Paramecium Genetics, Genomics, and Evolution.

The ciliate genus Paramecium served as one of the first model systems in microbial eukaryotic genetics, contributing much to the early understanding of phenomena as diverse as genome rearrangement, cryptic speciation, cytoplasmic inheritance, and endosymbiosis, as well as more recently to the evolution of mating types, introns, and roles of small RNAs in DNA processing. Substantial progress has recently been made in the area of comparative and population genomics. Paramecium species combine some of the lowest known mutation rates with some of the largest known effective populations, along with likely very high recombination rates, thereby harboring a population-genetic environment that promotes an exceptionally efficient capacity for selection. As a consequence, the genomes are extraordinarily streamlined, with very small intergenic regions combined with small numbers of tiny introns. The subject of the bulk of Paramecium research, the ancient Paramecium aurelia species complex, is descended from two whole-genome duplication events that retain high degrees of synteny, thereby providing an exceptional platform for studying the fates of duplicate genes. Despite having a common ancestor dating to several hundred million years ago, the known descendant species are morphologically indistinguishable, raising significant questions about the common view that gene duplications lead to the origins of evolutionary novelties.

Assembling a Hippo: the evolutionary emergence of an animal developmental signaling pathway.

Decades of work in developmental genetics has given us a deep mechanistic understanding of the fundamental signaling pathways underlying animal development. However, little is known about how these pathways emerged and changed over evolutionary time. Here, we review our current understanding of the evolutionary emergence of the Hippo pathway, a conserved signaling pathway that regulates tissue size in animals. This pathway has deep evolutionary roots, emerging piece by piece in the unicellular ancestors of animals, with a complete core pathway predating the origin of animals. Recent functional studies in close unicellular relatives of animals and early-branching animals suggest an ancestral function of the Hippo pathway in cytoskeletal regulation, which was subsequently co-opted to regulate proliferation and animal tissue size.

Mitochondrial genomes revisited: why do different lineages retain different genes?

The mitochondria contain their own genome derived from an alphaproteobacterial endosymbiont. From thousands of protein-coding genes originally encoded by their ancestor, only between 1 and about 70 are encoded on extant mitochondrial genomes (mitogenomes). Thanks to a dramatically increasing number of sequenced and annotated mitogenomes a coherent picture of why some genes were lost, or relocated to the nucleus, is emerging. In this review, we describe the characteristics of mitochondria-to-nucleus gene transfer and the resulting varied content of mitogenomes across eukaryotes. We introduce a 'burst-upon-drift' model to best explain nuclear-mitochondrial population genetics with flares of transfer due to genetic drift.

Archaeal actins and the origin of a multi-functional cytoskeleton.

Actin and actin-like proteins form filamentous polymers that carry out important cellular functions in all domains of life. In this review, we sketch a map of the function and regulation of actin-like proteins across bacteria, archaea, and eukarya, marking some of the terra incognita that remain in this landscape. We focus particular attention on archaea because mapping the structure and function of cytoskeletal systems across this domain promises to help us understand the evolutionary relationship between the (mostly) mono-functional actin-like filaments found in bacteria and the multi-functional actin cytoskeletons that characterize eukaryotic cells.

The cell morphological diversity of Saccharomycotina yeasts.

The ∼1 200 known species in subphylum Saccharomycotina are a highly diverse clade of unicellular fungi. During its lifecycle, a typical yeast exhibits multiple cell types with various morphologies; these morphologies vary across Saccharomycotina species. Here, we synthesize the evolutionary dimensions of variation in cellular morphology of yeasts across the subphylum, focusing on variation in cell shape, cell size, type of budding, and filament production. Examination of 332 representative species across the subphylum revealed that the most common budding cell shapes are ovoid, spherical, and ellipsoidal, and that their average length and width is 5.6 µm and 3.6 µm, respectively. 58.4% of yeast species examined can produce filamentous cells, and 87.3% of species reproduce asexually by multilateral budding, which does not require utilization of cell polarity for mitosis. Interestingly, ∼1.8% of species examined have not been observed to produce budding cells, but rather only produce filaments of septate hyphae and/or pseudohyphae. 76.9% of yeast species examined have sexual cycle descriptions, with most producing one to four ascospores that are most commonly hat-shaped (37.4%). Systematic description of yeast cellular morphological diversity and reconstruction of its evolution promises to enrich our understanding of the evolutionary cell biology of this major fungal lineage.

