Intermediary metabolism in protists: a sequence-based view of facultative anaerobic metabolism in evolutionarily diverse eukaryotes

Abstract

Protists account for the bulk of eukaryotic diversity. Through studies of gene and especially genome sequences the molecular basis for this diversity can be determined. Evident from genome sequencing are examples of versatile metabolism that go far beyond the canonical pathways described for eukaryotes in textbooks. In the last 2-3 years, genome sequencing and transcript profiling has unveiled several examples of heterotrophic and phototrophic protists that are unexpectedly well-equipped for ATP production using a facultative anaerobic metabolism, including some protists that can (Chlamydomonas reinhardtii) or are predicted (Naegleria gruberi, Acanthamoeba castellanii, Amoebidium parasiticum) to produce H(2) in their metabolism. It is possible that some enzymes of anaerobic metabolism were acquired and distributed among eukaryotes by lateral transfer, but it is also likely that the common ancestor of eukaryotes already had far more metabolic versatility than was widely thought a few years ago. The discussion of core energy metabolism in unicellular eukaryotes is the subject of this review. Since genomic sequencing has so far only touched the surface of protist diversity, it is anticipated that sequences of additional protists may reveal an even wider range of metabolic capabilities, while simultaneously enriching our understanding of the early evolution of eukaryotes.

Figure 1

The organisation of eukaryotes into six major groups, each of which contains protists. Identities of the six major supergroups are highlighted by the bold font. Dotted lines reflect the uncertainty regarding the relationship between Chromalveolata and Rhizaria (Burki et al. 2008; Hampl et al. 2009). The position of the root for eukaryotic evolution is not known.

Figure 2

A simple ‘textbook’ model for central energy metabolism in eukaryotes. Dotted lines indicate intermediary metabolites are linked by multiple enzyme-catalysed reactions. Abbreviations: I, II, III, IV, V, mitochondrial complexes I, II, III, IV, and V, respectively; Ialt, alternative mitochondrial NADH:ubiquinone oxidoreductase; c, mitochondrial cytochrome c; UQ, ubiquinone.

Figure 3

Pyruvate metabolism in anaerobic protists. A, Giardia lamblia and Entamoeba histolytica. B, Trichomonas vaginalis. The compartmentalisation of energy metabolism downstream of the formation of the glycolytic intermediate phospho-enolpyruvate is summarised. Enzymes considered specific to anaerobic energy metabolism (discussed further in the main text) are numbered as follows: 1, pyruvate phosphate dikinase (PPDK); 2, PFO; 3, FeFe-hydrogenase; 4, NADH oxidase; 5, alcohol dehydrogenase E; 6, acetyl-CoA synthetase [ADP-forming]; 7, acetate:succinate CoA transferase; 8, succinyl-CoA synthetase; 9, hydrogenosomal NADH dehydrogenase (derived from 51kDa and 24 kDa sub-units of mitochondrial complex I). Note, PPDK has widely been considered to be an enzyme associated with the transition to anaerobic metabolism, and displacement of pyruvate kinase, but is in fact found in a wide variety of eukaryotes, including plants, and can operate in forward and reverse directions (see Slamovits and Keeling 2006 for further discussion).

Figure 4

Phylogenetic distribution of enzymes associated with anaerobic energy metabolism in unicellular eukaryotes. Published draft or complete genome sequences were analysed using BLAST for the presence or absence of homologues related to each of the enzymes listed. Except for two exceptions (PFO and acetate:succinate CoA transferase), a black square indicates a homologue in a genome that detects the query as the top hit in a reciprocal BLAST analysis; open square, no genome sequence available; grey square, biochemical evidence for the presence of the stated enzyme; no symbol indicates that no homologue was detected in a complete or draft genome sequence. For PFO homologues, a black square indicates a homologue with the canonical eukaryotic/eubacterial PFO domain architecture (Rotte et al. 2001); black circle, homologue where the PFO domain architecture is C-terminally fused to a NADPH-cytochrome P450 reductase module providing either known or putative (Perkinsus marinus) PNO activity (as described in the main text); black triangle, homologue exhibits similarity to eukaryotic/eubacterial PFO domain architecture and contains a c-terminal ‘cysI’ domain. CysI is the domain characterising one of the sub-units of multi-meric assimilatory sulfite reductase; a requirement for the PFO-related CysI-containing homologue from Saccharomyces cerevisiae (Met5) in sulphur assimilation has recently been reported (Cordente et al. 2009). Phototrophs, including all of the algae surveyed here, possess CysI-containing polypeptides that lack the associated PFO-related domain architecture. The ‘mix’n’match’ combinations of domains in PFO-related proteins has been noted previously, and possibly reflects the co-option of the PFO domain for use in numerous redox reactions (Hug et al. 2010; Rotte et al. 2001). For acetate:succinate CoA transferase homologues, a black triangle indicates cloning and biochemical verification of a ‘sub-family 1A’ homologue (related to mammalian SCOT transferase; Tielens et al. 2010); grey triangle, homologue of a ‘sub-family 1A’ enzyme detected in a genome; grey circle, homologue of a ‘sub-family 1B’ acetate:succinate CoA transferase detected in a genome; black pentagon, cloning and biochemical verification of a ‘sub-family 1C’ acetate:succinate CoA transferase; grey pentagon, ‘sub-family 1C’ homologue detected in a genome. Note: Amoebidium parasiticum, Acanthamoeba castellanii, and Euglena gracilis have no genome sequence available, but are shown here because biochemical and/or molecular data indicate the presence of stated enzymes, and thus, point towards intriguing signatures of anaerobic metabolism. In contrast to the widespread distribution of enzymes required for anaerobic ATP production, the only heterotrophs that contain flavo-diiron protein as defence against oxygen toxicity are obligate anaerobes/microaerophiles. The presence of flavo-diiron proteins in some oxygenic phototrophs, and their putative role in photosystem II maintenance was noted previously (Zhang et al. 2009).