Endosymbiotic origins of sex

Understanding how complex sexual reproduction arose, and why sexual organisms have been more successful than otherwise similar asexual organisms, is a longstanding problem in evolutionary biology. Within this problem, the potential role of endosymbionts or intracellular pathogens in mediating primitive genetic transfers is a continuing theme. In recent years, several remarkable activities of mitochondria have been observed in the germline cells of complex eukaryotes, and it has been found that bacterial endosymbionts related to mitochondria are capable of manipulating diverse aspects of metazoan gametogenesis. An attempt is made here to rationalize these observations with an endosymbiotic model for the evolutionary origins of sex. It is hypothesized that the contemporary life cycle of germline cells has descended from the life cycle of the endosymbiotic ancestor of the mitochondrion. Through an actin-based motility that drove it from one cell to another, the rickettsial ancestor of mitochondria may have functioned as a primitive transducing particle, the evolutionary progenitor of sperm.     

Structure of the bc1 complex from Seculamonas ecuadoriensis,a jakobid flagellate with an ancestral mitochondrial genome

In eubacteria, the respiratory bc(1) complex (complex III) consists of three or four different subunits, whereas that of mitochondria, which have descended from an alpha-proteobacterial endosymbiont, contains about seven additional subunits. To understand better how mitochondrial protein complexes evolved from their simpler bacterial predecessors, we purified complex III of Seculamonas ecuadoriensis, a member of the jakobid protists, which possess the most bacteria-like mitochondrial genomes known. The S. ecuadoriensis complex III has an apparent molecular mass of 460 kDa and exhibits antimycin-sensitive quinol:cytochrome c oxidoreductase activity. It is composed of at least eight subunits between 6 and 46 kDa in size, including two large "core" subunits and the three "respiratory" subunits. The molecular mass of the S. ecuadoriensis bc(1) complex is slightly lower than that reported for other eukaryotes, but about 2x as large as complex III in bacteria. This indicates that the departure from the small bacteria-like complex III took place at an early stage in mitochondrial evolution, prior to the divergence of jakobids. We posit that the recruitment of additional subunits in mitochondrial respiratory complexes is a consequence of the migration of originally alpha-proteobacterial genes to the nucleus.     

Cytonuclear coevolution: the genomics of cooperation

Without mitochondria we would be in big trouble, and there would be a global biological energy crisis if it were not for chloroplasts. Fortunately, genomic evolution over the past two billion years has ensured that the functions of these key organelles are with us to stay. Whole-genome analyses have not only proven that mitochondria and chloroplasts are descended from formerly free-living bacteria, but have also shown that it is difficult to define eukaryotes without reference to the fusion and coevolution of host and endosymbiont genomes. Here, we review how the macro- and microevolutionary insights that follow from the genomics of cytonuclear interactions are uniting molecular evolution, structural proteomics, population genetics and problems in aging and disease. Our goals are to clarify the coevolutionary events that have governed nuclear and organelle evolution, and to encourage further critical analyses of these interactions as problems in the study of co-adapted gene complexes.     

Correlating single cell motility with population growth dynamics for flagellated bacteria

Many bacteria used for biotechnological applications are naturally motile. Their "bio-nanopropeller" driven movement allows searching for better environments in a process called chemotaxis. Since bacteria are extremely small in size compared to the bulk fluid volumes in bioreactors, single cell motility is not considered to influence bioreactor operations. However, with increasing interest in localized fluid flow inside reactors, it is important to ask whether individual motility characteristics of bacteria are important in bioreactor operations. The first step in this direction is to try to correlate single cell measurements with population data of motile bacteria in a bioreactor. Thus, we observed the motility behavior of individual bacterial cells, using video microscopy with 33 ms time resolution, as a function of population growth dynamics of batch cultures in shake flasks. While observing the motility behavior of the most intensively studied bacteria, Escherichia coli, we find that overall bacterial motility decreases with progression of the growth curve. Remarkably, this is due to a decrease in a specific motility behavior called "running". Our results not only have direct implications on biofilm formations, but also provide a new direction in bioprocess design research highlighting the role of individual bacterial cell motility as an important parameter.     

LUMP is a putative double-stranded RNA binding protein required for male fertility in Drosophila melanogaster

In animals, male fertility requires the successful development of motile sperm. During Drosophila melanogaster spermatogenesis, 64 interconnected spermatids descended from a single germline stem cell are resolved into motile sperm in a process termed individualization. Here we identify a putative double-stranded RNA binding protein LUMP that is required for male fertility. lump(1) mutants are male-sterile and lack motile sperm due to defects in sperm individualization. We show that one dsRNA binding domains (dsRBD) is essential for LUMP function in male fertility. These findings reveal LUMP is a novel factor required for late stages of male germline differentiation.     

