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Two long-standing challenges in theoretical population genetics and evolution are predicting the distribution of phenotype diversity generated by mutation and available for selection, and determining the interaction of mutation, selection and drift to characterize evolutionary equilibria and dynamics. More fundamental for enabling such predictions is the current inability to causally link genotype to phenotype. There are three major mechanistic mappings required for such a linking – genetic sequence to kinetic parameters of the molecular processes, kinetic parameters to biochemical system phenotypes, and biochemical phenotypes to organismal phenotypes. This article introduces a theoretical framework, the Phenotype Design Space (PDS) framework, for addressing these challenges by focusing on the mapping of kinetic parameters to biochemical system phenotypes. It provides a quantitative theory whose key features include (1) a mathematically rigorous definition of phenotype based on biochemical kinetics, (2) enumeration of the full phenotypic repertoire, and (3) functional characterization of each phenotype independent of its context-dependent selection or fitness contributions. This framework is built on Design Space methods that relate system phenotypes to genetically determined parameters and environmentally determined variables. It also has the potential to automate prediction of phenotype-specific mutation rate constants and equilibrium distributions of phenotype diversity in microbial populations undergoing steady-state exponential growth, which provides an ideal reference to which more realistic cases can be compared. Although the framework is quite general and flexible, the details will undoubtedly differ for different functions, organisms and contexts. Here a hypothetical case study involving a small molecular system, a primordial circadian clock, is used to introduce this framework and to illustrate its use in a particular case. The framework is built on fundamental biochemical kinetics. Thus, the foundation is based on linear algebra and reasonable physical assumptions, which provide numerous opportunities for experimental testing and further elaboration to deal with complex multicellular organisms that are currently beyond its scope. The discussion provides a comparison of results from the PDS framework with those from other approaches in theoretical population genetics.

 

ABSTRACT Although alcohols are toxic to many microorganisms, they are good carbon and energy sources for some bacteria, including many pseudomonads. However, most studies that have examined chemosensory responses to alcohols have reported that alcohols are sensed as repellents, which is consistent with their toxic properties. In this study, we examined the chemotaxis of Pseudomonas putida strain F1 to n -alcohols with chain lengths of 1 to 12 carbons. P. putida F1 was attracted to all n -alcohols that served as growth substrates (C 2 to C 12 ) for the strain, and the responses were induced when cells were grown in the presence of alcohols. By assaying mutant strains lacking single or multiple methyl-accepting chemotaxis proteins, the receptor mediating the response to C 2 to C 12 alcohols was identified as McfP, the ortholog of the P. putida strain KT2440 receptor for C 2 and C 3 carboxylic acids. Besides being a requirement for the response to n -alcohols, McfP was required for the response of P. putida F1 to pyruvate, l -lactate, acetate, and propionate, which are detected by the KT2440 receptor, and the medium- and long-chain carboxylic acids hexanoic acid and dodecanoic acid. β-Galactosidase assays of P. putida F1 carrying an mcfP-lacZ transcriptional fusion showed that the mcfP gene is not induced in response to alcohols. Together, our results are consistent with the idea that the carboxylic acids generated from the oxidation of alcohols are the actual attractants sensed by McfP in P. putida F1, rather than the alcohols themselves. IMPORTANCE Alcohols, released as fermentation products and produced as intermediates in the catabolism of many organic compounds, including hydrocarbons and fatty acids, are common components of the microbial food web in soil and sediments. Although they serve as good carbon and energy sources for many soil bacteria, alcohols have primarily been reported to be repellents rather than attractants for motile bacteria. Little is known about how alcohols are sensed by microbes in the environment. We report here that catabolizable n -alcohols with linear chains of up to 12 carbons serve as attractants for the soil bacterium Pseudomonas putida , and rather than being detected directly, alcohols appear to be catabolized to acetate, which is then sensed by a specific cell-surface chemoreceptor protein. 

ABSTRACT Soil bacteria adapt to diverse and rapidly changing environmental conditions by sensing and responding to environmental cues using a variety of sensory systems. Two-component systems are a widespread type of signal transduction system present in all three domains of life and typically are comprised of a sensor kinase and a response regulator. Many two-component systems function by regulating gene expression in response to environmental stimuli. The bacterial chemotaxis system is a modified two-component system with additional protein components and a response that, rather than regulating gene expression, involves behavioral adaptation and results in net movement toward or away from a chemical stimulus. Soil bacteria generally have 20 to 40 or more chemoreceptors encoded in their genomes. To simplify the identification of chemoeffectors (ligands) sensed by bacterial chemoreceptors, we constructed hybrid sensor proteins by fusing the sensor domains of Pseudomonas putida chemoreceptors to the signaling domains of the Escherichia coli NarX/NarQ nitrate sensors. Responses to potential attractants were monitored by β-galactosidase assays using an E. coli reporter strain in which the nitrate-responsive narG promoter was fused to lacZ . Hybrid receptors constructed from PcaY, McfR, and NahY, which are chemoreceptors for aromatic acids, tricarboxylic acid cycle intermediates, and naphthalene, respectively, were sensitive and specific for detecting known attractants, and the β-galactosidase activities measured in E. coli correlated well with results of chemotaxis assays in the native P. putida strain. In addition, a screen of the hybrid receptors successfully identified new ligands for chemoreceptor proteins and resulted in the identification of six receptors that detect propionate. IMPORTANCE Relatively few of the thousands of chemoreceptors encoded in bacterial genomes have been functionally characterized. More importantly, although methyl-accepting chemotaxis proteins, the major type of chemoreceptors present in bacteria, are easily identified bioinformatically, it is not currently possible to predict what chemicals will bind to a particular chemoreceptor. Chemotaxis is known to play roles in biodegradation as well as in host-pathogen and host-symbiont interactions, but many studies are currently limited by the inability to identify relevant chemoreceptor ligands. The use of hybrid receptors and this simple E. coli reporter system allowed rapid and sensitive screening for potential chemoeffectors. The fusion site chosen for this study resulted in a high percentage of functional hybrids, indicating that it could be used to broadly test chemoreceptor responses from phylogenetically diverse samples. Considering the wide range of chemical attractants detected by soil bacteria, hybrid receptors may also be useful as sensitive biosensors.