Early Eukaryotes Research Articles

A palaeoecological model for the late Mesoproterozoic – early Neoproterozoic Atar/El Mreïti Group, Taoudeni Basin, Mauritania, northwestern Africa

Reconstructing the spatial distribution of early eukaryotes in palaeoenvironments through Proterozoic sedimentary basins provides important information about their palaeoecology and taphonomic conditions. Here, we combine the geological context and a reconstruction of palaeoenvironmental redox conditions (using iron speciation) with quantitative analysis of microfossil assemblages (eukaryotes and incertae sedis), to provide the first palaeoecological model for the Atar/El Mreïti Group of the Taoudeni Basin. Our model suggests that in the late Mesoproterozoic – early Neoproterozoic, the availability of both molecular oxygen and nutrients controlled eukaryotic diversity, higher in oxic shallow marginal marine environments, while coccoidal colonies and benthic microbial mats dominated respectively in anoxic iron-rich and euxinic waters during marine highstands or away from shore where eukaryotes are lower or absent.

Organelle acidification: an ancient cellular leak detector

Intracellular membrane-bounded organelles of eukaryotic cells transiently contact the extracellular environment during endocytosis and secretion. Such contacts must be precisely timed to prevent leakage of cargo. I argue that early eukaryotes evolved organelle acidification as a way to detect and prevent leakage.

P 011 - Mycosporine-like amino acid activation of the Keap1-Nrf2 pathway

Oxidative stress is a contributing factor in the progression of numerous pathological conditions including neurodegeneration, cancer and ageing. The Keap1-Nrf2 pathway is a master regulator of oxidative stress: reactive oxygen species are sensed by Keap1 to release Nrf2 for transcriptional activation of protective genes controlled by the nuclear antioxidant response element. We have identified homologs of Nrf2 in early eukaryotes and used virtual screens to predict natural products able to activate Nrf2 by competitive inhibition of Keap1-Nrf2 binding. Mycosporine-like amino acids (MAAs) are water-soluble metabolites produced by taxonomically diverse organisms, particularly marine algae and seaweeds. These compounds absorb UV radiation – thus acting as “primary sunscreens” – and reported also to protect against oxidative damage. We have tested the MAAs, porphyra-334, shinorine, and palythine for in-vitro antioxidant activity using the DPPH free-radical quenching assay and report also their ability to activate the Keap1-Nrf2 pathway using fluorescence polarization and thermal shift assays to detect Keap1 receptor antagonism. Our results demonstrate that shinorine and porphyra-334 are competitive inhibitors of Keap1-Nrf2 binding having potential to protect against UVR via sunscreen absorption and transcriptional activation of endogenous defenses against UV-induced oxidative damage.

Contributions of U-Th-Pb dating on the diagenesis and sediment sources of the lower group (BI) of the Mbuji-Mayi Supergroup (Democratic Republic of Congo)

In this paper, we present new age constraints for the lower part of the Meso-Neoproterozoic sedimentary Mbuji-Mayi Supergroup (Democratic Republic of Congo, DRC). This Supergroup preserves a large diversity of organic-walled microfossils, evidencing the diversification of early eukaryotes for the first time in Central Africa. We use different methods such as in situ U-Pb geochronology by LA-ICP-MS and U-Th-Pb chemical datings by Electron Microprobe on diagenetic and detrital minerals such as xenotimes, monazites and zircons. We attempt to better constrain the provenance of the Mbuji-Mayi sediments and the minimum age of the Mbuji-Mayi Supergroup to constrain the age of the microfossils. Results with LA-ICP-MS and EMP provide new ages between 1030 and 1065Ma for the diagenesis of the lower part of the sedimentary sequence. These results are consistent with data on biostratigraphy supporting the occurrence of worldwide changes at the Mesoproterozoic/Neoproterozoic boundary.

Transitions between marine and freshwater environments provide new clues about the origins of multicellular plants and algae.

