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.

The Ancestral Mitotic State: Closed Orthomitosis With Intranuclear Spindles in the Syncytial Last Eukaryotic Common Ancestor.

All eukaryotes have linear chromosomes that are distributed to daughter nuclei during mitotic division, but the ancestral state of nuclear division in the last eukaryotic common ancestor (LECA) is so far unresolved. To address this issue, we have employed ancestral state reconstructions for mitotic states that can be found across the eukaryotic tree concerning the intactness of the nuclear envelope during mitosis (open or closed), the position of spindles (intranuclear or extranuclear), and the symmetry of spindles being either axial (orthomitosis) or bilateral (pleuromitosis). The data indicate that the LECA possessed closed orthomitosis with intranuclear spindles. Our reconstruction is compatible with recent findings indicating a syncytial state of the LECA, because it decouples three main processes: chromosome division, chromosome partitioning, and cell division (cytokinesis). The possession of closed mitosis using intranuclear spindles adds to the number of cellular traits that can now be attributed to LECA, providing insights into the lifestyle of this otherwise elusive biological entity at the origin of eukaryotic cells. Closed mitosis in a syncytial eukaryotic common ancestor would buffer mutations arising at the origin of mitotic division by allowing nuclei with viable chromosome sets to complement defective nuclei via mRNA in the cytosol.

Realistic Gene Transfer to Gene Duplication Ratios Identify Different Roots in the Bacterial Phylogeny Using a Tree Reconciliation Method.

The rooting of phylogenetic trees permits important inferences about ancestral states and the polarity of evolutionary events. Recently, methods that reconcile discordance between gene-trees and species-trees-tree reconciliation methods-are becoming increasingly popular for rooting species trees. Rooting via reconciliation requires values for a particular parameter, the gene transfer to gene duplication ratio (T:D), which in current practice is estimated on the fly from discordances observed in the trees. To date, the accuracy of T:D estimates obtained by reconciliation analyses has not been compared to T:D estimates obtained by independent means, hence the effect of T:D upon inferences of species tree roots is altogether unexplored. Here we investigated the issue in detail by performing tree reconciliations of more than 10,000 gene trees under a variety of T:D ratios for two phylogenetic cases: a bacterial (prokaryotic) tree with 265 species and a fungal-metazoan (eukaryotic) tree with 31 species. We show that the T:D ratios automatically estimated by a current tree reconciliation method, ALE, generate virtually identical T:D ratios across bacterial genes and fungal-metazoan genes. The T:D ratios estimated by ALE differ 10- to 100-fold from robust, ALE-independent estimates from real data. More important is our finding that the root inferences using ALE in both datasets are strongly dependent upon T:D. Using more realistic T:D ratios, the number of roots inferred by ALE consistently increases and, in some cases, clearly incorrect roots are inferred. Furthermore, our analyses reveal that gene duplications have a far greater impact on ALE's preferences for phylogenetic root placement than gene transfers or gene losses do. Overall, we show that obtaining reliable species tree roots with ALE is only possible when gene duplications are abundant in the data and the number of falsely inferred gene duplications is low. Finding a sufficient sample of true gene duplications for rooting species trees critically depends on the T:D ratios used in the analyses. T:D ratios, while being important parameters of genome evolution in their own right, affect the root inferences with tree reconciliations to an unanticipated degree.

Ancestral State Reconstructions Trace Mitochondria But Not Phagocytosis to the Last Eukaryotic Common Ancestor.

Two main theories have been put forward to explain the origin of mitochondria in eukaryotes: phagotrophic engulfment (undigested food) and microbial symbiosis (physiological interactions). The two theories generate mutually exclusive predictions about the order in which mitochondria and phagocytosis arose. To discriminate the alternatives, we have employed ancestral state reconstructions (ASR) for phagocytosis as a trait, phagotrophy as a feeding habit, the presence of mitochondria, the presence of plastids, and the multinucleated organization across major eukaryotic lineages. To mitigate the bias introduced by assuming a particular eukaryotic phylogeny, we reconstructed the appearance of these traits across 1789 different rooted gene trees, each having species from opisthokonts, mycetozoa, hacrobia, excavate, archeplastida, and Stramenopiles, Alveolates and Rhizaria. The trees reflect conflicting relationships and different positions of the root. We employed a novel phylogenomic test that summarizes ASR across trees which reconstructs a last eukaryotic common ancestor that possessed mitochondria, was multinucleated, lacked plastids, and was non-phagotrophic as well as non-phagocytic. This indicates that both phagocytosis and phagotrophy arose subsequent to the origin of mitochondria, consistent with findings from comparative physiology. Furthermore, our ASRs uncovered multiple origins of phagocytosis and of phagotrophy across eukaryotes, indicating that, like wings in animals, these traits are useful but neither ancestral nor homologous across groups. The data indicate that mitochondria preceded the origin of phagocytosis, such that phagocytosis cannot have been the mechanism by which mitochondria were acquired.

Evidence for a Syncytial Origin of Eukaryotes from Ancestral State Reconstruction.

Modern accounts of eukaryogenesis entail an endosymbiotic encounter between an archaeal host and a proteobacterial endosymbiont, with subsequent evolution giving rise to a unicell possessing a single nucleus and mitochondria. The mononucleate state of the last eukaryotic common ancestor (LECA) is seldom, if ever, questioned, even though cells harboring multiple (syncytia, coenocytes, and polykaryons) are surprisingly common across eukaryotic supergroups. Here, we present a survey of multinucleated forms. Ancestral character state reconstruction for representatives of 106 eukaryotic taxa using 16 different possible roots and supergroup sister relationships, indicate that LECA, in addition to being mitochondriate, sexual, and meiotic, was multinucleate. LECA exhibited closed mitosis, which is the rule for modern syncytial forms, shedding light on the mechanics of its chromosome segregation. A simple mathematical model shows that within LECA's multinucleate cytosol, relationships among mitochondria and nuclei were neither one-to-one, nor one-to-many, but many-to-many, placing mitonuclear interactions and cytonuclear compatibility at the evolutionary base of eukaryotic cell origin. Within a syncytium, individual nuclei and individual mitochondria function as the initial lower-level evolutionary units of selection, as opposed to individual cells, during eukaryogenesis. Nuclei within a syncytium rescue each other's lethal mutations, thereby postponing selection for viable nuclei and cytonuclear compatibility to the generation of spores, buffering transitional bottlenecks at eukaryogenesis. The prokaryote-to-eukaryote transition is traditionally thought to have left no intermediates, yet if eukaryogenesis proceeded via a syncytial common ancestor, intermediate forms have persisted to the present throughout the eukaryotic tree as syncytia but have so far gone unrecognized.