The Eukaryotic Last Common Ancestor Was Bifunctional for Hopanoid and Sterol Production

Steroid and hopanoid biomarkers can be found in ancient rocks and may give a glimpse of what life was present at that time. Sterols and hopanoids are produced by two related enzymes, though the evolutionary history of this protein family is complicated by losses and horizontal gene transfers, and appears to be widely misinterpretted. Here, I have added sequences from additional key species, and re-analysis of the phylogeny of SHC and OSC indicates a single origin of both enzymes among eukaryotes. This pattern is best explained by vertical inheritance of both enzymes from a bacterial ancestor, followed by widespread loss of SHC, and two subsequent HGT events to ferns and ascomycetes. Thus, the last common ancestor of eukaryotes would have been bifunctional for both sterol and hopanoid production. Later enzymatic innovations allowed diversification of sterols in eukaryotes. Contrary to previous interpretations, the LCA of eukaryotes potentially would have been able to produce hopanoids as a substitute for sterols in anaerobic conditions. Without invoking any other metabolic demand, the LCA of eukaryotes could have been a facultative aerobe, living in unstable conditions with respect to oxygen level.

The Eukaryotic Last Common Ancestor Was Bifunctional for Hopanoid and Sterol Production

Steroid and hopanoid biomarkers can be found in ancient rocks and, in principle, can give a glimpse of what life was present at that time. Sterols and hopanoids are produced by two related enzymes, though the evolutionary history of this protein family is complicated by losses and horizontal gene transfers, and appears to be widely misinterpretted. Here, I have added sequences from additional key species, and re-analysis of the phylogeny of SHC and OSC indicates a single origin of both enzymes among eukaryotes. This pattern is best explained by vertical inheritance of both enzymes from a bacterial ancestor, followed by widespread loss of SHC, and two subsequent HGT events to ferns and ascomycetes. Thus, the last common ancestor of eukaryotes would have been bifunctional for both sterol and hopanoid production. Later enzymatic innovations allowed diversification of sterols in eukaryotes. Contrary to previous interpretations, the LCA of eukaryotes potentially would have been able to produce hopanoids as a substitute for sterols in anaerobic conditions. Without invoking any other metabolic demand, the LCA of eukaryotes could have been a facultative aerobe, living in unstable conditions with respect to oxygen level.

Anoxybacillus voinovskiensis sp. nov., a moderately thermophilic bacterium from a hot spring in Kamchatka

A novel moderately thermophilic bacterium, strain TH13T, was isolated from a hot spring in Kamchatka. It was found to be a Gram-positive, facultative aerobe; the straight, non-motile rods grew at 30–64 °C (optimum 54 °C). The isolate was positive for catalase and oxidase tests and reduced nitrate to nitrite, but was negative for H2S production and growth in more than 3 % NaCl (w/v). The isolate grew at pH 7–8, but not at pH values higher than 9. The DNA G+C content was 43·9 mol%. Phylogenetic analysis based on 16S rRNA gene sequencing indicated that strain TH13T was a member of the genus Anoxybacillus. DNA–DNA hybridization revealed a low relatedness (less than 30·2 %) between the isolate and its close phylogenetic neighbours Anoxybacillus pushchinoensis and Anoxybacillus flavithermus. On the basis of phenotypic characteristics, phylogenetic data and DNA–DNA hybridization data, it was concluded that the isolate merited classification as a novel species, for which the name Anoxybacillus voinovskiensis sp. nov. is proposed. The type strain of this species is TH13T (=NCIMB 13956T=JCM 12111T).

Nitratireductor aquibiodomus gen. nov., sp. nov., a novel α-proteobacterium from the marine denitrification system of the Montreal Biodome (Canada)

The Montreal Biodome operates a methanol-fed denitrification system that treats the water in its three million litre marine mesocosm. An unknown bacterium, named strain NL21T, was isolated from this system on TSA and R2A agar. The organism is a Gram-negative, rod-shaped (1×3 μm) facultative aerobe. Optimal growth conditions on R2A agar are 30–35 °C, pH 7–7·5 and 1 % (w/w) NaCl. Phylogenetic analysis of the 16S rDNA sequence reveals that strain NL21T forms a novel lineage in the family ‘Phyllobacteriaceae’ within the α2 subgroup of the Proteobacteria. The closest related genera are Aminobacter, Pseudaminobacter, Mesorhizobium and Defluvibacter. Major cellular fatty acids are C18 : 1 ω7c (75 %), C19 : 0 ω8c cyclopropane (9·4 %) and C18 : 0 (4·2 %). The DNA G+C content of strain NL21T (57 mol%) differs from those of all other described members of the ‘Phyllobacteriaceae’ (60–64 mol%). Strain NL21T reduces nitrate to nitrite, but does not reduce nitrite to nitrogen gas. Only a few sugars and amino acids can serve as carbon sources. Strain NL21T is able to grow without salt and tolerates up to 5 % NaCl. Phylogenetic analysis, as well as physiological and biochemical tests, showed that strain NL21T was different from all other members of the ‘Phyllobacteriaceae’ with validly published names. Strain NL21T therefore represents a novel genus, for which the name Nitratireductor aquibiodomus gen. nov., sp. nov. is proposed, with the type strain NL21T (=DSM 15645T=ATCC BAA-762T).