Comparative genomic analysis illustrates evolutionary dynamics of multisubunit tethering complexes across green algal diversity.

The chlorophyte algae are a dominant group of photosynthetic eukaryotes. Although many are photoautotrophs, there are also mixotrophs, heterotrophs, and even parasites. The physical characteristics of green algae are also highly diverse, varying greatly in size, shape, and habitat. Given this morphological and trophic diversity, we postulated that diversity may also exist in the protein components controlling intracellular movement of material by vesicular transport. One such set is the multisubunit tethering complexes (MTCs)-components regulating cargo delivery. As they span endomembrane organelles and are well-conserved across eukaryotes, MTCs should be a good proxy for assessing the evolutionary dynamics across the diversity of Chlorophyta. Our results reveal that while green algae carry a generally conserved and unduplicated complement of MTCs, some intriguing variation exists. Notably, we identified incomplete sets of TRAPPII, exocyst, and HOPS/CORVET components in all Mamiellophyceae, and what is more, not a single subunit of Dsl1 was found in Cymbomonas tetramitiformis. As the absence of Dsl1 has been correlated with having unusual peroxisomes, we searched for peroxisome biogenesis machinery, finding very few components in Cymbomonas, suggestive of peroxisome degeneration. Overall, we demonstrate conservation of MTCs across green algae, but with notable taxon-specific losses suggestive of unusual endomembrane systems.

VAMP7j: A Splice Variant of Human VAMP7 That Modulates Neurite Outgrowth by Regulating L1CAM Transport to the Plasma Membrane.

The vesicle-associated membrane protein 7 (VAMP7) is a SNARE protein of the longin family involved in a wide range of subcellular trafficking events, including neurite sprouting and elongation. The expression of the human gene SYBL1, encoding VAMP7, is finely regulated by alternative splicing. Among the minor isoforms identified so far, VAMP7j is the one most expressed and modulated in the human brain. Therefore, we focused on gaining functional evidence on VAMP7j, which lacks a functional SNARE motif but retains both the longin and transmembrane domains. In human SH-SY5Y cells, we found VAMP7j to modulate neuritogenesis by mediating transport of L1CAM toward the plasma membrane, in a fashion regulated by phosphorylation of the longin domain. VAMP7-mediated regulation of L1CAM trafficking seems at least to differentiate humans from rats, with VAMP7j CNS expression being restricted to primates, including humans. Since L1CAM is a central player in neuritogenesis and axon guidance, these findings suggest the species-specific splicing of SYBL1 is among the fine tuners of human neurodevelopmental complexity.

Reverse Plasticity Underlies Rapid Evolution by Clonal Selection within Populations of Fibroblasts Propagated on a Novel Soft Substrate.

Mechanical properties such as substrate stiffness are a ubiquitous feature of a cell's environment. Many types of animal cells exhibit canonical phenotypic plasticity when grown on substrates of differing stiffness, in vitro and in vivo. Whether such plasticity is a multivariate optimum due to hundreds of millions of years of animal evolution, or instead is a compromise between conflicting selective demands, is unknown. We addressed these questions by means of experimental evolution of populations of mouse fibroblasts propagated for approximately 90 cell generations on soft or stiff substrates. The ancestral cells grow twice as fast on stiff substrate as on soft substrate and exhibit the canonical phenotypic plasticity. Soft-selected lines derived from a genetically diverse ancestral population increased growth rate on soft substrate to the ancestral level on stiff substrate and evolved the same multivariate phenotype. The pattern of plasticity in the soft-selected lines was opposite of the ancestral pattern, suggesting that reverse plasticity underlies the observed rapid evolution. Conversely, growth rate and phenotypes did not change in selected lines derived from clonal cells. Overall, our results suggest that the changes were the result of genetic evolution and not phenotypic plasticity per se. Whole-transcriptome analysis revealed consistent differentiation between ancestral and soft-selected populations, and that both emergent phenotypes and gene expression tended to revert in the soft-selected lines. However, the selected populations appear to have achieved the same phenotypic outcome by means of at least two distinct transcriptional architectures related to mechanotransduction and proliferation.

Diversification of CORVET tethers facilitates transport complexity in Tetrahymena thermophila.

