The origins of cellular life

All life on earth can be naturally classified into cellular life forms and virus-like selfish elements, the latter being fully dependent on the former for their reproduction. Cells are reproducers that not only replicate their genome but also reproduce the cellular organization that depends on semipermeable, energy-transforming membranes and cannot be recovered from the genome alone, under the famous dictum of Rudolf Virchow, Omnis cellula e cellula. In contrast, simple selfish elements are replicators that can complete their life cycles within the host cell starting from genomic RNA or DNA alone. The origin of the cellular organization is the central and perhaps the hardest problem of evolutionary biology. I argue that the origin of cells can be understood only in conjunction with the origin and evolution of selfish genetic elements. A scenario of precellular evolution is presented that involves cohesion of the genomes of the emerging cellular life forms from primordial pools of small genetic elements that eventually segregated into hosts and parasites. I further present a model of the coevolution of primordial membranes and membrane proteins, discuss protocellular and non-cellular models of early evolution, and examine the habitats on the primordial earth that could have been conducive to precellular evolution and the origin of cells.     

The origin of replicators and reproducers

Replicators are fundamental to the origin of life and evolvability. Their survival depends on the accuracy of replication and the efficiency of growth relative to spontaneous decay. Infrabiological systems are built of two coupled autocatalytic systems, in contrast to minimal living systems that must comprise at least a metabolic subsystem, a hereditary subsystem and a boundary, serving respective functions. Some scenarios prefer to unite all these functions into one primordial system, as illustrated in the lipid world scenario, which is considered as a didactic example in detail. Experimentally produced chemical replicators grow parabolically owing to product inhibition. A selection consequence is survival of everybody. The chromatographized replicator model predicts that such replicators spreading on surfaces can be selected for higher replication rate because double strands are washed away slower than single strands from the surface. Analysis of real ribozymes suggests that the error threshold of replication is less severe by about one order of magnitude than thought previously. Surface-bound dynamics is predicted to play a crucial role also for exponential replicators: unlinked genes belonging to the same genome do not displace each other by competition, and efficient and accurate replicases can spread. The most efficient form of such useful population structure is encapsulation by reproducing vesicles. The stochastic corrector model shows how such a bag of genes can survive, and what the role of chromosome formation and intragenic recombination could be. Prebiotic and early evolution cannot be understood without the models of dynamics.     

Chasing the Origin of Viruses: Capsid-Forming Genes as a Life-Saving Preadaptation within a Community of Early Replicators

Virus capsids mediate the transfer of viral genetic information from one cell to another, thus the origin of the first viruses arguably coincides with the origin of the viral capsid. Capsid genes are evolutionarily ancient and their emergence potentially predated even the origin of first free-living cells. But does the origin of the capsid coincide with the origin of viruses, or is it possible that capsid-like functionalities emerged before the appearance of true viral entities? We set to investigate this question by using a computational simulator comprising primitive replicators and replication parasites within a compartment matrix. We observe that systems with no horizontal gene transfer between compartments collapse due to the rapidly emerging replication parasites. However, introduction of capsid-like genes that induce the movement of randomly selected genes from one compartment to another rescues life by providing the non-parasitic replicators a mean to escape their current compartments before the emergence of replication parasites. Capsid-forming genes can mediate the establishment of a stable meta-population where parasites cause only local tragedies but cannot overtake the whole community. The long-term survival of replicators is dependent on the frequency of horizontal transfer events, as systems with either too much or too little genetic exchange are doomed to succumb to replication-parasites. This study provides a possible scenario for explaining the origin of viral capsids before the emergence of genuine viruses: in the absence of other means of horizontal gene transfer between compartments, evolution of capsid-like functionalities may have been necessary for early life to prevail.     