Analysis of Transcription of theStaphylococcus aureus Aerobic Class Ib and Anaerobic Class III Ribonucleotide Reductase Genes in Response to Oxygen

ABSTRACT Staphylococcus aureus is a gram-positive facultative aerobe that can grow in the absence of oxygen by fermentation or by using an alternative electron acceptor. To investigate the mechanism by which S. aureus is able to adapt to changes in oxygen concentration, we analyzed the transcriptional regulation of genes that encode the aerobic class Ib and anaerobic class III ribonucleotide reductase (RNR) systems that are responsible for the synthesis of deoxyribonucleotides needed for DNA synthesis. The S. aureus class Ib RNR nrdIEF and class III RNRnrdDG genes and their regulatory regions were cloned and sequenced. Inactivation of the nrdDG genes showed that the class III RNR is essential for anaerobic growth. Inhibition of aerobic growth by hydroxyurea showed that the class Ib RNR is an oxygen-dependent enzyme. Northern blot analysis and primer extension analysis demonstrated that transcription of class IIInrdDG genes is regulated by oxygen concentration and was at least 10-fold higher under anaerobic than under aerobic conditions. In contrast, no significant effect of oxygen concentration was found on the transcription of class Ib nrdIEF genes. Disruption or deletion of S. aureus nrdDG genes caused up to a fivefold increase in nrdDG and nrdIEFtranscription under anaerobic conditions but not under aerobic conditions. Similarly, hydroxyurea, an inhibitor of the class I RNRs, resulted in increased transcription of class Ib and class III RNR genes under aerobic conditions. These findings establish that transcription of class Ib and class III RNR genes is upregulated under conditions that cause the depletion of deoxyribonucleotide. Promoter analysis of class Ib and class III RNR operons identified several inverted-repeat elements that may account for the transcriptional response of thenrdIEF and nrdDG genes to oxygen.

Sulfate Reduction and Possible Aerobic Metabolism of the Sulfate-Reducing Bacterium Desulfovibrio oxyclinae in a Chemostat Coculture with Marinobacter sp. Strain MB under Exposure to Increasing Oxygen Concentrations

ABSTRACT A chemostat coculture of the sulfate-reducing bacteriumDesulfovibrio oxyclinae together with a facultative aerobe heterotroph tentatively identified as Marinobacter sp. strain MB was grown under anaerobic conditions and then exposed to a stepwise-increasing oxygen influx (0 to 20% O2 in the incoming gas phase). The coculture consumed oxygen efficiently, and no residual oxygen was detected with an oxygen supply of up to 5%. Sulfate reduction persisted at all levels of oxygen input, even at the maximal level, when residual oxygen in the growth vessel was 87 μM. The portion of D. oxyclinae cells in the coculture decreased gradually from 92% under anaerobic conditions to 27% under aeration. Both absolute cell numbers and viable cell counts of the organism were the same as or even higher than those observed in the absence of oxygen input. The patterns of consumption of electron donors and acceptors suggest that aerobic incomplete oxidation of lactate to acetate is performed by D. oxyclinae under high oxygen input. Both organisms were isolated from the same oxic zone of a cyanobacterial mat where they have to adapt to daily shifts from oxic to anoxic conditions. This type of syntrophic association may occur in natural habitats, enabling sulfate-reducing bacteria to cope with periodic exposure to oxygen.

Expression of a Tetraheme Protein,Desulfovibrio vulgaris Miyazaki F Cytochromec3, in Shewanella oneidensisMR-1

ABSTRACT Cytochrome c 3 from Desulfovibrio vulgaris Miyazaki F was successfully expressed in the facultative aerobe Shewanella oneidensis MR-1 under anaerobic, microaerophilic, and aerobic conditions, with yields of 0.3 to 0.5 mg of cytochrome/g of cells. A derivative of the broad-host-range plasmid pRK415 containing the cytochrome c 3 gene fromD. vulgaris Miyazaki F was used for transformation ofS. oneidensis MR-1, resulting in the production of protein product that was indistinguishable from that produced by D. vulgaris Miyazaki F, except for the presence of one extra alanine residue at the N terminus.

Oxygen sensing and the transcriptional regulation of oxygen-responsive genes in yeast.

The budding yeast Saccharomyces cerevisiae is a facultative aerobe that responds to changes in oxygen availability (and carbon source) by initiating a biochemically complex program that ensures that energy demands are met under two different physiological states: aerobic growth, supported by oxidative and fermentative pathways, and anaerobic growth, supported solely by fermentative processes. This program includes the differential expression of a large number of genes, many of which are involved in the direct utilization of oxygen. Research over the past decade has defined many of the cis-sites and trans-acting factors that control the transcription of these oxygen-responsive genes. However, the manner in which oxygen is sensed and the subsequent steps involved in the transduction of this signal have not been precisely determined. Heme is known to play a pivotal role in the expression of these genes, acting as a positive modulator for the transcription of the aerobic genes and as a negative modulator for the transcription of the hypoxic genes. Consequently, cellular concentrations of heme, whose biosynthesis is oxygen-dependent, are thought to provide a gauge of oxygen availability and dictate which set of genes will be transcribed. But the precise role of heme in oxygen sensing and the transcriptional regulation of oxygen-responsive genes is presently unclear. Here, we provide an overview of the transcriptional regulation of oxygen-responsive genes, address the functional roles that heme and hemoproteins may play in this regulation, and discuss possible mechanisms of oxygen sensing in this simple eukaryotic organism.