Three enzymes governed the rise of O2 on Earth.

Current views of O2 accumulation in Earth history depict three phases: The onset of O2 production by ∼2.4 billion years ago; 2 billion years of stasis at ∼1 % of modern atmospheric levels; and a rising phase, starting about 500 million years ago, in which oxygen eventually reached modern values. Purely geochemical mechanisms have been proposed to account for this tripartite time course of Earth oxygenation. In particular the second phase, the long period of stasis between the advent of O2 and the late rise to modern levels, has posed a puzzle. Proposed solutions involve Earth processes (geochemical, ecosystem, day length). Here we suggest that Earth oxygenation was not determined by geochemical processes. Rather it resulted from emergent biological innovations associated with photosynthesis and the activity of only three enzymes: 1) The oxygen evolving complex of cyanobacteria that makes O2; 2) Nitrogenase, with its inhibition by O2 causing two billion years of oxygen level stasis; 3) Cellulose synthase of land plants, which caused mass deposition and burial of carbon, thus removing an oxygen sink and therefore increasing atmospheric O2. These three enzymes are endogenously produced by, and contained within, cells that have the capacity for exponential growth. The catalytic properties of these three enzymes paved the path of Earth's atmospheric oxygenation, requiring no help from Earth other than the provision of water, CO2, salts, colonizable habitats, and sunlight.

The radical impact of oxygen on prokaryotic evolution-enzyme inhibition first, uninhibited essential biosyntheses second, aerobic respiration third.

Molecular oxygen is a stable diradical. All O2-dependent enzymes employ a radical mechanism. Generated by cyanobacteria, O2 started accumulating on Earth 2.4 billion years ago. Its evolutionary impact is traditionally sought in respiration and energy yield. We mapped 365 O2-dependent enzymatic reactions of prokaryotes to phylogenies for the corresponding 792 protein families. The main physiological adaptations imparted by O2-dependent enzymes were not energy conservation, but novel organic substrate oxidations and O2-dependent, hence O2-tolerant, alternative pathways for O2-inhibited reactions. Oxygen-dependent enzymes evolved in ancestrally anaerobic pathways for essential cofactor biosynthesis including NAD+, pyridoxal, thiamine, ubiquinone, cobalamin, heme, and chlorophyll. These innovations allowed prokaryotes to synthesize essential cofactors in O2-containing environments, a prerequisite for the later emergence of aerobic respiratory chains.

Catalysts, autocatalysis and the origin of metabolism.

If life on Earth started out in geochemical environments like hydrothermal vents, then it started out from gasses like CO2, N2 and H2. Anaerobic autotrophs still live from these gasses today, and they still inhabit the Earth's crust. In the search for connections between abiotic processes in ancient geological systems and biotic processes in biological systems, it becomes evident that chemical activation (catalysis) of these gasses and a constant source of energy are key. The H2-CO2 redox reaction provides a constant source of energy and anabolic inputs, because the equilibrium lies on the side of reduced carbon compounds. Identifying geochemical catalysts that activate these gasses en route to nitrogenous organic compounds and small autocatalytic networks will be an important step towards understanding prebiotic chemistry that operates only on the basis of chemical energy, without input from solar radiation. So, if life arose in the dark depths of hydrothermal vents, then understanding reactions and catalysts that operate under such conditions is crucial for understanding origins.

Nitrogenase Inhibition Limited Oxygenation of Earth's Proterozoic Atmosphere.

Cyanobacteria produced the oxygen that began to accumulate on Earth 2.5 billion years ago, at the dawn of the Proterozoic Eon. By 2.4 billion years ago, the Great Oxidation Event (GOE) marked the onset of an atmosphere containing oxygen. The oxygen content of the atmosphere then remained low for almost 2 billion years. Why? Nitrogenase, the sole nitrogen-fixing enzyme on Earth, controls the entry of molecular nitrogen into the biosphere. Nitrogenase is inhibited in air containing more than 2% oxygen: the concentration of oxygen in the Proterozoic atmosphere. We propose that oxygen inhibition of nitrogenase limited Proterozoic global primary production. Oxygen levels increased when upright terrestrial plants isolated nitrogen fixation in soil from photosynthetic oxygen production in shoots and leaves.

Molecular Recognition: How Photosynthesis Anchors the Mobile Antenna.

True to its name, light-harvesting complex II (LHC II) harvests light energy for photosystem II (PS II). However, LHC II can stray, harvesting light energy for photosystem I (PS I) instead. Cryo-electron microscopy (cryo-EM) now shows how this mobile antenna becomes so attached to its new partner.

An Algal Greening of Land.