DNA elements modulating the <Emphasis Type="Italic">KARS12</Emphasis> chromosomal replicator in <Emphasis Type="Italic">Kluyveromyces lactis</Emphasis>

Eukaryotic chromosomal DNA replication is initiated by a highly conserved set of proteins that interact with cis-acting elements on chromosomes called replicators. Despite the conservation of replication initiation proteins, replicator sequences show little similarity from species to species in the small number of organisms that have been examined. Examination of replicators in other species is likely to reveal common features of replicators. We have examined a Kluyeromyces lactis replicator, KARS12, that functions as origin of DNA replication on plasmids and in the chromosome. It contains a 50-bp region with similarity to two other K. lactis replicators, KARS101 and the pKD1 replication origin. Replacement of the 50-bp sequence with an EcoRI site completely abrogated the ability of KARS12 to support plasmid and chromosomal DNA replication origin activity, demonstrating this sequence is a common feature of K. lactis replicators and is essential for function, possibly as the initiator protein binding site. Additional sequences up to 1 kb in length are required for efficient KARS12 function. Within these sequences are a binding site for a global regulator, Abf1p, and a region of bent DNA, both of which contribute to the activity of KARS12. These elements may facilitate protein binding, protein/protein interaction and/or nucleosome positioning as has been proposed for other eukaryotic origins of DNA replication.     

Imitation, internal absorption and the reversal of local drift in stochastic evolutionary games

Evolutionary game dynamics in finite populations is typically subject to noise, inducing effects which are not present in deterministic systems, including fixation and extinction. In the first part of this paper we investigate the phenomenon of drift reversal in finite populations, taking into account that drift is a local quantity in strategy space. Secondly, we study a simple imitation dynamics, and show that it can lead to fixation at internal mixed-strategy fixed points even in finite populations. Imitation in infinite populations is adequately described by conventional replicator dynamics, and these equations are known to have internal fixed points. Internal absorption in finite populations on the other hand is a novel dynamic phenomenon. Due to an outward drift in finite populations this type of dynamic arrest is not found in other commonly studied microscopic dynamics, not even in those with the same deterministic replicator limit as imitation.     

Models of Replicator Proliferation Involving Differential Replicator Subunit Stability

Several models for the origin of life involve molecules that are capable of self-replication, such as self-replicating polymers composed of RNA or DNA or amino acids. Here we consider a hypothetical replicator (AB) composed of two subunits, A and B. Programs written in Python and C programming languages were used to model AB replicator abundance as a function of cycles of replication (iterations), under specified hypothetical conditions. Two non-exclusive models describe how a reduced stability for B relative to A can have an advantage for replicator activity and/or evolution by generating free A subunits. In model 1, free A subunits associate with AB replicators to create AAB replicators with greater activity. In simulations, reduced stability of B was beneficial when the replication activity of AAB was greater than two times the replication activity of AB. In model 2, the free A subunit is inactive for some number of iterations before it re-creates the B subunit. A re-creates the B subunit with an equal chance of creating B or B′, where B′ is a mutant that increases AB’ replicator activity relative to AB. In simulations, at moderate number of iterations (< 15), a shorter survival time for B is beneficial when the stability of B is greater than the inactive time of A. The results are consistent with the hypothesis that reduced stability for a replicator subunit can be advantageous under appropriate conditions.     

Group selection of early replicators and the origin of life

A major problem of the origin of life has been that of information integration. As Eigen (1971) has shown, a mutant distribution of RNAs replicating without the aid of a replicase cannot integrate sufficient information for the functioning of a higher-level unit utilizing several types of encoded enzymes. He proposed the hypercycle model to bridge this gap in prebiology. It can be shown by a nonlinear game model, incorporating mutation of a hypercycle, that the selection properties of hypercycles make them inefficient information integrators as they cannot compete favourably with all kinds of less efficient information carriers or mutationally coupled hypercycles. The stochastic corrector model is presented as an alternative resolution of Eigen's paradox. It assumes that replicative templates are competing within replicative compartments, whose selective values depend on the internal template composition via a catalytic acid in replication and "metabolism". The dynamics of template replication are analyzed by numerical simulation of master equations. Due to the stochasticity in replication and compartment fission the best compartment types recur. An Eigen equation at the compartment level is set up and calculated. Even selfish template mutants cannot destroy the system though they make it less efficient. The genetic information of templates is evaluated at both levels, and the higher (compartment) level successfully constrains the lower (template) one. Compartmentation together with stochastic effects is sufficient to integrate information dispersed in competitive replicators. Compartment selection is considered to be group selection of replicators. Implications for the origin of life are discussed.     