Bacterial proteins predisposed for targeting to mitochondria

Mitochondria evolved from an endosymbiotic proteobacterium in a process that required the transfer of genes from the bacterium to the host cell nucleus, and the translocation of proteins thereby made in the host cell cytosol into the internal compartments of the organelle. According to current models for this evolution, two highly improbable events are required to occur simultaneously: creation of a protein translocation machinery to import proteins back into the endosymbiont and creation of targeting sequences on the protein substrates themselves. Using a combination of two independent prediction methods, validated through tests on simulated genomes, we show that at least 5% of proteins encoded by an extant proteobacterium are predisposed for targeting to mitochondria, and propose we that mitochondrial targeting information was preexisting for many proteins of the endosymbiont. We analyzed a family of proteins whose members exist both in bacteria and in mitochondria of eukaryotes and show that the amino-terminal extensions occasionally found in bacterial family members can function as a crude import sequence when the protein is presented to isolated mitochondria. This activity leaves the development of a primitive translocation channel in the outer membrane of the endosymbiont as a single hurdle to initiating the evolution of mitochondria.     

Using experimental evolution to explore natural patterns between bacterial motility and resistance to bacteriophages

Resistance of bacteria to phages may be gained by alteration of surface proteins to which phages bind, a mechanism that is likely to be costly as these molecules typically have critical functions such as movement or nutrient uptake. To address this potential trade-off, we combine a systematic study of natural bacteria and phage populations with an experimental evolution approach. We compare motility, growth rate and susceptibility to local phages for 80 bacteria isolated from horse chestnut leaves and, contrary to expectation, find no negative association between resistance to phages and bacterial motility or growth rate. However, because correlational patterns (and their absence) are open to numerous interpretations, we test for any causal association between resistance to phages and bacterial motility using experimental evolution of a subset of bacteria in both the presence and absence of naturally associated phages. Again, we find no clear link between the acquisition of resistance and bacterial motility, suggesting that for these natural bacterial populations, phage-mediated selection is unlikely to shape bacterial motility, a key fitness trait for many bacteria in the phyllosphere. The agreement between the observed natural pattern and the experimental evolution results presented here demonstrates the power of this combined approach for testing evolutionary trade-offs.     

Live cell imaging of mitochondrial movement along actin cables in budding yeast

BACKGROUND: Mitochondrial inheritance is essential for cell division. In budding yeast, mitochondrial movement from mother to daughter requires (1) actin cables, F-actin bundles that undergo retrograde movement during elongation from buds into mother cells; (2) the mitochore, a mitochondrial protein complex implicated in linking mitochondria to actin cables; and (3) Arp2/3 complex-mediated force generation on mitochondria. RESULTS: We observed three new classes of mitochondrial motility: anterograde movement at velocities of 0.2-0.33 microm/s, retrograde movement at velocities of 0.26-0.51 microm/s, and no net anterograde or retrograde movement. In all cases, motile mitochondria were associated with actin cables undergoing retrograde flow at velocities of 0.18-0.62 microm/s. Destabilization of actin cables or mutations of the mitochore blocked all mitochondrial movements. In contrast, mutations in the Arp2/3 complex affected anterograde but not retrograde mitochondrial movements. CONCLUSIONS: Actin cables are required for movement of mitochondria, secretory vesicles, mRNA, and spindle alignment elements in yeast. We provide the first direct evidence that one of the proposed cargos use actin cables as tracks. In the case of mitochondrial inheritance, anterograde movement drives transfer of the organelle from mothers to buds, while retrograde movement contributes to retention of the organelle in mother cells. Interaction of mitochondria with actin cables is required for anterograde and retrograde movement. In contrast, force generation on mitochondria is required only for anterograde movement. Finally, we propose a novel mechanism in which actin cables serve as "conveyor belts" that drive retrograde organelle movement.     