Photosynthetic eukaryotes arose ∼1.5 billion years ago by endosymbiosis with a cyanobacterium. Algae then evolved for a billion years before one lineage finally colonized land. Why the wait? The Chara braunii genome details a decisive step linking plant origins with Earth's history.

Oligomeric states in sodium ion-dependent regulation of cyanobacterial histidine kinase-2.

Two-component signal transduction systems (TCSs) consist of sensor histidine kinases and response regulators. TCSs mediate adaptation to environmental changes in bacteria, plants, fungi and protists. Histidine kinase 2 (Hik2) is a sensor histidine kinase found in all known cyanobacteria and as chloroplast sensor kinase in eukaryotic algae and plants. Sodium ions have been shown to inhibit the autophosphorylation activity of Hik2 that precedes phosphoryl transfer to response regulators, but the mechanism of inhibition has not been determined. We report on the mechanism of Hik2 activation and inactivation probed by chemical cross-linking and size exclusion chromatography together with direct visualisation of the kinase using negative-stain transmission electron microscopy of single particles. We show that the functional form of Hik2 is a higher-order oligomer such as a hexamer or octamer. Increased NaCl concentration converts the active hexamer into an inactive tetramer. The action of NaCl appears to be confined to the Hik2 kinase domain.

Why we need to know the structure of phosphorylated chloroplast light-harvesting complex II.

In oxygenic photosynthesis there are two 'light states' - adaptations of the photosynthetic apparatus to spectral composition that otherwise favours either photosystem I or photosystem II. In chloroplasts of green plants the transition to light state 2 depends on phosphorylation of apoproteins of a membrane-intrinsic antenna, the chlorophyll-a/b-binding, light-harvesting complex II (LHC II), and on the resulting redistribution of absorbed excitation energy from photosystem II to photosystem I. The transition to light state 1 reverses these events and requires a phospho-LHC II phosphatase. Current structures of LHC II reveal little about possible steric effects of phosphorylation. The surface-exposed N-terminal domain of an LHC II polypeptide contains its phosphorylation site and is disordered in its unphosphorylated form. A molecular recognition hypothesis proposes that state transitions are a consequence of movement of LHC II between binding sites on photosystems I and II. In state 1, LHC II forms part of the antenna of photosystem II. In state 2, a unique but as yet unidentified 3-D structure of phospho-LHC II may attach it instead to photosystem I. One possibility is that the LHC II N-terminus becomes ordered upon phosphorylation, adopting a local alpha-helical secondary structure that initiates changes in LHC II tertiary and quaternary structure that sever contact with photosystem II while securing contact with photosystem I. In order to understand redistribution of absorbed excitation energy in photosynthesis we need to know the structure of LHC II in its phosphorylated form, and in its complex with photosystem I.

The CoRR hypothesis for genes in organelles.

Chloroplasts and mitochondria perform energy transduction in photosynthesis and respiration. These processes can be described in physico-chemical terms with no obvious requirement for co-located genetic systems, separat from those of the rest of the cell. Accordingly, biochemists once tended to regard endosymbiosis as untestable evolutionary speculation. Lynn Sagan's seminal 1967 paper "On the Origin of Mitosing Cells" outlined the evolution of eukaryotic cells by endosymbiosis of prokaryotes. The endosymbiont hypothesis is consistent with presence of DNA in chloroplasts and mitochondria, but does not assign it a function. Biochemistry and molecular biology now show that Sagan's proposal has an explanatory reach far beyond that originally envisaged. Prokaryotic origins of photosynthetic and respiratory mechanisms are apparent in protein structural insights into energy coupling. Genome sequencing confirms the underlying, prokaryotic architecture of chloroplasts and mitochondria and illustrates the profound influence of the original mergers of their ancestors' genes and proteins with those of their host cells. Peter Mitchell's 1961 chemiosmotic hypothesis applied the concept of vectorial catalysis that underlies biological energy transduction and cell structure, function, and origins. Continuity of electrical charge separation and membrane sidedness requires compartments within compartments, together with intricate mechanisms for transport within and between them. I suggest that the reason for the persistence of distinct genetic systems within bioenergetic organelles is the selective advantage of subcellular co-location of specific genes with their gene products. Co-location for Redox Regulation - CoRR - provides for a dialogue between chemical reduction-oxidation and the action of genes encoding its protein catalysts. These genes and their protein products are in intimate contact, and cannot be isolated from each other without loss of an essential mechanism of adaptation of electron transport to change in the external environment.

Redox Tuning in Photosystem II.

In photosynthesis, oxygen is liberated from water, not from CO2; however, this model has been silent on why photosynthesis requires bicarbonate. Rutherford and colleagues solve this problem elegantly: bicarbonate tunes water-oxidising photosystem II to make onward electron transfer efficient; an absence of bicarbonate retunes, redirects, and safely shuts down energy flow.

