Marek Mentel

Biochemistry and evolution of anaerobic energy metabolism in eukaryotes.

SUMMARY Major insights into the phylogenetic distribution, biochemistry, and evolutionary significance of organelles involved in ATP synthesis (energy metabolism) in eukaryotes that thrive in anaerobic environments for all or part of their life cycles have accrued in recent years. All known eukaryotic groups possess an organelle of mitochondrial origin, mapping the origin of mitochondria to the eukaryotic common ancestor, and genome sequence data are rapidly accumulating for eukaryotes that possess anaerobic mitochondria, hydrogenosomes, or mitosomes. Here we review the available biochemical data on the enzymes and pathways that eukaryotes use in anaerobic energy metabolism and summarize the metabolic end products that they generate in their anaerobic habitats, focusing on the biochemical roles that their mitochondria play in anaerobic ATP synthesis. We present metabolic maps of compartmentalized energy metabolism for 16 well-studied species. There are currently no enzymes of core anaerobic energy metabolism that are specific to any of the six eukaryotic supergroup lineages; genes present in one supergroup are also found in at least one other supergroup. The gene distribution across lineages thus reflects the presence of anaerobic energy metabolism in the eukaryote common ancestor and differential loss during the specialization of some lineages to oxic niches, just as oxphos capabilities have been differentially lost in specialization to anoxic niches and the parasitic life-style. Some facultative anaerobes have retained both aerobic and anaerobic pathways. Diversified eukaryotic lineages have retained the same enzymes of anaerobic ATP synthesis, in line with geochemical data indicating low environmental oxygen levels while eukaryotes arose and diversified.

Formation of new chromosomes as a virulence mechanism in yeast Candida glabrata.

In eukaryotes, the number and rough organization of chromosomes is well preserved within isolates of the same species. Novel chromosomes and loss of chromosomes are infrequent and usually associated with pathological events. Here, we analyzed 40 pathogenic isolates of a haploid and asexual yeast, Candida glabrata, for their genome structure and stability. This organism has recently become the second most prevalent yeast pathogen in humans. Although the gene sequences were well conserved among different strains, their chromosome structures differed drastically. The most frequent events reshaping chromosomes were translocations of chromosomal arms. However, also larger segmental duplications were frequent and occasionally we observed novel chromosomes. Apparently, this yeast can generate a new chromosome by duplication of chromosome segments carrying a centromere and subsequently adding novel telomeric ends. We show that the observed genome plasticity is connected with antifungal drug resistance and it is likely an advantage in the human body, where environmental conditions fluctuate a lot.

Energy metabolism among eukaryotic anaerobes in light of Proterozoic ocean chemistry

Recent years have witnessed major upheavals in views about early eukaryotic evolution. One very significant finding was that mitochondria, including hydrogenosomes and the newly discovered mitosomes, are just as ubiquitous and defining among eukaryotes as the nucleus itself. A second important advance concerns the readjustment, still in progress, about phylogenetic relationships among eukaryotic groups and the roughly six new eukaryotic supergroups that are currently at the focus of much attention. From the standpoint of energy metabolism (the biochemical means through which eukaryotes gain their ATP, thereby enabling any and all evolution of other traits), understanding of mitochondria among eukaryotic anaerobes has improved. The mainstream formulations of endosymbiotic theory did not predict the ubiquity of mitochondria among anaerobic eukaryotes, while an alternative hypothesis that specifically addressed the evolutionary origin of energy metabolism among eukaryotic anaerobes did. Those developments in biology have been paralleled by a similar upheaval in the Earth sciences regarding views about the prevalence of oxygen in the oceans during the Proterozoic (the time from ca 2.5 to 0.6 Ga ago). The new model of Proterozoic ocean chemistry indicates that the oceans were anoxic and sulphidic during most of the Proterozoic. Its proponents suggest the underlying geochemical mechanism to entail the weathering of continental sulphides by atmospheric oxygen to sulphate, which was carried into the oceans as sulphate, fuelling marine sulphate reducers (anaerobic, hydrogen sulphide-producing prokaryotes) on a global scale. Taken together, these two mutually compatible developments in biology and geology underscore the evolutionary significance of oxygen-independent ATP-generating pathways in mitochondria, including those of various metazoan groups, as a watermark of the environments within which eukaryotes arose and diversified into their major lineages.