In endolysosomal networks, two hetero-hexameric tethers called HOPS and CORVET are found widely throughout eukaryotes. The unicellular ciliate Tetrahymena thermophila possesses elaborate endolysosomal structures, but curiously both it and related protozoa lack the HOPS tether and several other trafficking proteins, while retaining the related CORVET complex. Here, we show that Tetrahymena encodes multiple paralogs of most CORVET subunits, which assemble into six distinct complexes. Each complex has a unique subunit composition and, significantly, shows unique localization, indicating participation in distinct pathways. One pair of complexes differ by a single subunit (Vps8), but have late endosomal versus recycling endosome locations. While Vps8 subunits are thus prime determinants for targeting and functional specificity, determinants exist on all subunits except Vps11. This unprecedented expansion and diversification of CORVET provides a potent example of tether flexibility, and illustrates how 'backfilling' following secondary losses of trafficking genes can provide a mechanism for evolution of new pathways.This article has an associated First Person interview with the first author of the paper.

Evolutionary bioenergetics of ciliates.

Understanding why various organisms evolve alternative ways of living requires information on both the fitness advantages of phenotypic modifications and the costs of constructing and operating cellular features. Although the former has been the subject of a myriad of ecological studies, almost no attention has been given to how organisms allocate resources to alternative structures and functions. We address these matters by capitalizing on an array of observations on diverse ciliate species and from the emerging field of evolutionary bioenergetics. A relatively robust and general estimator for the total cost of a cell per cell cycle (in units of ATP equivalents) is provided, and this is then used to understand how the magnitudes of various investments scale with cell size. Among other things, we examine the costs associated with the large macronuclear genomes of ciliates, as well as ribosomes, various internal membranes, osmoregulation, cilia, and swimming activities. Although a number of uncertainties remain, the general approach taken may serve as blueprint for expanding this line of work to additional traits and phylogenetic lineages.

Unexpected organellar locations of ESCRT machinery in Giardia intestinalis and complex evolutionary dynamics spanning the transition to parasitism in the lineage Fornicata.

BACKGROUND: Comparing a parasitic lineage to its free-living relatives is a powerful way to understand how that evolutionary transition to parasitism occurred. Giardia intestinalis (Fornicata) is a leading cause of gastrointestinal disease world-wide and is famous for its unusual complement of cellular compartments, such as having peripheral vacuoles instead of typical endosomal compartments. Endocytosis plays an important role in Giardia's pathogenesis. Endosomal sorting complexes required for transport (ESCRT) are membrane-deforming proteins associated with the late endosome/multivesicular body (MVB). MVBs are ill-defined in G. intestinalis, and roles for identified ESCRT-related proteins are not fully understood in the context of its unique endocytic system. Furthermore, components thought to be required for full ESCRT functionality have not yet been documented in this species. RESULTS: We used genomic and transcriptomic data from several Fornicata species to clarify the evolutionary genome streamlining observed in Giardia, as well as to detect any divergent orthologs of the Fornicata ESCRT subunits. We observed differences in the ESCRT machinery complement between Giardia strains. Microscopy-based investigations of key components of ESCRT machinery such as GiVPS36 and GiVPS25 link them to peripheral vacuoles, highlighting these organelles as simplified MVB equivalents. Unexpectedly, we show ESCRT components associated with the endoplasmic reticulum and, for the first time, mitosomes. Finally, we identified the rare ESCRT component CHMP7 in several fornicate representatives, including Giardia and show that contrary to current understanding, CHMP7 evolved from a gene fusion of VPS25 and SNF7 domains, prior to the last eukaryotic common ancestor, over 1.5 billion years ago. CONCLUSIONS: Our findings show that ESCRT machinery in G. intestinalis is far more varied and complete than previously thought, associates to multiple cellular locations, and presents changes in ESCRT complement which pre-date adoption of a parasitic lifestyle.

Seeing the endomembrane system for the trees: Evolutionary analysis highlights the importance of plants as models for eukaryotic membrane-trafficking.

Plant cells show many signs of a unique evolutionary history. This is seen in the system of intracellular organelles and vesicle transport pathways plants use to traffic molecular cargo. Bioinformatic and cell biological work in this area is beginning to tackle the question of how plant cells have evolved, and what this tells us about the evolution of other eukaryotes. Key protein families with membrane trafficking function, including Rabs, SNAREs, vesicle coat proteins, and ArfGAPs, show patterns of evolution that indicate both specialization and conservation in plants. These changes are accompanied by changes at the level of organelles and trafficking pathways between them. Major specializations include losses of several ancient Rabs, novel functions of many proteins, and apparent modification of trafficking in endocytosis and cytokinesis. Nevertheless, plants show extensive conservation of ancestral membrane trafficking genes, and conservation of their ancestral function in most duplicates. Moreover, plants have retained several ancient membrane trafficking genes lost in the evolution of animals and fungi. Considering this, plants such as Arabidopsis are highly valuable for investigating not only plant-specific aspects of membrane trafficking, but also general eukaryotic mechanisms.