Microbiome‐Germline Interactions and Their Transgenerational Implications

It is becoming increasingly clear that most, if not all, animals and plants are associated with a diverse array of resident gut microbiota. This symbiosis is regulated by host‐microbiome interactions which influence the development, homeostasis, adaptation and evolution of the host. Recent evidence indicated that these interactions can also affect the host germline and have a potential of supporting transgenerational effects, including inheritance of acquired characteristics. Taken together, the influence of gut bacteria on the host soma and germline could potentially give rise to emergent phenotypes, which may be partially inherited by three distinguishable modes of transgenerational influence of gut bacteria: 1) “soma‐to‐soma” 2) “soma‐to‐germline” and 3) “soma‐germline‐soma”. Here, we discuss these possibilities in light of evidence supporting bacterial‐mediated modes of transgenerational inheritance.

     

Phylogenetic analyses of fat body endosymbionts reveal differences in invasion times of blaberid wood-feeding cockroaches (Blaberidae: Panesthiinae) into the Japanese archipelago

Cockroaches have endosymbiotic bacteria in their fat bodies. Recent molecular phylogenetic analyses on both hosts and endosymbionts have revealed that co-evolution has occurred throughout the history of cockroaches and termites. Co-cladogenesis was also shown among closely related taxa (woodroach genus Cryptocercus; Cryptocercidae), and thus endosymbiont data are likely to be useful for biogeographical analyses. To test the possibility of co-cladogenesis among inter-and intraspecific taxa, as well as the utility of endosymbiont data for inferring biogeographical scenarios, we analyzed rRNA genes of endosymbionts of Japanese and Taiwanese Panesthiinae (Salganea and Panesthia; Blaberidae), on which phylogenetic analyses previously had been performed based on the mitochondrial genes. Statistical analyses on the topologies inferred from both endosymbiont and host mitochondria genes showed that co-cladogenesis has occurred. The endosymbiont sequences examined appear to have evolved in a clock-like manner, and their rate of evolution based on the host fossil data showed a major difference in the time of invasion of the two Japanese genera, that is congruent with the recent analyses of their mitochondrial genes.     

Origins of hydrogenosomes and mitochondria

Complete genome sequences for many oxygen-respiring mitochondria, as well as for some bacteria, leave no doubt that mitochondria are descendants of alpha-proteobacteria, a finding for which the endosymbiont hypothesis can easily account. Yet a wealth of data indicate that mitochondria and hydrogenosomes - the ATP-producing organelles of many anaerobic protists - share a common ancestry, a finding that traditional formulations of the endosymbiont hypothesis less readily accommodates. Available evidence suggests that a more in-depth understanding of the origins of eukaryotes and their organelles will hinge upon data from the genomes of protists that synthesize ATP without the need for oxygen.     

Translocation of precytochrome C2 into intracytoplasmic membrane vesicles of Rhodobacter capsulatus requires a peripheral membrane protein

Rhodobacter capsulatus is a member of the group α-purple bacteria which are closely related to the ancestral endosymbiont that gave rise to mitochondria. It has therefore been hypothesized that the molecular mechanisms governing protein export in α-purple bacteria have been conserved during the evolution of mitochondria. To enable analysis of protein export in α-purple bacteria we describe here the development of a homologous cell-free synthesis/export system consisting entirely of components of R. capsulatus. Translocation of precytochrome C₂ into intracytoplasmic membrane vesicles of this organism was found to require the proton-motive force and proceed at a significantly higher efficiency when membranes were present during protein synthesis. Furthermore, we show that, in this cell-free system, translocation depends on a preparation of peripheral membrane proteins Which do not possess detectable SecA- and SecB-like actvities.     

Biogenesis of beta-barrel membrane proteins of mitochondria

beta-Barrel membrane proteins have several important functions in outer membranes of Gram-negative bacteria and in the organelles of endosymbiotic origin, mitochondria and chloroplasts. The biogenesis of beta-barrel membrane proteins was, until recently, an unresolved process. A breakthrough was achieved when a specific pathway for the insertion of beta-barrel outer-membrane proteins was identified in both mitochondria and Gram-negative bacteria. The key component of this pathway is Tob55 (also known as Sam50) in mitochondria and Omp85 in bacteria, both beta-barrel membrane proteins themselves. Tob55 is part of the hetero-oligomeric TOB (topogenesis of mitochondrial outer-membrane beta-barrel proteins) or SAM (sorting and assembly of mitochondria) complex, which is present in the mitochondrial outer membrane. Tob55 belongs to an evolutionarily conserved protein family, the members of which are present in almost all eukaryotes and in Gram-negative bacteria and chloroplasts. Thus, is it emphasized that the insertion pathway of mitochondrial beta-barrel membrane proteins was conserved during evolution of mitochondria from endosymbiotic bacterial ancestors.     

