The Essentials of Protein Import in the Degenerate Mitochondrion of Entamoeba histolytica

Abstract

Several essential biochemical processes are situated in mitochondria. The metabolic transformation of mitochondria in distinct lineages of eukaryotes created proteomes ranging from thousands of proteins to what appear to be a much simpler scenario. In the case of Entamoeba histolytica, tiny mitochondria known as mitosomes have undergone extreme reduction. Only recently a single complete metabolic pathway of sulfate activation has been identified in these organelles. The E. histolytica mitosomes do not produce ATP needed for the sulfate activation pathway and for three molecular chaperones, Cpn60, Cpn10 and mtHsp70. The already characterized ADP/ATP carrier would thus be essential to provide cytosolic ATP for these processes, but how the equilibrium of inorganic phosphate could be maintained was unknown. Finally, how the mitosomal proteins are translocated to the mitosomes had remained unclear. We used a hidden Markov model (HMM) based search of the E. histolytica genome sequence to discover candidate (i) mitosomal phosphate carrier complementing the activity of the ADP/ATP carrier and (ii) membrane-located components of the protein import machinery that includes the outer membrane translocation channel Tom40 and membrane assembly protein Sam50. Using in vitro and in vivo systems we show that E. histolytica contains a minimalist set up of the core import components in order to accommodate a handful of mitosomal proteins. The anaerobic and parasitic lifestyle of E. histolytica has produced one of the simplest known mitochondrial compartments of all eukaryotes. Comparisons with mitochondria of another amoeba, Dictystelium discoideum, emphasize just how dramatic the reduction of the protein import apparatus was after the loss of archetypal mitochondrial functions in the mitosomes of E. histolytica.

NIF-type iron-sulfur cluster assembly system is duplicated and distributed in the mitochondria and cytosol of Mastigamoeba balamuthi

“…However, following proteomic analysis of E. histolytica mitosomes has not confirmed the presence of NIF machinery (20). In our cell localization analyses, we detected components of the E. histolytica NIF machinery, particularly in the cytosolic fraction, which had been suggested previously (19). Moreover, the E. histolytica NIF components were found to be encoded by single copy genes, which lack the mitochondrial targeting signals.…”

“…Neither the cytosolic NIF components of M. balamuthi nor the NIF components of E. histolytica were imported into the mitochondria in yeast, thereby suggesting an absence of inner mitochondrial targeting signals in these proteins. However, we cannot exclude the potential recognition of a small amount of NifS and NifU through the translocation machinery of the mitosomes, which are considerably modified compared with yeast mitochondria (19), although the present experiments do not provide support for this possibility.…”

“…However, these organelles do not possess the ISC machinery, and no component of either the ISC or SUF machinery has been identified in the Entamoeba genome. NIF components have been predominantly identified in the cytosol of Entamoeba (18,19). However, it remains controversial whether these components are present exclusively in the cytosol or also operate in the mitosomes (18)(19)(20).…”

Functional Complementation Analyses Reveal that the Single PRAT Family Protein of Trypanosoma brucei Is a Divergent Homolog of Tim17 in Saccharomyces cerevisiae

“…Thus, it is likely that the microsporidian PRAT protein will also be more similar to Tim17 than to Tim23 in fungi. Parasitic protozoa, such as Giardia and Entamoeba, also possess mitosomes (52,53). Surprisingly, no member of the PRAT family of proteins has been identified in the genomes of these protozoa (52,53).…”

Biochemistry and Evolution of Anaerobic Energy Metabolism in Eukaryotes

“…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). A free-living relative of Entamoeba, Mastigamoeba balmuthii, has several enzymes of core energy metabolism in common with Entamoeba (183,564) and also possesses mitochondria that import Cpn60 (159).…”