Emerging Frontiers in the Study of Molecular Evolution.

A collection of the editors of Journal of Molecular Evolution have gotten together to pose a set of key challenges and future directions for the field of molecular evolution. Topics include challenges and new directions in prebiotic chemistry and the RNA world, reconstruction of early cellular genomes and proteins, macromolecular and functional evolution, evolutionary cell biology, genome evolution, molecular evolutionary ecology, viral phylodynamics, theoretical population genomics, somatic cell molecular evolution, and directed evolution. While our effort is not meant to be exhaustive, it reflects research questions and problems in the field of molecular evolution that are exciting to our editors.

Identification and characterisation of a cryptic Golgi complex in Naegleria gruberi.

Although the Golgi complex has a conserved morphology of flattened stacked cisternae in most eukaryotes, it has lost the stacked organisation in several lineages, raising the question of what range of morphologies is possible for the Golgi. In order to understand this diversity, it is necessary to characterise the Golgi in many different lineages. Here, we identify the Golgi complex in Naegleria, one of the first descriptions of an unstacked Golgi organelle in a non-parasitic eukaryote, other than fungi. We provide a comprehensive list of Golgi-associated membrane trafficking genes encoded in two species of Naegleria and show that nearly all are expressed in mouse-passaged N. fowleri cells. We then study distribution of the Golgi marker (Ng)CopB by fluorescence in Naegleria gruberi, identifying membranous structures that are disrupted by Brefeldin A treatment, consistent with Golgi localisation. Confocal and immunoelectron microscopy reveals that NgCOPB localises to tubular membranous structures. Our data identify the Golgi organelle for the first time in this major eukaryotic lineage, and provide the rare example of a tubular morphology, representing an important sampling point for the comparative understanding of Golgi organellar diversity.This article has an associated First Person interview with the first author of the paper.

Whence Blobs? Phylogenetics of functional protein condensates.

What do we know about the molecular evolution of functional protein condensation? The capacity of proteins to form biomolecular condensates (compact, protein-rich states, not bound by membranes, but still separated from the rest of the contents of the cell) appears in many cases to be bestowed by weak, transient interactions within one or between proteins. Natural selection is expected to remove or fix amino acid changes, insertions or deletions that preserve and change this condensation capacity when doing so is beneficial to the cell. A few recent studies have begun to explore this frontier of phylogenetics at the intersection of biophysics and cell biology.

Mitochondrial protein import in trypanosomes: Expect the unexpected.

Mitochondria have many different functions, the most important one of which is oxidative phosphorylation. They originated from an endosymbiotic event between a bacterium and an archaeal host cell. It was the evolution of a protein import system that marked the boundary between the endosymbiotic ancestor of the mitochondrion and a true organelle that is under the control of the nucleus. In present day mitochondria more than 95% of all proteins are imported from the cytosol in a proces mediated by hetero-oligomeric protein complexes in the outer and inner mitochondrial membranes. In this review we compare mitochondrial protein import in the best studied model system yeast and the parasitic protozoan Trypanosoma brucei. The 2 organisms are phylogenetically only remotely related. Despite the fact that mitochondrial protein import has the same function in both species, only very few subunits of their import machineries are conserved. Moreover, while yeast has 2 inner membrane protein translocases, one specialized for presequence-containing and one for mitochondrial carrier proteins, T. brucei has a single inner membrane translocase only, that mediates import of both types of substrates. The evolutionary implications of these findings are discussed.

Evolutionary dynamics of the kinetochore network in eukaryotes as revealed by comparative genomics.