Marine-freshwater and freshwater-marine transitions have been key events in the evolution of life, and most major groups of organisms have independently undergone such events at least once in their history. Here, we first compile an inventory of bidirectional freshwater and marine transitions in multicellular photosynthetic eukaryotes. While green and red algae have mastered multiple transitions in both directions, brown algae have colonized freshwater on a maximum of six known occasions, and angiosperms have made the transition to marine environments only two or three times. Next, we review the early evolutionary events leading to the colonization of current habitats. It is commonly assumed that the conquest of land proceeded in a sequence from marine to freshwater habitats. However, recent evidence suggests that early photosynthetic eukaryotes may have arisen in subaerial or freshwater environments and only later colonized marine environments as hypersaline oceans were diluted to the contemporary level. Although this hypothesis remains speculative, it is important to keep these alternative scenarios in mind when interpreting the current habitat distribution of plants and algae. Finally, we discuss the roles of structural and functional adaptations of the cell wall, reactive oxygen species scavengers, osmoregulation, and reproduction. These are central for acclimatization to freshwater or to marine environments. We observe that successful transitions appear to have occurred more frequently in morphologically simple forms and conclude that, in addition to physiological studies of euryhaline species, comparative studies of closely related species fully adapted to one or the other environment are necessary to better understand the adaptive processes.

Erratum to: Mitochondrial clock: moderating evolution of early eukaryotes in light of the Proterozoic oceans

Due to a misunderstanding caused by the author, Dr. Shamik Dasgupta, the name of the co-author, Dr. Jiasong Fang (who has been acknowledged in the manuscript), was omitted in the original publication. The correct authorship of this article is below.

Nonsense-mediated mRNA decay in Tetrahymena is EJC independent and requires a protozoa-specific nuclease.

Nonsense-mediated mRNA decay (NMD) is essential for removing premature termination codon-containing transcripts from cells. Studying the NMD pathway in model organisms can help to elucidate the NMD mechanism in humans and improve our understanding of how this biologically important process has evolved. Ciliates are among the earliest branching eukaryotes; their NMD mechanism is poorly understood and may be primordial. We demonstrate that highly conserved Upf proteins (Upf1a, Upf2 and Upf3) are involved in the NMD pathway of the ciliate, Tetrahymena thermophila. We further show that a novel protozoa-specific nuclease, Smg6L, is responsible for destroying many NMD-targeted transcripts. Transcriptome-wide identification and characterization of NMD-targeted transcripts in vegetative Tetrahymena cells showed that many have exon–exon junctions downstream of the termination codon. However, Tetrahymena may lack a functional exon junction complex (EJC), and the Tetrahymena ortholog of an EJC core component, Mago nashi (Mag1), is dispensable for NMD. Therefore, NMD is EJC independent in this early branching eukaryote.

The Major Protein Arginine Methyltransferase in Trypanosoma brucei Functions as an Enzyme-Prozyme Complex

Prozymes are catalytically inactive enzyme paralogs that dramatically stimulate the function of weakly active enzymes through complex formation. The two prozymes described to date reside in the polyamine biosynthesis pathway of the human parasite Trypanosoma brucei, an early branching eukaryote that lacks transcriptional regulation and regulates its proteome through posttranscriptional and posttranslational means. Arginine methylation is a common posttranslational modification in eukaryotes catalyzed by protein arginine methyltransferases (PRMTs) that are typically thought to function as homodimers. We demonstrate that a major T. brucei PRMT, TbPRMT1, functions as a heterotetrameric enzyme-prozyme pair. The inactive PRMT paralog, TbPRMT1PRO, is essential for catalytic activity of the TbPRMT1ENZ subunit. Mutational analysis definitively demonstrates that TbPRMT1ENZ is the cofactor-binding subunit and carries all catalytic activity of the complex. Our results are the first demonstration of an obligate heteromeric PRMT, and they suggest that enzyme-prozyme organization is expanded in trypanosomes as a posttranslational means of enzyme regulation.

Proximity Interactions among Basal Body Components in Trypanosoma brucei Identify Novel Regulators of Basal Body Biogenesis and Inheritance.