Protein Import into Hydrogenosomes of Trichomonas vaginalis Involves both N-Terminal and Internal Targeting Signals: a Case Study of Thioredoxin Reductases

ABSTRACT The parabasalian flagellate Trichomonas vaginalis harbors mitochondrion-related and H2-producing organelles of anaerobic ATP synthesis, called hydrogenosomes, which harbor oxygen-sensitive enzymes essential to its pyruvate metabolism. In the human urogenital tract, however, T. vaginalis is regularly exposed to low oxygen concentrations and therefore must possess antioxidant systems protecting the organellar environment against the detrimental effects of molecular oxygen and reactive oxygen species. We have identified two closely related hydrogenosomal thioredoxin reductases (TrxRs), the hitherto-missing component of a thioredoxin-linked hydrogenosomal antioxidant system. One of the two hydrogenosomal TrxR isoforms, TrxRh1, carried an N-terminal extension resembling known hydrogenosomal targeting signals. Expression of hemagglutinin-tagged TrxRh1 in transfected T. vaginalis cells revealed that its N-terminal extension was necessary to import the protein into the organelles. The second hydrogenosomal TrxR isoform, TrxRh2, had no N-terminal targeting signal but was nonetheless efficiently targeted to hydrogenosomes. N-terminal presequences from hydrogenosomal proteins with known processing sites, i.e., the alpha subunit of succinyl coenzyme A synthetase (SCSα) and pyruvate:ferredoxin oxidoreductase A, were investigated for their ability to direct mature TrxRh1 to hydrogenosomes. Neither presequence directed TrxRh1 to hydrogenosomes, indicating that neither extension is, by itself, sufficient for hydrogenosomal targeting. Moreover, SCSα lacking its N-terminal extension was efficiently imported into hydrogenosomes, indicating that this extension is not required for import of this major hydrogenosomal protein. The finding that some hydrogenosomal enzymes require N-terminal signals for import but that in others the N-terminal extension is not necessary for targeting indicates the presence of additional targeting signals within the mature subunits of several hydrogenosome-localized proteins.

The Physiology of Phagocytosis in the Context of Mitochondrial Origin

SUMMARY How mitochondria came to reside within the cytosol of their host has been debated for 50 years. Though current data indicate that the last eukaryote common ancestor possessed mitochondria and was a complex cell, whether mitochondria or complexity came first in eukaryotic evolution is still discussed. In autogenous models (complexity first), the origin of phagocytosis poses the limiting step at eukaryote origin, with mitochondria coming late as an undigested growth substrate. In symbiosis-based models (mitochondria first), the host was an archaeon, and the origin of mitochondria was the limiting step at eukaryote origin, with mitochondria providing bacterial genes, ATP synthesis on internalized bioenergetic membranes, and mitochondrion-derived vesicles as the seed of the eukaryote endomembrane system. Metagenomic studies are uncovering new host-related archaeal lineages that are reported as complex or phagocytosing, although images of such cells are lacking. Here we review the physiology and components of phagocytosis in eukaryotes, critically inspecting the concept of a phagotrophic host. From ATP supply and demand, a mitochondrion-lacking phagotrophic archaeal fermenter would have to ingest about 34 times its body weight in prokaryotic prey to obtain enough ATP to support one cell division. It would lack chemiosmotic ATP synthesis at the plasma membrane, because phagocytosis and chemiosmosis in the same membrane are incompatible. It would have lived from amino acid fermentations, because prokaryotes are mainly protein. Its ATP yield would have been impaired relative to typical archaeal amino acid fermentations, which involve chemiosmosis. In contrast, phagocytosis would have had great physiological benefit for a mitochondrion-bearing cell.