Coexistence and error propagation in pre-biotic vesicle models: a group selection approach

Compartmentalization of unlinked, competing templates is widely accepted as a necessary step towards the evolution of complex organisms. However, preservation of information by templates confined to isolated vesicles of finite size faces much harder obstacles than by free templates: random drift allied to mutation pressure wipe out any template that does not replicate perfectly, no matter how small the error probability might be. In addition, drift alone hinders the coexistence of distinct templates in a same compartment. Here, we investigate the conditions for group selection to prevail over drift and mutation and hence to guarantee the maintenance and coexistence of distinct templates in a vesicle. Group selection is implemented through a vesicle survival probability that depends on the template composition. By considering the limit case of an infinite number of vesicles, each one carrying a finite number of templates, we were able to derive a set of recursion equations for the frequencies of vesicles with different template compositions. Numerical iteration of these recursions allows the exact characterization of the steady state of the vesicle population-a quasispecies of vesicles-thus revealing the values of the mutation and group selection intensities for which template coexistence is possible. Within the main assumption of the model-a fixed, finite or infinite, number of vesicles-we find no fundamental impediment to the coexistence of an arbitrary number of template types with the same replication rate inside a vesicle, except of course for the vesicle capacity. Group selection in the form of vesicle selection is a must for compartmentalized primordial genetic systems even in the absence of intra-genomic competition of different templates.     

The replicon revisited: an old model learns new tricks in metazoan chromosomes

The origins of DNA replication were proposed in the replicon model to be specified genetically by replicator elements that coordinate the initiation of DNA synthesis with gene expression and cell growth. Recent studies have identified DNA sequences in mammalian cells that fulfil the genetic criteria for replicators and are beginning to uncover the sequence requirements for the initiation of DNA replication. Mammalian replicators are com- posed of non-redundant modules that cooperate to direct initiation to specific chromosomal sites. Conversely, replicators do not show strong sequence similarity, and their ability to initiate replication depends on the chromosomal context and epigenetic factors, as well as their primary sequence. Here, we review the properties of metazoan replicators, and discuss the genetic and epigenetic factors that determine where and when DNA replication is initiated.     

A role for a replicator dominance mechanism in silencing.

The role of the natural HMR-E silencer in modulating replication initiation and silencing by the origin recognition complex (ORC) was examined. When natural HMR-E was the only silencer controlling HMR, the silencer's ORC-binding site (ACS) was dispensable for replication initiation but essential for silencing, indicating that a non-silencer chromosomal replicator(s) existed in close proximity to the silencer. Further analysis revealed that regions flanking both sides of HMR-E contained replicators. In contrast to replication initiation by the intact silencer, initiation by the non-silencer replicator(s) was abolished in an orc2-1 mutant, indicating that these replicators were extremely sensitive to defects in ORC. Remarkably, the activity of one of the non-silencer replicators correlated with reduced silencing; inactivation of these replicators caused by either the orc2-1 mutation or the deletion of flanking sequences enhanced silencing. These data were consistent with a role for the ORC bound to the HMR-E silencer ACS in suppressing the function of neighboring ORC molecules capable of inhibiting silencing, and indicated that differences in ORC-binding sites within HMR itself had profound effects on ORC function. Moreover, replication initiation by natural HMR-E was inefficient, suggesting that closely spaced replicators within HMR contributed to an inhibition of replication initiation.     