ABSTRACTThe basal body shares similar architecture with centrioles in animals and is involved in nucleating flagellar axonemal microtubules in flagellated eukaryotes. The early-branching Trypanosoma brucei possesses a motile flagellum nucleated from the basal body that consists of a mature basal body and an adjacent pro-basal body. Little is known about the basal body proteome and its roles in basal body biogenesis and flagellar axoneme assembly in T. brucei. Here, we report the identification of 14 conserved centriole/basal body protein homologs and 25 trypanosome-specific basal body proteins. These proteins localize to distinct subdomains of the basal body, and several of them form a ring-like structure surrounding the basal body barrel. Functional characterization of representative basal body proteins revealed distinct roles in basal body duplication/separation and flagellar axoneme assembly. Overall, this work identified novel proteins required for basal body duplication and separation and uncovered new functions of conserved basal body proteins in basal body duplication and separation, highlighting an unusual mechanism of basal body biogenesis and inheritance in this early divergent eukaryote.

The MAP kinase MAPKLK1 is essential to Trypanosoma brucei proliferation and regulates proteins involved in mRNA metabolism

Protein phosphorylation and dephosphorylation events regulate many cellular processes. The identification of all phosphorylation sites and their association to a respective protein kinase or phosphatase is a challenging and crucial step to have a deeper understanding of the effects of signaling networks on cells. Pathogenic trypanosomatids have a large number of protein kinases and phosphatases in comparison to other organisms, which reinforces the relevance of the phosphorylation process in these early eukaryotes, nevertheless little is known about protein phosphorylation in these protozoa. In this context, the role of a MAP kinase-like kinase (MAPKLK1), observed to be essential to proliferation of procyclic Trypanosoma brucei, was studied. After silencing MAPKLK1 expression by RNAi, the cells were evaluated by SILAC MS-based proteomics and RNA-Seq. We identified 1756 phosphorylation sites of which 384 were not previously described in T. brucei. Despite being essential, few modulations were observed at the phosphorylation patterns and gene expression levels of MAPKLK1 knockdown. These indirect targets and potential substrates of MAPKLK1 are related to key cellular processes enriched to mRNA processing and stability control. SignificanceThe field of cell signaling is a promising topic of study for trypanosomatids, since little is known about this topic and the gene expression regulation occurs at post-transcriptional level. In this sense, the present work increases the knowledge on protein phosphorylation process in Trypanosoma brucei. We depleted one MAP kinase (MAPKLK1) of T. brucei and evaluated the effects on the cell. We showed that MAPKLK1 is essential to the cell, while few modulations on phosphoproteome, proteome and transcriptome are observed with its depletion. Although in low number, the changes in phosphoproteome were significant, presenting possible substrate candidates of MAPKLK1 and indirect targets related to mRNA processing and stability control, metabolic pathways, among others. This result provides insights in the phosphorylation network of T. brucei, a model organism that impacts human and animal health.

What can we infer about the origin of sex in early eukaryotes?

Current analysis shows that the last eukaryotic common ancestor (LECA) was capable of full meiotic sex. The original eukaryotic life cycle can probably be described as clonal, interrupted by episodic sex triggered by external or internal stressors. The cycle could have started in a highly flexible form, with the interruption of either diploid or haploid clonal growth determined by stress signals only. Eukaryotic sex most likely evolved in response to a high mutation rate, arising from the uptake of the endosymbiont, as this (proto) mitochondrion generated internal reactive oxygen species. This is consistent with the likely development of full meiotic sex from a diverse set of existing archaeal (the host of the endosymbiont) repair and signalling mechanisms. Meiotic sex could thus have been one of the fruits of symbiogenesis at the basis of eukaryotic origins: a product of the merger by which eukaryotic cells arose. Symbiogenesis also explains the large-scale migration of organellar DNA to the nucleus. I also discuss aspects of uniparental mitochondrial inheritance and mitonuclear interactions in the light of the previous analysis.This article is part of the themed issue 'Weird sex: the underappreciated diversity of sexual reproduction'.

A widely employed germ cell marker is an ancient disordered protein with reproductive functions in diverse eukaryotes.

The advent of sexual reproduction and the evolution of a dedicated germline in multicellular organisms are critical landmarks in eukaryotic evolution. We report an ancient family of GCNA (germ cell nuclear antigen) proteins that arose in the earliest eukaryotes, and feature a rapidly evolving intrinsically disordered region (IDR). Phylogenetic analysis reveals that GCNA proteins emerged before the major eukaryotic lineages diverged; GCNA predates the origin of a dedicated germline by a billion years. Gcna gene expression is enriched in reproductive cells across eukarya - either just prior to or during meiosis in single-celled eukaryotes, and in stem cells and germ cells of diverse multicellular animals. Studies of Gcna-mutant C. elegans and mice indicate that GCNA has functioned in reproduction for at least 600 million years. Homology to IDR-containing proteins implicated in DNA damage repair suggests that GCNA proteins may protect the genomic integrity of cells carrying a heritable genome.

