Dependence of post-segregational killing mediated by Type II restriction-modification systems on the lifetime of restriction endonuclease effective activity.

Plasmid-borne Type II restriction-modification (RM) systems mediate post-segregational killing (PSK). PSK is thought to be caused by the dilution of restriction and modification enzymes during cell division, resulting in accumulation of unmethylated DNA recognition sites and their cleavage by restriction endonucleases. PSK is the likely reason for stabilization of plasmids carrying RM systems in the absence of selection for plasmid maintenance. In this study, we developed a CRISPR interference-based method to eliminate RM-carrying plasmids and study PSK-related phenomena with minimal perturbation to the Escherichia coli host. Plasmids carrying the EcoRV, Eco29kI, and EcoRI RM systems were highly stable, and their loss resulted in SOS response and PSK. In contrast, plasmids carrying the Esp1396I system were poorly stabilized; their loss led to a temporary cessation of growth, followed by full recovery. We demonstrate that this unusual behavior is due to a limited lifetime of the Esp1396I restriction endonuclease activity, which, upon Esp1396I plasmid loss, disappears approximately after two cycles of cell division, i.e., before unmethylated sites appear in significant numbers. Our results indicate that whenever PSK induced by a loss of RM systems, and, possibly, other toxin-antitoxin systems, is considered, the lifetimes of individual system components and the growth rate of host cells shall be taken in account. Mathematical modeling shows, that unlike the situation with classical toxin-antitoxin systems, RM system-mediated PSK is possible when the lifetimes of restriction endonuclease and methyltransferase activities are similar, as long as the toxic restriction endonuclease activity persists for more than two chromosome replication cycles.IMPORTANCEIt is widely accepted that many Type II restriction-modification (RM) systems mediate post-segregational killing (PSK) if plasmids that encode them are lost. In this study, we harnessed an inducible CRISPR-Cas system to remove RM plasmids from Escherichia coli cells to study PSK while minimally perturbing cell physiology. We demonstrate that PSK depends on restriction endonuclease activity lifetime and is not observed when it is less than two replication cycles. We present a mathematical model that explains experimental data and shows that unlike the case of toxin-antitoxin-mediated PSK, the loss of an RM system induced PSK even when the RM enzymes have identical lifetimes.

Fission as a source of variation for group selection.

Without heritable variation, natural selection cannot effect evolutionary change. In the case of group selection, there must be variation in the population of groups. Where does this variation come from? One source of variation is from the stochastic birth-death processes that occur within groups. This is where variation between groups comes from in most mathematical models of group selection. Here we argue that another important source of variation between groups is fission, the (generally random) group-level reproduction where parent groups split into two or more offspring groups. We construct a simple model of the fissioning process with a parameter that controls how much variation is produced among the offspring groups. We then illustrate the effect of that parameter with some examples. In most models of group selection in the literature, no variation is produced during group reproduction events, i.e., groups "clone" themselves when they reproduce. Fission is often a more biologically realistic method of group reproduction, and it can significantly increase the efficacy of group selection.

Interference Requirements of Type III CRISPR-Cas Systems from Thermus thermophilus.

Among the diverse prokaryotic adaptive immunity mechanisms, the Type III CRISPR-Cas systems are the most complex. The multisubunit Type III effectors recognize RNA targets complementary to CRISPR RNAs (crRNAs). Target recognition causes synthesis of cyclic oligoadenylates that activate downstream auxiliary effectors, which affect cell physiology in complex and poorly understood ways. Here, we studied the ability of III-A and III-B CRISPR-Cas subtypes from Thermus thermophilus to interfere with plasmid transformation. We find that for both systems, requirements for crRNA-target complementarity sufficient for interference depend on the target transcript abundance, with more abundant targets requiring shorter complementarity segments. This result and thermodynamic calculations indicate that Type III effectors bind their targets in a simple bimolecular reaction with more extensive crRNA-target base pairing compensating for lower target abundance. Since the targeted RNA used in our work is non-essential for either the host or the plasmid, the results also establish that a certain number of target-bound effector complexes must be present in the cell to interfere with plasmid establishment. For the more active III-A system, we determine the minimal length of RNA-duplex sufficient for interference and show that the position of this minimal duplex can vary within the effector. Finally, we show that the III-A immunity is dependent on the HD nuclease domain of the Cas10 subunit. Since this domain is absent from the III-B system the result implies that the T. thermophilus III-B system must elicit a more efficient cyclic oligoadenylate-dependent response to provide the immunity.

On the evolutionary emergence of predation.

In models for the evolution of predation from initially purely competitive species interactions, the propensity of predation is most often assumed to be a direct consequence of the relative morphological and physiological traits of interacting species. Here we explore a model in which predation ability is an independently evolving phenotypic feature, so that even when the relative morphological or physiological traits allow for predation, predation only occurs if the predation ability of individuals has independently evolved to high enough values. In addition to delineating the conditions for the evolutionary emergence of predation, the model reproduces stationary and non-stationary multilevel food webs with the top predators not necessarily having size superiority.

Maximal ecological diversity exceeds evolutionary diversity in model ecosystems.