A Proposal for Formation of Archaean Stromatolites before the Advent of Oxygenic Photosynthesis.

Stromatolites are solid, laminar structures of biological origin. Living examples are sparsely distributed and formed by cyanobacteria, which are oxygenic phototrophs. However, stromatolites were abundant between 3.4 and 2.4 Gyr, prior to the advent of cyanobacteria and oxygenic photosynthesis. Here I propose that many Archaean stromatolites were seeded at points of efflux of hydrogen sulfide from hydrothermal fields into shallow water, while their laminar composition arose from alternating modes of strictly anoxygenic photosynthetic metabolism. These changes were a redox regulatory response of gene expression to changing hydrogen sulfide concentration, which fluctuated with intermittent dilution by tidal action or by rainfall into surface waters. The proposed redox switch between modes of metabolism deposited sequential microbial mats. These mats gave rise to alternating carbonate sediments predicted to retain evidence of their origin in differing ratios of isotopes of carbon and sulfur and in organic content. The mats may have arisen either by replacement of microbial populations or by continuous lineages of protocyanobacteria in which a redox genetic switch selected between Types I and II photosynthetic reaction centers, and thus between photolithoautotrophic and photoorganoheterotrophic metabolism. In the latter case, and by 2.4 Gyr at the latest, a mutation had disabled the redox genetic switch to give simultaneous constitutive expression of both Types I and II reaction centers, and thus to the ability to extract electrons from manganese and then water. By this simple step, the first cyanobacterium had the dramatic advantage of emancipation from limiting supplies of inorganic electron donors, produced free molecular oxygen as a waste product, and initiated the Great Oxidation Event in Earth's history at the transition from the Archaean to the Paleoproterozoic.

Lokiarchaeon is hydrogen dependent.

The nature of the host that acquired the mitochondrion at the eukaryote origin is an important microbial evolutionary issue. Modern phylogenetics indicates that the host was an archaeon. The metagenome sequence of Candidatus Lokiarchaeon has identified it as being the closest relative of the host yet known. Here, we report comparative genomic evidence indicating that Lokiarchaeon is hydrogen dependent, as one theory for the eukaryote origin-the hydrogen hypothesis-predicts for the host lineage.

Why Have Organelles Retained Genomes?

Genes in mitochondria and chloroplasts are co-located with their gene products to permit regulation of trans-membrane electron transport at the energetic boundary of the cell.

A Two-Component Regulatory System in Transcriptional Control of Photosystem Stoichiometry: Redox-Dependent and Sodium Ion-Dependent Phosphoryl Transfer from Cyanobacterial Histidine Kinase Hik2 to Response Regulators Rre1 and RppA.

Two-component systems (TCSs) are ubiquitous signaling units found in prokaryotes. A TCS consists of a sensor histidine kinase and a response regulator protein as signal transducers. These regulatory systems mediate acclimation to various environmental changes by coupling environmental cues to gene expression. Hik2 is a sensor histidine kinase and its gene is found in all cyanobacteria. Hik2 is the homolog of Chloroplast Sensor Kinase (CSK), a protein involved in redox regulation of chloroplast gene expression during changes in light quality in plants and algae. Here we describe biochemical characterization of the signaling mechanism of Hik2 and its phosphotransferase activity. Results presented here indicate that Hik2 undergoes autophosphorylation on a conserved histidine residue, and becomes rapidly dephosphorylated by the action of response regulators Rre1 and RppA. We also show that the autophosphorylation of Hik2 is specifically inhibited by sodium ions.

Probing the nucleotide-binding activity of a redox sensor: two-component regulatory control in chloroplasts.

Two-component signal transduction systems mediate adaptation to environmental changes in bacteria, plants, fungi, and protists. Each two-component system consists of a sensor histidine kinase and a response regulator. Chloroplast sensor kinase (CSK) is a modified sensor histidine kinase found in chloroplasts-photosynthetic organelles of plants and algae. CSK regulates the transcription of chloroplast genes in response to changes in photosynthetic electron transport. In this study, the full-length and truncated forms of Arabidopsis CSK proteins were overexpressed and purified in order to characterise their kinase and redox sensing activities. Our results show that CSK contains a modified kinase catalytic domain that binds ATP with high affinity and forms a quinone adduct that may confer redox sensing activity.

Why chloroplasts and mitochondria retain their own genomes and genetic systems: Colocation for redox regulation of gene expression.