Scission,spores, and apoptosis: a proposal for the evolutionary origin of mitochondria in cell death induction

Mitochondria fragment prior to caspase activation during many pathways of apoptosis. Inhibition of the machinery that normally regulates mitochondrial morphology in healthy cells inhibits the fission that occurs during apoptosis and actually delays the process of cell death. Interestingly, there are certain parallels between mitochondrial fission and bacterial sporulation. As bacterial sporulation can be considered a stress response we suggest that a primordial stress response of endosymbiont mitochondrial progenitors may have been adopted for the stress response of early eukaryotes. Thus, the mitochondrial fission process may represent an early stress response of primitive mitochondria that could have integrated the stress signals and acted as an initial sensor for the eukaryotic response system. The fact that mitochondria fragment during apoptosis using the machinery descended from or that superceded the bacterial stress response of sporulation is consistent with this hypothesis. This hypothesis would explain why what is generally considered the "power house" of the cell came to integrate the cell death response and regulate apoptosis.     

Step-wise loss of bacterial flagellar torsion confers progressive phagocytic evasion

Phagocytosis of bacteria by innate immune cells is a primary method of bacterial clearance during infection. However, the mechanisms by which the host cell recognizes bacteria and consequentially initiates phagocytosis are largely unclear. Previous studies of the bacterium Pseudomonas aeruginosa have indicated that bacterial flagella and flagellar motility play an important role in colonization of the host and, importantly, that loss of flagellar motility enables phagocytic evasion. Here we use molecular, cellular, and genetic methods to provide the first formal evidence that phagocytic cells recognize bacterial motility rather than flagella and initiate phagocytosis in response to this motility. We demonstrate that deletion of genes coding for the flagellar stator complex, which results in non-swimming bacteria that retain an initial flagellar structure, confers resistance to phagocytic binding and ingestion in several species of the gamma proteobacterial group of Gram-negative bacteria, indicative of a shared strategy for phagocytic evasion. Furthermore, we show for the first time that susceptibility to phagocytosis in swimming bacteria is proportional to mot gene function and, consequently, flagellar rotation since complementary genetically- and biochemically-modulated incremental decreases in flagellar motility result in corresponding and proportional phagocytic evasion. These findings identify that phagocytic cells respond to flagellar movement, which represents a novel mechanism for non-opsonized phagocytic recognition of pathogenic bacteria.     

Interaction with type IV pili induces structural changes in the bacterial outer membrane secretin PilQ

Type IV pili are cell surface organelles found on many Gram-negative bacteria. They mediate a variety of functions, including adhesion, twitching motility, and competence for DNA uptake. The type IV pilus is a helical polymer of pilin protein subunits and is capable of rapid polymerization or depolymerization, generating large motor forces in the process. Here we show that a specific interaction between the outer membrane secretin PilQ and the type IV pilus fiber can be detected by far-Western analysis and sucrose density gradient centrifugation. Transmission electron microscopy of preparations of purified pili, to which the purified PilQ oligomer had been added, showed that PilQ was uniquely located at one end of the pilus fiber, effectively forming a "mallet-type" structure. Determination of the three-dimensional structure of the PilQ-type IV pilus complex at 26-angstroms resolution showed that the cavity within the protein complex was filled. Comparison with a previously determined structure of PilQ at 12-angstroms resolution indicated that binding of the pilus fiber induced a dissociation of the "cap" feature and lateral movement of the "arms" of the PilQ oligomer. The results demonstrate that the PilQ structure exhibits a dynamic response to the binding of its transported substrate and suggest that the secretin could play an active role in type IV pilus assembly as well as secretion.     

Ancestral and derived protein import pathways in the mitochondrion of Reclinomonas americana

The evolution of mitochondria from ancestral bacteria required that new protein transport machinery be established. Recent controversy over the evolution of these new molecular machines hinges on the degree to which ancestral bacterial transporters contributed during the establishment of the new protein import pathway. Reclinomonas americana is a unicellular eukaryote with the most gene-rich mitochondrial genome known, and the large collection of membrane proteins encoded on the mitochondrial genome of R. americana includes a bacterial-type SecY protein transporter. Analysis of expressed sequence tags shows R. americana also has components of a mitochondrial protein translocase or "translocase in the inner mitochondrial membrane complex." Along with several other membrane proteins encoded on the mitochondrial genome Cox11, an assembly factor for cytochrome c oxidase retains sequence features suggesting that it is assembled by the SecY complex in R. americana. Despite this, protein import studies show that the RaCox11 protein is suited for import into mitochondria and functional complementation if the gene is transferred into the nucleus of yeast. Reclinomonas americana provides direct evidence that bacterial protein transport pathways were retained, alongside the evolving mitochondrial protein import machinery, shedding new light on the process of mitochondrial evolution.     

