Metastasis is the defining feature of advanced malignancy, yet remains challenging to study in laboratory environments. Here we describe a high-throughput zebrafish system for comprehensive, in vivo assessment of metastatic biology. First, we generated several stable cell lines from melanomas of transgenic mitfa-BRAFV600E;p53−/− fish. We then transplanted the melanoma cells into the transparent casper strain to enable highly quantitative measurement of the metastatic process at single cell resolution. Using computational image analysis of the resulting metastases, we generated a metastasis score, μ, that can be applied to quantitative comparison of metastatic capacity between experimental conditions. Furthermore, image analysis also provided estimates of the frequency of metastasis-initiating cells (~1/120,000 cells). Finally, we determined that the degree of pigmentation is a key feature defining cells with metastatic capability. The small size and rapid generation of progeny combined with superior imaging tools make zebrafish ideal for unbiased high-throughput investigations of cell-intrinsic or microenvironmental modifiers of metastasis. The approaches described here are readily applicable to other tumor types and thus serve to complement studies also employing murine and human cell culture systems.

Cellular plasticity is a state in which cancer cells exist along a reversible phenotypic spectrum, and underlies key traits such as drug resistance and metastasis. Melanoma plasticity is linked to phenotype switching, where the microenvironment induces switches between invasive/MITFLO versus proliferative/MITFHI states. Since MITF also induces pigmentation, we hypothesize that macrometastatic success should be favoured by microenvironments that induce a MITFHI/differentiated/proliferative state. Zebrafish imaging demonstrates that after extravasation, melanoma cells become pigmented and enact a gene expression program of melanocyte differentiation. We screened for microenvironmental factors leading to phenotype switching, and find that EDN3 induces a state that is both proliferative and differentiated. CRISPR-mediated inactivation of EDN3, or its synthetic enzyme ECE2, from the microenvironment abrogates phenotype switching and increases animal survival. These results demonstrate that after metastatic dissemination, the microenvironment provides signals to promote phenotype switching and provide proof that targeting tumour cell plasticity is a viable therapeutic opportunity.

Many Proteobacteria use acyl-homoserine lactone (AHL)-mediated quorum sensing to activate the production of antibiotics at high cell density. Extracellular factors like antibiotics can be considered public goods shared by individuals within a group. Quorum-sensing control of antibiotic production may be important for protecting a niche or competing for limited resources in mixed bacterial communities. To begin to investigate the role of quorum sensing in interspecies competition, we developed a dual-species co-culture model using the soil saprophytes Burkholderia thailandensis (Bt) and Chromobacterium violaceum (Cv). These bacteria require quorum sensing to activate the production of antimicrobial factors that inhibit growth of the other species. We demonstrate that quorum-sensing-dependent antimicrobials can provide a competitive advantage to either Bt or Cv by inhibiting growth of the other species in co-culture. Although the quorum-sensing signals differ for each species, we show that the promiscuous signal receptor encoded by Cv can sense signals produced by Bt, and that this ability to eavesdrop on Bt can provide Cv an advantage in certain situations. We use an in silico approach to investigate the effect of eavesdropping in competition, and show conditions where early activation of antibiotic production resulting from eavesdropping can promote competitiveness. Our work supports the idea that quorum sensing is important for interspecies competition and that promiscuous signal receptors allow eavesdropping on competitors in mixed microbial habitats.

Bacteriophage are voracious predators of bacteria and a major determinant in shaping bacterial life strategies. Many phage species are virulent, meaning that infection leads to certain death of the host and immediate release of a large batch of phage progeny. Despite this apparent voraciousness, bacteria have stably coexisted with virulent phages for eons. Here, using individual-based stochastic spatial models, we study the conditions for achieving coexistence on the edge between two habitats, one of which is a bacterial refuge with conditions hostile to phage whereas the other is phage friendly. We show how bacterial density-dependent, or quorum-sensing, mechanisms such as the formation of biofilm can produce such refuges and edges in a self-organized manner. Coexistence on these edges exhibits the following properties, all of which are observed in real phage–bacteria ecosystems but difficult to achieve together in nonspatial ecosystem models: ( i ) highly efficient virulent phage with relatively long lifetimes, high infection rates and large burst sizes; ( ii ) large, stable, and high-density populations of phage and bacteria; ( iii ) a fast turnover of both phage and bacteria; and ( iv ) stability over evolutionary timescales despite imbalances in the rates of phage vs. bacterial evolution.