A revisionist replicon model for higher eukaryotic genomes

The replicon model devised to explain replication control in bacteria has served as the guiding paradigm in the search for origins of replication in the more complex genomes of eukaryotes. In Saccharomyces cerevisiae, this model has proved to be extremely useful, leading to the identification of specific genetic elements (replicators) and the interacting initiator proteins that activate them. However, replication control in organisms ranging from Schizosaccharomyces pombe to mammals is far more fluid: only a small number of origins seem to represent classic replicators, while the majority correspond to zones of inefficient, closely spaced start sites none of which are indispensable for origin activity. In addition, it is apparent that the epigenetic state of a given sequence largely determines its ability to be used as a replication initiation site. These conclusions were arrived at over a period of three decades, and required the development of several novel replicon mapping techniques, as well as new ways of examining the chromatin architecture of any sequence of interest. Recently, methods have been elaborated for isolating all of the active origins in the genomes of higher eukaryotes en masse. Microarray analyses and more recent high-throughput sequencing technology will allow all the origins to be mapped onto the chromosomes of any organism whose genome has been sequenced. With the advent of whole-genome studies on gene expression and chromatin composition, the field is now positioned to define both the genetic and epigenetic rules that govern origin activity.     

ORC- and Cdc6-dependent complexes at active and inactive chromosomal replication origins in Saccharomyces cerevisiae.

We have developed a genomic footprinting protocol which allows us to examine protein-DNA interactions at single copy chromosomal origins of DNA replication in the budding yeast Saccharomyces cerevisiae. We show that active replication origins oscillate between two chromatin states during the cell cycle: an origin recognition complex (ORC)-dependent post-replicative state and a Cdc6p-dependent pre-replicative state. Furthermore, we show that both post- and pre-replicative complexes can form efficiently on closely apposed replicators. Surprisingly, ARS301 which is active as an origin on plasmids but not in its normal chromosomal location, forms ORC- and Cdc6p-dependent complexes in both its active and inactive contexts. Thus, although ORC and Cdc6p are essential for initiation, their binding is not sufficient to dictate origin use.     

Interplay between Pleiotropy and Secondary Selection Determines Rise and Fall of Mutators in Stress Response

Mutators are clones whose mutation rate is about two to three orders of magnitude higher than the rate of wild-type clones and their roles in adaptive evolution of asexual populations have been controversial. Here we address this problem by using an ab initio microscopic model of living cells, which combines population genetics with a physically realistic presentation of protein stability and protein-protein interactions. The genome of model organisms encodes replication controlling genes (RCGs) and genes modeling the mismatch repair (MMR) complexes. The genotype-phenotype relationship posits that the replication rate of an organism is proportional to protein copy numbers of RCGs in their functional form and there is a production cost penalty for protein overexpression. The mutation rate depends linearly on the concentration of homodimers of MMR proteins. By simulating multiple runs of evolution of populations under various environmental stresses—stationary phase, starvation or temperature-jump—we find that adaptation most often occurs through transient fixation of a mutator phenotype, regardless of the nature of stress. By contrast, the fixation mechanism does depend on the nature of stress. In temperature jump stress, mutators take over the population due to loss of stability of MMR complexes. In contrast, in starvation and stationary phase stresses, a small number of mutators are supplied to the population via epigenetic stochastic noise in production of MMR proteins (a pleiotropic effect), and their net supply is higher due to reduced genetic drift in slowly growing populations under stressful environments. Subsequently, mutators in stationary phase or starvation hitchhike to fixation with a beneficial mutation in the RCGs, (second order selection) and finally a mutation stabilizing the MMR complex arrives, returning the population to a non-mutator phenotype. Our results provide microscopic insights into the rise and fall of mutators in adapting finite asexual populations.     

Multiple DNA elements in ARS305 determine replication origin activity in a yeast chromosome.