During eukaryotic cell division, the sister chromatids of duplicated chromosomes are pulled apart by microtubules, which connect via kinetochores. The kinetochore is a multiprotein structure that links centromeres to microtubules, and that emits molecular signals in order to safeguard the equal distribution of duplicated chromosomes over daughter cells. Although microtubule-mediated chromosome segregation is evolutionary conserved, kinetochore compositions seem to have diverged. To systematically inventory kinetochore diversity and to reconstruct its evolution, we determined orthologs of 70 kinetochore proteins in 90 phylogenetically diverse eukaryotes. The resulting ortholog sets imply that the last eukaryotic common ancestor (LECA) possessed a complex kinetochore and highlight that current-day kinetochores differ substantially. These kinetochores diverged through gene loss, duplication, and, less frequently, invention and displacement. Various kinetochore components co-evolved with one another, albeit in different manners. These co-evolutionary patterns improve our understanding of kinetochore function and evolution, which we illustrated with the RZZ complex, TRIP13, the MCC, and some nuclear pore proteins. The extensive diversity of kinetochore compositions in eukaryotes poses numerous questions regarding evolutionary flexibility of essential cellular functions.

A sophisticated, differentiated Golgi in the ancestor of eukaryotes.

BACKGROUND: The Golgi apparatus is a central meeting point for the endocytic and exocytic systems in eukaryotic cells, and the organelle's dysfunction results in human disease. Its characteristic morphology of multiple differentiated compartments organized into stacked flattened cisternae is one of the most recognizable features of modern eukaryotic cells, and yet how this is maintained is not well understood. The Golgi is also an ancient aspect of eukaryotes, but the extent and nature of its complexity in the ancestor of eukaryotes is unclear. Various proteins have roles in organizing the Golgi, chief among them being the golgins. RESULTS: We address Golgi evolution by analyzing genome sequences from organisms which have lost stacked cisternae as a feature of their Golgi and those that have not. Using genomics and immunomicroscopy, we first identify Golgi in the anaerobic amoeba Mastigamoeba balamuthi. We then searched 87 genomes spanning eukaryotic diversity for presence of the most prominent proteins implicated in Golgi structure, focusing on golgins. We show some candidates as animal specific and others as ancestral to eukaryotes. CONCLUSIONS: None of the proteins examined show a phyletic distribution that correlates with the morphology of stacked cisternae, suggesting the possibility of stacking as an emergent property. Strikingly, however, the combination of golgins conserved among diverse eukaryotes allows for the most detailed reconstruction of the organelle to date, showing a sophisticated Golgi with differentiated compartments and trafficking pathways in the common eukaryotic ancestor.

The Paradigms They Are a-Changin': past, present and future of PVC bacteria research.

These are exciting times for PVC researchers! The PVC superphylum is composed of the bacterial phyla Planctomycetes, Verrucomicrobia, Chlamydiae (those three founders giving it its name), Lentisphaerae and Kirimatiellaeota as well as some uncultured candidate phyla, such as the Candidatus Omnitrophica (previously known as OP3). Despite early debates, most of the disagreements that surround this group of bacteria have been recently resolved. In this article, we review the history of the study of PVC bacteria, with a particular focus on the misinterpretations that emerged early in the field and their resolution. We begin with a historical perspective that describes the relevant facts of PVC research from the early times when they were not yet termed PVC. Those were controversial times and we refer to them as the "discovery age" of the field. We continue by describing new discoveries due to novel techniques and data that combined with the reinterpretations of old ones have contributed to solve most of the discordances and we refer to these times as the "illumination age" of PVC research. We follow by arguing that we are just entering the "golden age" of PVC research and that the future of this growing community is looking bright. We finish by suggesting a few of the directions that PVC researches might take in the future.

The evolution of ERMIONE in mitochondrial biogenesis and lipid homeostasis: An evolutionary view from comparative cell biology.

The ER-mitochondria organizing network (ERMIONE) in Saccharomyces cerevisiae is involved in maintaining mitochondrial morphology and lipid homeostasis. ERMES and MICOS are two scaffolding complexes of ERMIONE that contribute to these processes. ERMES is ancient but has been lost in several lineages including animals, plants, and SAR (stramenopiles, alveolates and rhizaria). On the other hand, MICOS is ancient and has remained present in all organisms bearing mitochondrial cristae. The ERMIONE precursor evolved in the α-proteobacterial ancestor of mitochondria which had the central subunit of MICOS, Mic60. The subsequent evolution of ERMIONE and its interactors in eukaryotes reflects the integrative co-evolution of mitochondria and their hosts and the adaptive paths that some lineages have followed in their specialization to certain environments. By approaching the ERMIONE from a perspective of comparative evolutionary cell biology, we hope to shed light on not only its evolutionary history, but also how ERMIONE components may function in organisms other than S. cerevisiae. This article is part of a Special Issue entitled: The cellular lipid landscape edited by Tim P. Levine and Anant K. Menon.