Understanding community saturation is fundamental to ecological theory. While investigations of the diversity of evolutionary stable states (ESSs) are widespread, the diversity of communities that have yet to reach an evolutionary endpoint is poorly understood. We use Lotka-Volterra dynamics and trait-based competition to compare the diversity of randomly assembled communities to the diversity of the ESS. We show that, with a large enough founding diversity (whether assembled at once or through sequential invasions), the number of long-time surviving species exceeds that of the ESS. However, the excessive founding diversity required to assemble a saturated community increases rapidly with the dimension of phenotype space. Additionally, traits present in communities resulting from random assembly are more clustered in phenotype space compared to random, although still markedly less ordered than the ESS. By combining theories of random assembly and ESSs we bring a new viewpoint to both the saturation and random assembly literature.

Adaptive diversification and niche packing on rugged fitness landscapes.

Explaining the emergence of diversity and the coexistence of competing types has long been one of the main goals of ecological theory. Rugged fitness landscapes have often been used to explain diversity through the presence of local peaks, or adaptive zones, in the fitness landscape acting as available niches for different species. Alternatively, niche-packing and theories based on limiting similarity describe frequency-dependent selection leading to the organic differentiation of a continuous phenotype space into multiple coexisting types. By combining rugged carrying capacity landscapes with frequency-dependent selection, here we investigate the effects of ruggedness on adaptive diversification and stably maintained diversity. We show that while increased ruggedness often leads to a decreased opportunity for adaptive diversification, it is the shape of the global carrying capacity function, not the local ruggedness, that determines the diversity of the ESS and the total diversity a system can stably maintain.

Cells with stochastically increased methyltransferase to restriction endonuclease ratio provide an entry for bacteriophage into protected cell population.

The action of Type II restriction-modification (RM) systems depends on restriction endonuclease (REase), which cleaves foreign DNA at specific sites, and methyltransferase (MTase), which protects host genome from restriction by methylating the same sites. We here show that protection from phage infection increases as the copy number of plasmids carrying the Type II RM Esp1396I system is increased. However, since increased plasmid copy number leads to both increased absolute intracellular RM enzyme levels and to a decreased MTase/REase ratio, it is impossible to determine which factor determines resistance/susceptibility to infection. By controlled expression of individual Esp1396I MTase or REase genes in cells carrying the Esp1396I system, we show that a shift in the MTase to REase ratio caused by overproduction of MTase or REase leads, respectively, to decreased or increased protection from infection. Consistently, due to stochastic variation of MTase and REase amount in individual cells, bacterial cells that are productively infected by bacteriophage have significantly higher MTase to REase ratios than cells that ward off the infection. Our results suggest that cells with transiently increased MTase to REase ratio at the time of infection serve as entry points for unmodified phage DNA into protected bacterial populations.

Persistence of plasmids targeted by CRISPR interference in bacterial populations.

CRISPR-Cas systems provide prokaryotes with an RNA-guided defense against foreign mobile genetic elements (MGEs) such as plasmids and viruses. A common mechanism by which MGEs avoid interference by CRISPR consists of acquisition of escape mutations in regions targeted by CRISPR. Here, using microbiological, live microscopy and microfluidics analyses we demonstrate that plasmids can persist for multiple generations in some Escherichia coli cell lineages at conditions of continuous targeting by the type I-E CRISPR-Cas system. We used mathematical modeling to show how plasmid persistence in a subpopulation of cells mounting CRISPR interference is achieved due to the stochastic nature of CRISPR interference and plasmid replication events. We hypothesize that the observed complex dynamics provides bacterial populations with long-term benefits due to continuous maintenance of mobile genetic elements in some cells, which leads to diversification of phenotypes in the entire community and allows rapid changes in the population structure to meet the demands of a changing environment.

Evolution of diversity in metabolic strategies.

Understanding the origin and maintenance of biodiversity is a fundamental problem. Many theoretical approaches have been investigating ecological interactions, such as competition, as potential drivers of diversification. Classical consumer-resource models predict that the number of coexisting species should not exceed the number of distinct resources, a phenomenon known as the competitive exclusion principle. It has recently been argued that including physiological tradeoffs in consumer-resource models can lead to violations of this principle and to ecological coexistence of very high numbers of species. Here, we show that these results crucially depend on the functional form of the tradeoff. We investigate the evolutionary dynamics of resource use constrained by tradeoffs and show that if the tradeoffs are non-linear, the system either does not diversify or diversifies into a number of coexisting species that do not exceed the number of resources. In particular, very high diversity can only be observed for linear tradeoffs.

Boom-bust population dynamics increase diversity in evolving competitive communities.