A yeast autonomously replicating sequence, ARS305, shares essential components with a chromosome III replicator, ORI305. Known components include an ARS consensus sequence (ACS) element, presumed to bind the origin recognition complex (ORC), and a broad 3'-flanking sequence which contains a DNA unwinding element. Here linker substitution mutagenesis of ARS305 and analysis of plasmid mitotic stability identified three short sequence elements within the broad 3'-flanking sequence. The major functional element resides directly 3' of the ACS and the two remaining elements reside further downstream, all within non-conserved ARS sequences. To determine the contribution of the elements to replication origin function in the chromosome, selected linker mutations were transplaced into the ORI305 locus and two-dimensional gel electrophoresis was used to analyze replication bubble formation and fork directions. Mutation of the major functional element identified in the plasmid mitotic stability assay inactivated replication origin function in the chromosome. Mutation of each of the two remaining elements diminished both plasmid ARS and chromosomal origin activities to similar levels. Thus multiple DNA elements identified in the plasmid ARS are determinants of replication origin function in the natural context of the chromosome. Comparison with two other genetically defined chromosomal replicators reveals a conservation of functional elements known to bind ORC, but no two replicators are identical in the arrangement of elements downstream of ORC binding elements or in the extent of functional sequences adjacent to the ACS.     

The evolution of replicators

Replicators of interest in chemistry, biology and culture are briefly surveyed from a conceptual point of view. Systems with limited heredity have only a limited evolutionary potential because the number of available types is too low. Chemical cycles, such as the formose reaction, are holistic replicators since replication is not based on the successive addition of modules. Replicator networks consisting of catalytic molecules (such as reflexively autocatalytic sets of proteins, or reproducing lipid vesicles) are hypothetical ensemble replicators, and their functioning rests on attractors of their dynamics. Ensemble replicators suffer from the paradox of specificity: while their abstract feasibility seems to require a high number of molecular types, the harmful effect of side reactions calls for a small system size. No satisfactory solution to this problem is known. Phenotypic replicators do not pass on their genotypes, only some aspects of the phenotype are transmitted. Phenotypic replicators with limited heredity include genetic membranes, prions and simple memetic systems. Memes in human culture are unlimited hereditary, phenotypic replicators, based on language. The typical path of evolution goes from limited to unlimited heredity, and from attractor-based to modular (digital) replicators.     

A model of prebiotic replication: survival of the fittest versus extinction of the unfittest.

A model of prebiotic replication discussed by Szathmary and Maynard Smith (1997) is refined in accordance with Lifson's (1997) theory on the crucial stages in the origin of animate matter. The refined model accounts for two processes associated with replication: (1) Replicators always decompose at some rate. (2) Replicators deplete their reactants at a rate equal to the rate of their own replication. Consequently, replicators and their reactants are linked by a non-linear feedback process that keeps them within limits and leads them toward a steady state. The model suggests that: (a) All competing replicators, including those which replicate at sub-exponential "parabolic" and super-exponential "hyperbolic" rates, are subject to natural selection. (b) Survival/extinction is determined by positive/negative net-replication irrespective of the mechanism of replication. (c) Being fit/unfit is the consequence of survival/extinction rather than its cause. In other words, natural selection and survival of the fit is the outcome of continual extinction of the unfit.     

Eco-evolutionary dynamics,coding structure and the information threshold

Background  

The amount of information that can be maintained in an evolutionary system of replicators is limited by genome length, the number of errors during replication (mutation rate) and various external factors that influence the selection pressure. To date, this phenomenon, known as the information threshold, has been studied (both genotypically and phenotypically) in a constant environment and with respect to maintenance (as opposed to accumulation) of information. Here we take a broader perspective on this problem by studying the accumulation of information in an ecosystem, given an evolvable coding structure. Moreover, our setup allows for individual based as well as ecosystem based solutions. That is, all functions can be performed by individual replicators, or complementing functions can be performed by different replicators. In this setup, where both the ecosystem and the individual genomes can evolve their structure, we study how populations cope with high mutation rates and accordingly how the information threshold might be alleviated.