobacterial, and ancestral eukaryotic origins. These results suggest that sulfate activation is the major function of mitosomes in E. histolytica and that E. histolytica mitosomes represent a unique mitochondrion-related organelle with remarkable diversity.anaerobic protozoa ͉ evolution ͉ mitochondria ͉ organelle ͉ proteomics D iversification of mitochondrial structure and function has occurred during eukaryotic evolution, and was especially observed in anaerobic/microaerophilic environments. Most extant anaerobic eukaryotes, which were previously considered to lack mitochondria, are now regarded to possess reduced and highly divergent forms of mitochondrion-related organelles (1, 2). The hydrogenosome is an organelle in which hydrogen and ATP are produced, and is found in anaerobic protists and fungi such as Trichomonas vaginalis (3, 4), Neocallimastix patriciarum (5, 6), and Nyctotherus ovalis (7). The mitosome, typically demonstrated in parasitic and free-living protists such as E. histolytica (2,(8)(9)(10)(11)(12)(13)(14)(15), Giardia intestinalis (16, 17), diverse microsporidian species (18)(19)(20), and Cryptosporidium parvum (21), generally has reduced functions and does not produce hydrogen or ATP. In Mastigamoeba balamuthi, a mitochondrion-related organelle was discovered and presumed to possess a unique array of biochemical properties, although it remains unclear whether the organelle is more similar to either hydrogenosomes or mitosomes (22). In contrast, the mitochondrion-related organelle in Blastocystis contains DNA and shows characteristics for both hydrogenosomes and mitochondria of higher eukaryotes (23). Organisms that possess hydrogenosomes and mitosomes do not cluster together in eukaryote phylogenies, indicating that secondary losses and changes in mitochondrial functions have independently occurred multiple times i...

Biochemistry and Evolution of Anaerobic Energy Metabolism in Eukaryotes

“…The Entamoeba mitosome possesses a mitochondrial-type ADP/ATP translocator (80); its function is still under investigation (520). Recently, the enzymes of sulfate activation were localized to the mitosomes of Entamoeba (332,333), providing a new perspective on mitosomal functions. Other possible functions might have to do with FeS cluster maturation and hydroperoxide detoxification via the rubrerythrin system (302).…”

“…Mitosomes of some lineages have retained components of FeS cluster assembly (163,508), and others have retained components of sulfate activation (332).…”

“…Pyruvate:ferredoxin oxidoreductase is used for pyruvate decarboxylation/oxidation, as in parabasalian parasites; ATP is synthesized through substrate-level phosphorylation via acetyl-CoA synthetase (ADP forming) (431). Although not involved in energy metabolism, mitosomes in E. histolytica harbor enzymes of sulfate activation as a major function (332). Abbreviations: Fd, ferredoxin; [1], pyruvate:ferredoxin oxidoreductase; [5], malic enzyme; [9], pyruvate kinase; [11], malate dehydrogenase; [18], alanine aminotransferase; [25], alcohol dehydrogenase E; [28], phosphoenolpyruvate carboxytransferase (PP i dependent); [29], pyruvate:orthophosphate dikinase; [30] Because of the potential of the mitosome to harbor drug targets, protein import into Entamoeba mitosomes has been intensely studied as of late (118).…”

Evolution of the Cytosolic Iron-Sulfur Cluster Assembly Machinery in Blastocystis Species and Other Microbial Eukaryotes

“…For example, the microaerophiles Entamoeba and Mastigamoeba both possess functionally reduced MROs that do not contain the typical ISC pathway. Instead, their ge-nomes encode NIF system components that were acquired by lateral gene transfer (LGT) from epsilonproteobacteria and that localize both in the cytosol and in their organelles (14)(15)(16). Another example is found in Blastocystis, an obligate anaerobic parasite that encodes a functional fused version of the SufC and SufB proteins, SufCB, that was also acquired by LGT, in this case, from a methanoarchaeal lineage, and that functions in the cytosol of the organism (17).…”

Sawyeria marylandensis (Heterolobosea) Has a Hydrogenosome with Novel Metabolic Properties

“…Some of these mitosomes, such as those of Giardia intestinalis (54) and some Microsporidia (22), possess mitochondrion-like protein import apparatuses and seem to function in Fe-S cluster biogenesis. Other mitosomes, such as those of Entamoeba, retain mitochondrial heat shock and chaperonin proteins (2, 32) yet seem to carry out biochemical functions, such as sulfur activation (34), that are not typically associated with aerobic mitochondria. Recent investigations of little-studied protists have further revealed a continuum of diversity of function in MROs, from organelles that are complex metabolic intermediates between mitochondria and hydrogenosomes (e.g., Nyctotherus ovalis [7] and Blastocystis sp.…”