The processes and mechanisms underlying the origin and maintenance of biological diversity have long been of central importance in ecology and evolution. The competitive exclusion principle states that the number of coexisting species is limited by the number of resources, or by the species' similarity in resource use. Natural systems such as the extreme diversity of unicellular life in the oceans provide counter examples. It is known that mathematical models incorporating population fluctuations can lead to violations of the exclusion principle. Here we use simple eco-evolutionary models to show that a certain type of population dynamics, boom-bust dynamics, can allow for the evolution of much larger amounts of diversity than would be expected with stable equilibrium dynamics. Boom-bust dynamics are characterized by long periods of almost exponential growth (boom) and a subsequent population crash due to competition (bust). When such ecological dynamics are incorporated into an evolutionary model that allows for adaptive diversification in continuous phenotype spaces, desynchronization of the boom-bust cycles of coexisting species can lead to the maintenance of high levels of diversity.

Acculturation drives the evolution of intergroup conflict.

Conflict between groups of individuals is a prevalent feature in human societies. A common theoretical explanation for intergroup conflict is that it provides benefits to individuals within groups in the form of reproduction-enhancing resources, such as food, territory, or mates. However, it is not always the case that conflict results from resource scarcity. Here, we show that intergroup conflict can evolve, despite not providing any benefits to individuals or their groups. The mechanism underlying this process is acculturation: the adoption, through coercion or imitation, of the victor's cultural traits. Acculturation acts as a cultural driver (in analogy to meiotic drivers) favoring the transmission of conflict, despite a potential cost to both the host group as a whole and to individuals in that group. We illustrate this process with a two-level model incorporating state-dependent event rates and evolving traits for both individuals and groups. Individuals can become "warriors" who specialize in intergroup conflicts, but are costly otherwise. Additionally, groups are characterized by cultural traits, such as their tendency to engage in conflict with other groups and their tendency for acculturation. We show that, if groups engage in conflicts, group selection will favor the production of warriors. Then, we show that group engagement can evolve if it is associated with acculturation. Finally, we study the coevolution of engagement and acculturation. Our model shows that horizontal transmission of culture between interacting groups can act as a cultural driver and lead to the maintenance of costly behaviors by both individuals and groups.

Natural diversity of CRISPR spacers of Thermus: evidence of local spacer acquisition and global spacer exchange.

We investigated the diversity of CRISPR spacers of Thermus communities from two locations in Italy, two in Chile and one location in Russia. Among the five sampling sites, a total of more than 7200 unique spacers belonging to different CRISPR-Cas systems types and subtypes were identified. Most of these spacers are not found in CRISPR arrays of sequenced Thermus strains. Comparison of spacer sets revealed that samples within the same area (separated by few to hundreds of metres) have similar spacer sets, which appear to be largely stable at least over the course of several years. While at further distances (hundreds of kilometres and more) the similarity of spacer sets is decreased, there are still multiple common spacers in Thermus communities from different continents. The common spacers can be reconstructed in identical or similar CRISPR arrays, excluding their independent appearance and suggesting an extensive migration of thermophilic bacteria over long distances. Several new Thermus phages were isolated in the sampling sites. Mapping of spacers to bacteriophage sequences revealed examples of local acquisition of spacers from some phages and distinct patterns of targeting of phage genomes by different CRISPR-Cas systems. This article is part of a discussion meeting issue 'The ecology and evolution of prokaryotic CRISPR-Cas adaptive immune systems'.

Lily Pollen Tubes Pulse According to a Simple Spatial Oscillator.

Polar growth is a fundamental mode of cell morphogenesis observed in nearly all major groups of organisms. Among polarly growing cells, the angiosperm pollen tubes have emerged as powerful experimental systems in large part because of their oscillatory growth, which provides a window into the network of interactions regulating morphogenesis. Empirical studies of oscillatory pollen tubes have sought to uncover the temporal sequence of cellular and molecular events that constitutes an oscillatory cycle. Here we show that in lily pollen tubes the distance or wavelength (λ = 6.3 ± 1.7 µm) over which an oscillatory cycle unfolds is more robust than the period of oscillation (τ = 39.1 ± 17.6 s) (n = 159 cells). Moreover, the oscillatory cycle is divided into slow and fast phases, with each phase unfolding over precisely one half of the wavelength. Using these observations, we show that a simple spatial bi-oscillator predicts the most common modes of oscillation observed in pollen tubes. These results call into question the traditional view of pollen tube morphogenesis as a temporal succession of cellular events. Space, not time, may be the most natural metric to inteprete the morphogenetic dynamics of these cells.

Towards a mechanistic foundation of evolutionary theory.

Most evolutionary thinking is based on the notion of fitness and related ideas such as fitness landscapes and evolutionary optima. Nevertheless, it is often unclear what fitness actually is, and its meaning often depends on the context. Here we argue that fitness should not be a basal ingredient in verbal or mathematical descriptions of evolution. Instead, we propose that evolutionary birth-death processes, in which individuals give birth and die at ever-changing rates, should be the basis of evolutionary theory, because such processes capture the fundamental events that generate evolutionary dynamics. In evolutionary birth-death processes, fitness is at best a derived quantity, and owing to the potential complexity of such processes, there is no guarantee that there is a simple scalar, such as fitness, that would describe long-term evolutionary outcomes. We discuss how evolutionary birth-death processes can provide useful perspectives on a number of central issues in evolution.

Computing in fish schools.

A model based on shoaling fish suggests how a group can show decision-making properties beyond those of any one individual.