Mitochondrial clock: moderating evolution of early eukaryotes in light of the Proterozoic oceans

AbstractEvolution of early eukaryotes in the primitive Earth relied heavily on the origin and evolution of mitochondria. Understanding the structure and origin of mitochondria has a germane relation to understanding origin and evolution of eukaryotes. In light of the extreme conditions and the then existing Proterozoic ocean chemistry, eukaryotic cells developed adaptive adjustments for energy management. Apart from mitochondria, more reduced homologues like hydrogenosomes and mitosomes facilitated the metabolic activities of such eukaryotic life. In this short review, I highlight the importance of mitochondria in pushing eukaryotes to the peak of the evolutionary pyramid. Our knowledge has expanded but studying recent eukaryotic extremophiles and mitochondrial genomics in more details will enable us to estimate the position of the mitochondrial clock, understand its role better, and possibly find new eukaryotic lineages.

A diverse and exquisitely preserved organic-walled microfossil assemblage from the Meso–Neoproterozoic Mbuji-Mayi Supergroup (Democratic Republic of Congo) and implications for Proterozoic biostratigraphy

A well preserved and diversified microfossil assemblage is reported from the Meso–Neoproterozoic Mbuji-Mayi Supergroup in the Kasai oriental Province, central part of Democratic Republic of Congo. A total of 49 taxa belonging to 27 genera were identified, including 11 species of unambiguous eukaryotes, 10 species of possible eukaryotes or prokaryotes and 28 species of probable bacteria. This assemblage is more diverse than previously reported but includes taxa reported in coeval worldwide assemblages. It is characterized by abundant sphaeromorphs, filamentous colonial aggregates and filamentous forms, as well as a relatively low diversity of acanthomorphs including the Late Mesoproterozoic and Early Neoproterozoic index fossil – Trachyhystrichosphaera aimika – reported for the first time in Central Africa. This species co-occurs with other taxa also reported for the first time in Africa: Trachyhystrichosphaera botula, Jacutianema solubila, cf. Tappania sp., Valeria elongata and numerous other taxa. Correlation with other geochronologically constrained successions that contain Trachyhystrichosphaera confirms T. aimika as promising index fossil to define the Late Mesoproterozoic–Early Neoproterozoic interval. The available biostratigraphic data enable to suggest a minimum Tonian age for the Mbuji-Mayi Supergroup. This age is consistent with the published and new geochronological data. Comparison with worldwide Proterozoic assemblages permits to define microfossil assemblages useful for biostratigraphy. This study significantly improves our understanding of the diversity of the Late Mesoproterozoic–Early Neoproterozoic biosphere, and in particular the diversification of early eukaryotes, preserved in the Democratic Republic of Congo rock record and more broadly in Africa where micropaleontological investigations are sparse.