Effects of intermediate filaments on actin-based motility of Listeria monocytogenes.

How does subcellular architecture influence the intracellular movements of large organelles and macromolecular assemblies? To investigate the effects of mechanical changes in cytoplasmic structure on intracellular motility, we have characterized the actin-based motility of the intracellular bacterial pathogen Listeria monocytogenes in normal mouse fibroblasts and in fibroblasts lacking intermediate filaments. The apparent diffusion coefficient of L. monocytogenes was two-fold greater in vimentin-null fibroblasts than in wild-type fibroblasts, indicating that intermediate filaments significantly restrict the Brownian motion of bacteria. However, the mean speed of L. monocytogenes actin-based motility was statistically identical in vimentin-null and wild-type cells. Thus, environmental drag is not rate limiting for bacterial motility. Analysis of the temporal variations in speed measurements indicated that bacteria in vimentin-null cells displayed larger fluctuations in speed than did trajectories in wild-type cells. Similarly, the presence of the vimentin meshwork influenced the turning behavior of the bacteria; in the vimentin-null cells, bacteria made sharper turns than they did in wild-type cells. Taken together, these results suggest that a network of intermediate filaments constrains bacterial movement and operates over distances of several microns to reduce fluctuations in motile behavior.     

Mitochondrial toxicity detected in a health product with a boar spermatozoan bioassay

Seaweed and organic alfalfa capsules sold as "health promoting" products had repeatedly caused emesis in a consumer. Using the boar spermatozoan bioassay, the capsule contents were found to contain a toxic substance that inhibited boar sperm motility and depolarised mitochondria at low exposure concentrations of 10 microg/ml. The capsule also contained high amounts (10(5)-10(7) cfu/g), of endospore-forming bacteria and Streptomyces-like bacteria. Bacteria from the capsule produced toxic substances when cultured in the laboratory. Three different toxic responses were provoked in the spermatozoa exposed to extracts from the Streptomyces-like isolates: a) hyperpolarisation of the plasma membrane and depolarisation of the mitochondria; b) depolarisation of mitochondria similar to that caused by the capsule content extract; and c) motility inhibition, with no observed change of any cytosolic transmembrane potential. Membrane potential changes in the sperm cells exposed to the bacterial extracts were similar to those provoked by exposure to valinomycin and bafilomycin A1, to nigericin, and to oligomycin and ionomycin, respectively. Extracts prepared from Bacillus isolated from the capsule non-specifically depolarised all the cellular transmembrane potentials. The results demonstrate the potential value of a cell toxicity assay with boar spermatozoa for detecting hazardous substances in products intended for human consumption, without whole-animal exposure or using fetal calf serum for cell cultures.     

Dynamics of reductive genome evolution in mitochondria and obligate intracellular microbes

Reductive evolution in mitochondria and obligate intracellular microbes has led to a significant reduction in their genome size and guanine plus cytosine content (GC). We show that genome shrinkage during reductive evolution in prokaryotes follows an exponential decay pattern and provide a method to predict the extent of this decay on an evolutionary timescale. We validated predictions by comparison with estimated extents of genome reduction known to have occurred in mitochondria and Buchnera aphidicola, through comparative genomics and by drawing on available fossil evidences. The model shows how the mitochondrial ancestor would have quickly shed most of its genome, shortly after its incorporation into the protoeukaryotic cell and prior to codivergence subsequent to the split of eukaryotic lineages. It also predicts that the primary rickettsial parasitic event would have occurred between 180 and 425 million years ago (MYA), an event of relatively recent evolutionary origin considering the fact that Rickettsia and mitochondria evolved from a common alphaproteobacterial ancestor. This suggests that the symbiotic events of Rickettsia and mitochondria originated at different time points. Moreover, our model results predict that the ancestor of Wigglesworthia glossinidia brevipalpis, dated around the time of origin of its symbiotic association with the tsetse fly (50-100 MYA), was likely to have been an endosymbiont itself, thus supporting an earlier proposition that Wigglesworthia, which is currently a maternally inherited primary endosymbiont, evolved from a secondary endosymbiont.