Many bacteria secrete compounds which act as public goods. Such compounds are often under quorum sensing (QS) regulation, yet it is not understood exactly when bacteria may gain from having a public good under QS regulation. Here, we show that the optimal public good production rate per cell as a function of population size (the optimal production curve, OPC) depends crucially on the cost and benefit functions of the public good and that the OPC will fall into one of two categories: Either it is continuous or it jumps from zero discontinuously at a critical population size. If, e.g., the public good has accelerating returns and linear cost, then the OPC is discontinuous and the best strategy thus to ramp up production sharply at a precise population size. By using the example of public goods with accelerating and diminishing returns (and linear cost) we are able to determine how the two different categories of OPSs can best be matched by production regulated through a QS signal feeding back on its own production. We find that the optimal QS parameters are different for the two categories and specifically that public goods which provide accelerating returns, call for stronger positive signal feedback.

Virulent phages and their bacterial hosts represent an unusual sort of predator-prey system where each time a prey is eaten, hundreds of new predators are born. It is puzzling how, despite the apparent effectiveness of the phage predators, they manage to avoid driving their bacterial prey to extinction. Here we consider a phage-bacterial ecosystem on a two-dimensional (2-d) surface and show that homogeneous space in itself enhances coexistence. We analyze different behavioral mechanisms that can facilitate coexistence in a spatial environment. For example, we find that when the latent times of the phage are allowed to evolve, selection favors "mediocre killers," since voracious phage rapidly deplete local resources and go extinct. Our model system thus emphasizes the differences between short-term proliferation and long-term ecosystem sustainability.The replication strategies of phages fall into two major categories: virulent and temperate. A temperate phage has the ability to integrate its DNA into the host chromosome, where it is then replicated along with the bacterial DNA during cell division. This strategy allows the phage to slow down or completely stop exploitation of the bacteria, thus reducing the risk of driving its host to extinction. A virulent phage lacks this ability, and it is not fully understood how they manage to coexist with their bacterial prey (4, 19). Consider, for example, the highly effective T4 phage. For the sake of argument, let us assume a burst size of 100 offspring upon lysis. On average, not more than a single phage out of each burst of 100 should survive to infect another bacterium, or else the phage would rapidly outgrow the bacteria and drive them to extinction. The half-life (t 1/2 ) of a free T4 phage particle has been measured to be approximately 10 days in LB at 37°C (6). Therefore, on average, at least t 1/2 ϫ log 2 (100) Ϸ 2 months should pass between infections to prevent runaway phage growth-a time span that seems highly unreasonable for many of the environments where phage and bacteria interact, such as soil or biofilm. Even a more considered calculation, inserting the above half-life measurement into more realistic Lotka-Volterra-like predator-prey models (9) does not change the conclusion that T4 and other virulent phages appear to be far too effective predators for coexistence to be feasible. It is, however, an undisputed fact that virulent phages and bacteria have coexisted for eons and do so still, everywhere around us and inside us. One possible explanation for this puzzle is that bacteria constantly evolve resistance to existing phages and that the phages evolve to attack resistant bacteria in a continuous arms race. This "Red Queen" argument (23) has, however, been criticized on the grounds that the rates of evolution of phages and bacteria are not symmetric (17, 12). Recent measurements support this: in soil, phages appear to be "ahead of the bacteria in the coevolutionary arms race" (24). We therefore wish to explore mechanisms other than bacterial resistance ...

Bacteria are highly social organisms that communicate via signaling molecules, move collectively over surfaces and make biofilm communities. Nonetheless, our main line of defense against pathogenic bacteria consists of antibiotics – drugs that target individual-level traits of bacterial cells and thus, regrettably, select for resistance against their own action. A possible solution lies in targeting the mechanisms by which bacteria interact with each other within biofilms. The emerging field of microbial social evolution combines molecular microbiology with evolutionary theory to dissect the molecular mechanisms and the evolutionary pressures underpinning bacterial sociality. This exciting new research can ultimately lead to new therapies against biofilm infections that exploit evolutionary cheating or the trade-off between biofilm formation and dispersal.

Graphical Abstract Highlights d p120ctn expression segregates pancreatic progenitors into fate determining niches d p120ctn-low progenitors segregate into peripheral domains and form pro-acinar tips d p120ctn-high progenitors remain in the trunk to become duct and endocrine cells d p120ctn downregulation in endocrine cells drives delamination