Essential Functions of the N‐terminal Domain of Tim17 in Trypanosoma brucei

The majority of proteins essential for mitochondrial functions are encoded in the nuclear DNA and subsequently imported from the cytosol by translocases of the mitochondrial outer and inner membranes, termed TOM and TIM, respectively. These translocases are multimeric protein complexes, which are overall conserved from fungi to human. However, mitochondrial protein import machinery in trypanosomatids, a group of ancient and early branching unicellular eukaryotes that causes a fatal human disease African trypanosomiasis, is significantly different. As a part of this machinery, we previously characterized Tim17 from Trypanosoma brucei. TbTim17 is critical for mitochondrial protein import and thus essential for parasite survival. Here we further investigate the function of the divergent N‐terminal domain of this protein. We found that deletion of the first 20 or 30 amino acid residues (Δ20‐ and Δ30‐) of TbTim17 is detrimental for its function. These mutant proteins neither complement the growth defect of TbTim17 RNAi cells nor the import defect of other nuclear encoded proteins into TbTim17 knockdown mitochondria. However, knock‐in of the full‐length protein (FL‐TbTim17) completely restored these functions in RNAi cells. Further analysis showed that both of these mutants of TbTim17 were properly targeted to mitochondria, however, Δ20‐TbTim17 was not membrane integrated and Δ30‐mutant, though integrated into the mitochondrial membrane was sensitive to protease digestion of the isolated mitochondria, indicating that the N‐terminal region is necessary for proper integration of TbTim17 into the mitochondrial inner membrane. Analysis of the protein complexes in the wild type and mutant mitochondria by Blue‐native gel electrophoresis revealed that none of these mutants were assembled into the TbTim17 protein complex. Co‐immunoprecipitation analysis showed that unlike the wild type TbTim17, the mutant proteins were not associated with the endogenous TbTim17 as well as its interacting partner TbTim62, a novel trypanosome specific Tim. Together, these results show that the N‐terminal domain of TbTim17 is critical for its interaction with TbTim62 and required for the formation of a functional TbTim17 protein complex. To elucidate the assembly of the TIM complex in T. brucei, analysis is under progress to identify essential amino acids residues of this region by site‐directed mutagenesis.Support or Funding InformationNIH Grants 2SC1GM081146‐05, T32HL007727, and G12RR003032

Novel insights on ENTH domain-containing proteins in apicomplexan parasites.

The phylum Apicomplexa includes a large group of early branching eukaryotes having significant medical and economical importance. The molecular machinery responsible for protein trafficking is poorly understood in these apicomplexans. One of the most important proteins involved in clathrin-mediated protein trafficking is Epsin, which contains ENTH domain, a conserved domain crucial for membrane bending leading to vesicle formation. We undertook homology searching and phylogenetic analyses to produce a rigorously annotated set of Epsin homologs retrieved from diverse apicomplexan genomes. Genomic and phylogenetic comparisons revealed that apicomplexans contain unusual Epsin homologs that are distinct from those observed in mammals and yeast. Although there are four Epsin genes in mammalian system and five in the yeast genome, apicomplexan parasites consist only a single Epsin gene. The apicomplexan Epsin contains the conserved ENTH domain consisting of phosphoinositide (PtdIns)-binding sites which indicate about their functional significance in the formation of vesicles; however, the absence of ubiquitin-interacting motif (UIM) suggests a possible different mechanism for protein trafficking. The existence of dileucine motif in Plasmodium, Cryptosporidum parvum and Eimeria tenella Epsins might solve their functionality while lacking a lot of conserved motifs as this motif is known to interact with different adaptor protein complexes (AP1, AP2 and AP3). Other Epsin homologs are also shown to have different peptide motifs reported for possible interaction with α-ear appendage, γ-ear appendage and EH domain present in different adaptors. Bioinformatic and phylogenetic analyses suggest that the apicomplexan Epsins have unusual functionality from that of the mammalian Epsins. This detailed study may greatly facilitate future molecular cell biological investigation for the role of Epsins in these parasites.

Extremely low oxygen concentration in mid-Proterozoic shallow seawaters

The mid-Proterozoic (1.8–0.8Ga) witnessed the first appearance but unusually low diversification of eukaryotes. The stagnant biotic evolution during this billion-year-long period (commonly referred to as the “Boring Billion”) was arguably ascribed to low oxygen levels in atmosphere and ocean. However, evidence supporting low oxygen in shallow-marine environments where early eukaryotes first evolved is generally lacking or insufficient. Here we report Rare Earth Element and yttrium (REE+Y) data, particularly cerium (Ce) anomalies, from a suite of mid-Proterozoic sedimentary rocks of the North China platform. The new data from North China, in combination with available Ce anomaly data from other Proterozoic successions, demonstrate that during mid-Proterozoic, negative Ce anomalies did not occur until ∼1.54Ga and after ∼1.54Ga, only episodic negative Ce anomalies were present in shallow-water carbonates. Trace element enrichments (UEF, VEF, and MoEF) remained at the average continental crust level before ∼1.54Ga but showed a sudden increase at ∼1.54Ga. The data suggest that oxygen concentration in shallow-marine environments of the mid-Proterozoic ocean was extremely low, probably <0.2μM prior to ∼1.54Ga (based on minimal oxygen concentration requirement for Ce(III) oxidation) and fluctuating around 0.2μM afterwards. The low oxygen concentration (∼0.2μM) in shallow waters of the mid-Proterozoic ocean accounts for only ∼0.1% of the modern surface ocean oxygen level (∼280μM) and may help explain the evolutionary stasis of eukaryotes during the mid-Proterozoic.