Of early animals, anaerobic mitochondria, and a modern sponge

The origin and early evolution of animals marks an important event in lifes history. This event is historically associated with an important variable in Earth history – oxygen. One view has it that an increase in oceanic oxygen levels at the end of the Neoproterozoic Era (roughly 600 million years ago) allowed animals to become large and leave fossils. How important was oxygen for the process of early animal evolution? New data show that some modern sponges can survive for several weeks at low oxygen levels. Many groups of animals have mechanisms to cope with low oxygen or anoxia, and very often, mitochondria – organelles usually associated with oxygen – are involved in anaerobic energy metabolism in animals. It is a good time to refresh our memory about the anaerobic capacities of mitochondria in modern animals and how that might relate to the ecology of early metazoans.

BH3‐only proteins Noxa, Bik, Bmf, and Bid activate Bax and Bak indirectly when studied in yeast model

BH3-only proteins of the Bcl-2 family regulate programmed cell death in mammals through activation of multidomain proapoptotic proteins Bax and Bak in response to various proapoptotic stimuli by mechanism that remains under dispute. Here, we report that the cell death-promoting activity of BH3-only proteins Bik, Bmf, Noxa, and tBid can only be reconstituted in yeast when both multidomain proapoptotic and antiapoptotic Bcl-2 family proteins are present. Inability of these proteins to induce cell death in the absence of antiapoptotic proteins suggests that all tested BH3-only proteins likely activate Bax and Bak indirectly by inhibiting antiapoptotic proteins.

Transfer of genetic material between pathogenic and food-borne yeasts.

ABSTRACT Many pathogenic yeast species are asexual and therefore not involved in intra- or interspecies mating. However, high-frequency transfer of plasmid DNA was observed when pathogenic and food-borne yeasts were grown together. This property could play a crucial role in the spread of virulence and drug resistance factors among yeasts.

BH3-only protein Bim inhibits activity of antiapoptotic members of Bcl-2 family when expressed in yeast.

Proteins of the Bcl‐2 family regulate programmed cell death in mammals by promoting the release of cytochrome c from mitochondria in response to various proapoptotic stimuli. The mechanism by which BH3‐only members of the family activate multidomain proapoptotic proteins Bax and Bak to form a pore in mitochondrial membranes remains under dispute. We report that cell death promoting activity of BH3‐only protein Bim can be reconstituted in yeast when both Bax and antiapoptotic protein Bcl‐XL are present, suggesting that Bim likely activates Bax indirectly by inhibiting antiapoptotic proteins.

Isolation and expression of the gene encoding mitochondrial ADP/ATP carrier (AAC) from the pathogenic yeast Candida parapsilosis.

A gene homologous to Saccharomyces cerevisiae AAC genes coding for mitochondrial ADP/ATP carriers has been cloned from the pathogenic yeast Candida parapsilosis. A probe obtained by PCR amplification from C. parapsilosis DNA, using primers derived from the conserved transmembrane region of yeast ADP/ATP carriers, was used for screening of the C. parapsilosis genomic library. The cloned gene was sequenced and found to encode a polypeptide of 303 amino acids that shows homology with other yeast and fungal mitochondrial ADP/ATP carriers. The gene was designated CpAAC1 and was able to complement the growth phenotypes of S. cerevisiae double deletion mutant (Δaac2; Δaac3). The expression of the CpAAC1 gene was reduced under semi‐anaerobic conditions and it was affected at normal aerobic conditions by the nature of carbon sources used for growth. Hybridization experiments indicate that C. parapsilosis possesses a single gene encoding a mitochondrial ADP/ATP carrier. The GenBank Accession No. for the C. parapsilosis CpAAC1 gene is AF085429. Copyright