Chloroplasts and mitochondria are subcellular bioenergetic organelles with their own genomes and genetic systems. DNA replication and transmission to daughter organelles produces cytoplasmic inheritance of characters associated with primary events in photosynthesis and respiration. The prokaryotic ancestors of chloroplasts and mitochondria were endosymbionts whose genes became copied to the genomes of their cellular hosts. These copies gave rise to nuclear chromosomal genes that encode cytosolic proteins and precursor proteins that are synthesized in the cytosol for import into the organelle into which the endosymbiont evolved. What accounts for the retention of genes for the complete synthesis within chloroplasts and mitochondria of a tiny minority of their protein subunits? One hypothesis is that expression of genes for protein subunits of energy-transducing enzymes must respond to physical environmental change by means of a direct and unconditional regulatory control--control exerted by change in the redox state of the corresponding gene product. This hypothesis proposes that, to preserve function, an entire redox regulatory system has to be retained within its original membrane-bound compartment. Colocation of gene and gene product for redox regulation of gene expression (CoRR) is a hypothesis in agreement with the results of a variety of experiments designed to test it and which seem to have no other satisfactory explanation. Here, I review evidence relating to CoRR and discuss its development, conclusions, and implications. This overview also identifies predictions concerning the results of experiments that may yet prove the hypothesis to be incorrect.

Massively convergent evolution for ribosomal protein gene content in plastid and mitochondrial genomes.

Plastid and mitochondrial genomes have undergone parallel evolution to encode the same functional set of genes. These encode conserved protein components of the electron transport chain in their respective bioenergetic membranes and genes for the ribosomes that express them. This highly convergent aspect of organelle genome evolution is partly explained by the redox regulation hypothesis, which predicts a separate plastid or mitochondrial location for genes encoding bioenergetic membrane proteins of either photosynthesis or respiration. Here we show that convergence in organelle genome evolution is far stronger than previously recognized, because the same set of genes for ribosomal proteins is independently retained by both plastid and mitochondrial genomes. A hitherto unrecognized selective pressure retains genes for the same ribosomal proteins in both organelles. On the Escherichia coli ribosome assembly map, the retained proteins are implicated in 30S and 50S ribosomal subunit assembly and initial rRNA binding. We suggest that ribosomal assembly imposes functional constraints that govern the retention of ribosomal protein coding genes in organelles. These constraints are subordinate to redox regulation for electron transport chain components, which anchor the ribosome to the organelle genome in the first place. As organelle genomes undergo reduction, the rRNAs also become smaller. Below size thresholds of approximately 1,300 nucleotides (16S rRNA) and 2,100 nucleotides (26S rRNA), all ribosomal protein coding genes are lost from organelles, while electron transport chain components remain organelle encoded as long as the organelles use redox chemistry to generate a proton motive force.

Female and male gamete mitochondria are distinct and complementary in transcription, structure, and genome function.

Respiratory electron transport in mitochondria is coupled to ATP synthesis while generating mutagenic oxygen free radicals. Mitochondrial DNA mutation then accumulates with age, and may set a limit to the lifespan of individual, multicellular organisms. Why is this mutation not inherited? Here we demonstrate that female gametes-oocytes-have unusually small and simple mitochondria that are suppressed for DNA transcription, electron transport, and free radical production. By contrast, male gametes-sperm-and somatic cells of both sexes transcribe mitochondrial genes for respiratory electron carriers and produce oxygen free radicals. This germ-line division between mitochondria of sperm and egg is observed in both the vinegar fruitfly and the zebrafish-species spanning a major evolutionary divide within the animal kingdom. We interpret these findings as an evidence that oocyte mitochondria serve primarily as genetic templates, giving rise, irreversibly and in each new generation, to the familiar energy-transducing mitochondria of somatic cells and male gametes. Suppressed mitochondrial metabolism in the female germ line may therefore constitute a mechanism for increasing the fidelity of mitochondrial DNA inheritance.

Mitochondrial genome function and maternal inheritance.

The persistence of mtDNA to encode a small subset of mitochondrial proteins reflects the selective advantage of co-location of key respiratory chain subunit genes with their gene products. The disadvantage of this co-location is exposure of mtDNA to mutagenic ROS (reactive oxygen species), which are by-products of aerobic respiration. The resulting 'vicious circle' of mitochondrial mutation has been proposed to underlie aging and its associated degenerative diseases. Recent evidence is consistent with the hypothesis that oocyte mitochondria escape the aging process by acting as quiescent genetic templates, transcriptionally and bioenergetically repressed. Transmission of unexpressed mtDNA in the female germline is considered as a reason for the existence of separate sexes, i.e. male and female. Maternal inheritance then circumvents incremental accumulation of age-related disease in each new generation.