Sex, health and behaviour: Sexual reproduction might have emerged to provide better immunity against pathogens and further evolved to select for behaviour.

Understanding the emergence of sexual reproduction has been one of nature's long‐standing mysteries that even now is only partially understood. There is a clear distinction between the reason sex evolved in the first place in early eukaryotes—and even before that in prokaryotes—and why it has been so stubbornly maintained in virtually all higher species. After all, asexual reproduction is superior in a strict Darwinian sense, since it yields more offspring per unit time without having to waste energy on finding mates. On the question of the origin of sex, there is little progress, with plenty of competing theories but scant evidence. But a clearer picture has emerged of why sex has resisted the pull towards asexual reproduction and what has driven its evolution. According to Manfred Milinski, Director of the Max Planck Institute for Evolutionary Biology in Plon, Germany, the bottom line is that the constant threat from parasites and pathogens has maintained the need for sex. “In a world without infectious diseases, asexual reproduction would prevail,” he explained. “However, with asexual reproduction, there is no evolutionary improvement, only genetic degeneration through fatal mutation accumulation. Infectious diseases are indeed drivers of sexual reproduction and thus evolution. So if we removed all infectious diseases, we would return to asexual reproduction and disappear as a species after about 200 generations because of inevitable mutation accumulation. Infectious diseases help us survive.” One big advantage of infectious pathogens compared with their larger and longer lived hosts, including humans, is that they evolve much faster through mutation. The only recourse is a highly diverse immune system capable of quickly adapting to new and mutating pathogens, according to Daniel Davis, Director of Research at the Manchester Collaborative Centre for Inflammation Research in the UK. “The genes that vary the most between individuals are those to do …

A loopy view of telomere evolution.

About a decade ago, I proposed that t-loops, the lariat structures adopted by many eukaryotic telomeres, could explain how the transition from circular to linear chromosomes was successfully negotiated by early eukaryotes. Here I reconsider this loopy hypothesis in the context of the idea that eukaryotes evolved through a period of genome invasion by Group II introns.

Melatonin: an ancient molecule that makes oxygen metabolically tolerable.

Melatonin is remarkably functionally diverse with actions as a free radical scavenger and antioxidant, circadian rhythm regulator, anti-inflammatory and immunoregulating molecule, and as an oncostatic agent. We hypothesize that the initial and primary function of melatonin in photosynthetic cyanobacteria, which appeared on Earth 3.5-3.2 billion years ago, was as an antioxidant. The evolution of melatonin as an antioxidant by this organism was necessary as photosynthesis is associated with the generation of toxic-free radicals. The other secondary functions of melatonin came about much later in evolution. We also surmise that mitochondria and chloroplasts may be primary sites of melatonin synthesis in all eukaryotic cells that possess these organelles. This prediction is made on the basis that mitochondria and chloroplasts of eukaryotes developed from purple nonsulfur bacteria (which also produce melatonin) and cyanobacteria when they were engulfed by early eukaryotes. Thus, we speculate that the melatonin-synthesizing actions of the engulfed bacteria were retained when these organelles became mitochondria and chloroplasts, respectively. That mitochondria are likely sites of melatonin formation is supported by the observation that this organelle contains high levels of melatonin that are not impacted by blood melatonin concentrations. Melatonin has a remarkable array of means by which it thwarts oxidative damage. It, as well as its metabolites, is differentially effective in scavenging a variety of reactive oxygen and reactive nitrogen species. Moreover, melatonin and its metabolites modulate a large number of antioxidative and pro-oxidative enzymes, leading to a reduction in oxidative damage. The actions of melatonin on radical metabolizing/producing enzymes may be mediated by the Keap1-Nrf2-ARE pathway. Beyond its direct free radical scavenging and indirect antioxidant effects, melatonin has a variety of physiological and metabolic advantages that may enhance its ability to limit oxidative stress.