Population Dynamics In A Model Closed Ecosystem

For almost any species in any environment, it is nearly impossible to predict its fitness from molecular knowledge. If fitness is not to be a mere tautology, reproducible measurements of the survival and reproduction of populations are needed over many generations. Laboratory microbial ecosystems afford the short time and length scales required for such measurements. Their conventional implementations, batch cultures with period refreshment of growth medium or chemostats with continuous refreshment, have a number of disadvantages, such as the introduction of additional frequencies, selection for surface growth and the distortion of chemical interactions. In closed ecosystems free energy is instead supplied as light, allowing for simpler, replicable protocols and a consistent interpretation of interactions, independent of their mode or timescale. Here, I describe a model closed ecosystem consisting of three singlecelled microbes, Escherichia coli, Chlamydomonas reinhardtii and Tetrahymena thermophila and show that these species can coexist for hundreds of days under closure. Using a custom built in situ fluorescence microscopy set up, the densities of these three species can be measured automatically and noninvasively over months with low classification error and large dynamical range. When kept under identical boundary conditions, these ecosystems reproducibly diverge in composition, with characteristic divergence times of ~20 days for T. thermophila, ~40 days for the other two species, and an approximately linear increase of an aggregate divergence measure over the first ~60 days. For two ecosystems, densities were measured continuously under constant conditions and their dynamics shown to be nonstationary for all three species \u3e100 days after closure. As a consequence, conventional time series methods assuming stationarity are inadequate and wavelet analysis is proposed as an alternative. Species-species interactions are further investigated using oscillations in illumination intensity. Densities of C. reinhardtii and, surprisingly, E. coli respond to modest perturbations of light intensity. Variation of the modulation frequency strongly implicates the circadian clock of C. reinhardtii in its response. The nonlinearity of the E. coli response suggests that it depends on C. reinhardtii density or spatial distribution rather than directly responds to the modulation of illumination. Further improvements in the detection of interactions are proposed

Contingency and Statistical Laws in Replicate Microbial Closed Ecosystems

SummaryContingency, the persistent influence of past random events, pervades biology. To what extent, then, is each course of ecological or evolutionary dynamics unique, and to what extent are these dynamics subject to a common statistical structure? Addressing this question requires replicate measurements to search for emergent statistical laws. We establish a readily replicated microbial closed ecosystem (CES), sustaining its three species for years. We precisely measure the local population density of each species in many CES replicates, started from the same initial conditions and kept under constant light and temperature. The covariation among replicates of the three species densities acquires a stable structure, which could be decomposed into discrete eigenvectors, or “ecomodes.” The largest ecomode dominates population density fluctuations around the replicate-average dynamics. These fluctuations follow simple power laws consistent with a geometric random walk. Thus, variability in ecological dynamics can be studied with CES replicates and described by simple statistical laws

Absolute mRNA concentrations from sequence-specific calibration of oligonucleotide arrays

Oligonucleotide microarrays are based on the hybridization of labeled mRNA molecules to short length oligonucleotide probes on a glass surface. Two effects have been shown to affect the raw data: the sequence dependence of the probe hybridization properties and the chemical saturation resulting from surface adsorption processes. We address both issues simultaneously using a physically motivated hybridization model. Based on publicly available calibration data sets, we show that Langmuir adsorption accurately describes GeneChip hybridization, with model parameters that we predict from the sequence composition of the probes. Because these parameters have physical units, we are able to estimate absolute mRNA concentrations in picomolar. Additionally, by accounting for chemical saturation, we substantially reduce the compressive bias of differential expression estimates that normally occurs toward high concentrations

Polymeric peptide pigments with sequence-encoded properties

Melanins are a family of heterogeneous polymeric pigments that provide ultraviolet (UV) light protection, structural support, coloration, and free radical scavenging. Formed by oxidative oligomerization of catecholic small molecules, the physical properties of melanins are influenced by covalent and noncovalent disorder. We report the use of tyrosine-containing tripeptides as tunable precursors for polymeric pigments. In these structures, phenols are presented in a (supra-)molecular context dictated by the positions of the amino acids in the peptide sequence. Oxidative polymerization can be tuned in a sequence-dependent manner, resulting in peptide sequence–encoded properties such as UV absorbance, morphology, coloration, and electrochemical properties over a considerable range. Short peptides have low barriers to application and can be easily scaled, suggesting near-term applications in cosmetics and biomedicine

Microbial population dynamics by digital in-line holographic microscopy

Measurements of population dynamics are ubiquitous in experiments with microorganisms. Studies with microbes elucidating adaptation, selection, and competition rely on measurements of changing populations in time. Despite this importance, quantitative methods for measuring population dynamics microscopically, with high time resolution, across many replicates remain limited. Here we present a new noninvasive method to precisely measure microbial spatiotemporal population dynamics based on digital in-line holographic (DIH) microscopy. Our inexpensive, replicate DIH microscopes imaged hundreds of swimming algae in three dimensions within a volume of several microliters on a time scale of minutes over periods of weeks

Inhibitory action of certain cyclophosphate derivatives of cAMP on cAMP-dependent protein kinases

A series cAMP derivatives with modifications in the adenine, ribose and cyclophosphate moiety were screened for their binding affinity for the two types of cAMP-binding sites in mammalian protein kinase type I. In addition, the activation of the kinase by these analogs was monitored. The binding data indicate that cAMP is bound to both sites in a comparable manner: the adenine appears to have no hydrogen-bond interactions with the binding sites, whereas the ribose may be bound by three hydrogen bonds involving the 2’, 3’ and 5’ positions of cAMP. The binding data are not conclusive about the nature of the interaction with the exocyclic oxygen atoms on phosphorus, though a charge interaction seems to be absent. The cAMP molecule seems to be bound in the syn conformation. The results of activation experiments show that modifications in the adenine and ribose moiety do not affect the maximal activation level, while alteration of the two exocyclic oxygen atoms may result in a reduced maximal activation level and in one case, (Rp)-adenosine 3’,5’-monophosphorothioate [Rp-cAMPS], in total absence of activation even at concentrations at which the analog saturates both binding sites. Since occupancy of the cAMP-binding sites by this derivative apparently did not lead to activation of the enzyme, we examined whether this compound could antagonize the activation by cAMP. Indeed (Rp)-cAMPS was found to inhibit cAMP stimulated kinase activity at concentrations compatible to its binding affinity. Also with mammalian protein kinase type II (Rp)-cAMPS showed antagonistic activity, while with a cAMP-dependent protein kinase from Dictyostelium discoideum partial agonistic activity was observed. Previously a mechanism for activation of protein kinase type I was proposed involving a charge interaction between the equatorial exocyclic oxygen atom and the binding site. This was based on measurements with impure preparations of (Rp)-cAMPS and the Rp and Sp isomers adenosine 3’,5’-monophosphodimethylamidate, cAMPN(CH3)2. The present work using highly purified compounds suggests the absence of a charge interaction, since the uncharged analog (Sp)-cAMPN(CH3)2 activates the kinase effectively. The data seem compatible with an activation model involving the formation of a covalent bond with phosphorus in both cAMP binding sites.