Phenotype Design Space Provides a Mechanistic Framework Relating Molecular Parameters to Phenotype Diversity Available for Selection.

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.

Phenotype-centric modeling for rational metabolic engineering.

Phenotype-centric modeling enables a paradigm shift in the analysis of mechanistic models. It brings the focus to a network's biochemical phenotypes and their relationship with measurable traits (e.g.

Mechanistic Modeling of Biochemical Systems without A Priori Parameter Values Using the Design Space Toolbox v.3.0.

Mechanistic models of biochemical systems provide a rigorous description of biological phenomena. They are indispensable for making predictions and elucidating biological design principles. To date, mathematical analysis and characterization of these models encounter a bottleneck consisting of large numbers of unknown parameter values.

TaxisPy: A Python-based software for the quantitative analysis of bacterial chemotaxis.

Several species of bacteria are able to modify their swimming behavior in response to chemical attractants or repellents. Methods for the quantitative analysis of bacterial chemotaxis such as quantitative capillary assays are tedious and time-consuming. Computer-based video analysis of swimming bacteria represents a valuable method to directly assess their chemotactic response.

Molecular Mechanisms Driving Switch Behavior in Xylem Cell Differentiation.

Plant xylem cells conduct water and mineral nutrients. Although most plant cells are totipotent, xylem cells are unusual and undergo terminal differentiation. Many genes regulating this process are well characterized, including the Vascular-related NAC Domain 7 (VND7), MYB46, and MYB83 transcription factors, which are proposed to act in interconnected feedforward loops (FFLs).

Phenotype-centric modeling for elucidation of biological design principles.

A recently developed 'phenotype-centric' modeling strategy combines four innovations with the potential to advance our understanding of complex biological systems: (1) a rigorous mathematical definition of biochemical phenotypes, (2) a method for enumerating the phenotypic repertoire based on the biomolecular network architecture, (3) an integrated suite of computational algorithms for the efficient prediction of parameter values and analysis of the phenotypic repertoire, and (4) a user-focused environment for navigating the resulting space of phenotypes and identifying biologically relevant features and system design principles. These innovations will facilitate deterministic and stochastic simulations that require parameter values, will accelerate both hypothesis discrimination in systems biology and the design cycle in synthetic biology. Here we first review the fundamental definition of biochemical phenotype that enables this new modeling strategy and give an overview of the strategy using a simple system from phage λ to provide an example of a global design principle.

Rapid Discrimination Among Putative Mechanistic Models of Biochemical Systems.

An overarching goal in molecular biology is to gain an understanding of the mechanistic basis underlying biochemical systems. Success is critical if we are to predict effectively the outcome of drug treatments and the development of abnormal phenotypes. However, data from most experimental studies is typically noisy and sparse.

Design Space Toolbox V2: Automated Software Enabling a Novel Phenotype-Centric Modeling Strategy for Natural and Synthetic Biological Systems.

Mathematical models of biochemical systems provide a means to elucidate the link between the genotype, environment, and phenotype. A subclass of mathematical models, known as mechanistic models, quantitatively describe the complex non-linear mechanisms that capture the intricate interactions between biochemical components. However, the study of mechanistic models is challenging because most are analytically intractable and involve large numbers of system parameters.

Unrelated toxin-antitoxin systems cooperate to induce persistence.

Persisters are drug-tolerant bacteria that account for the majority of bacterial infections. They are not mutants, rather, they are slow-growing cells in an otherwise normally growing population. It is known that the frequency of persisters in a population is correlated with the number of toxin-antitoxin systems in the organism.

Design principles of a conditional futile cycle exploited for regulation.

In this report, we characterize the design principles of futile cycling in providing rapid adaptation by regulatory proteins that act as environmental sensors. In contrast to the energetically wasteful futile cycles that are avoided in metabolic pathways, here we describe a conditional futile cycle exploited for a regulatory benefit. The FNR (fumarate and nitrate reduction) cycle in Escherichia coli operates under two regimes - a strictly futile cycle in the presence of O2 and as a pathway under anoxic conditions.

Elucidating the genotype-phenotype map by automatic enumeration and analysis of the phenotypic repertoire.

Background: The gap between genotype and phenotype is filled by complex biochemical systems most of which are poorly understood. Because these systems are complex, it is widely appreciated that quantitative understanding can only be achieved with the aid of mathematical models. However, formulating models and measuring or estimating their numerous rate constants and binding constants is daunting.

Evolution of a genome-encoded bias in amino acid biosynthetic pathways is a potential indicator of amino acid dynamics in the environment.

Overcoming the stress of starvation is one of an organism's most challenging phenotypic responses. Those organisms that frequently survive the challenge, by virtue of their fitness, will have evolved genomes that are shaped by their specific environments. Understanding this genotype-environment-phenotype relationship at a deep level will require quantitative predictive models of the complex molecular systems that link these aspects of an organism's existence.

Strategy revealing phenotypic differences among synthetic oscillator designs.

Considerable progress has been made in identifying and characterizing the component parts of genetic oscillators, which play central roles in all organisms. Nonlinear interaction among components is sufficiently complex that mathematical models are required to elucidate their elusive integrated behavior. Although natural and synthetic oscillators exhibit common architectures, there are numerous differences that are poorly understood.

Relative amino acid composition signatures of organisms and environments.

Background: Identifying organism-environment interactions at the molecular level is crucial to understanding how organisms adapt to and change the chemical and molecular landscape of their habitats. In this work we investigated whether relative amino acid compositions could be used as a molecular signature of an environment and whether such a signature could also be observed at the level of the cellular amino acid composition of the microorganisms that inhabit that environment.

Methodologies/principal Findings: To address these questions we collected and analyzed environmental amino acid determinations from the literature, and estimated from complete genomic sequences the global relative amino acid abundances of organisms that are cognate to the different types of environment.

Phenotypic deconstruction of gene circuitry.

It remains a challenge to obtain a global perspective on the behavioral repertoire of complex nonlinear gene circuits. In this paper, we describe a method for deconstructing complex systems into nonlinear sub-systems, based on mathematically defined phenotypes, which are then represented within a system design space that allows the repertoire of qualitatively distinct phenotypes of the complex system to be identified, enumerated, and analyzed. This method efficiently characterizes large regions of system design space and quickly generates alternative hypotheses for experimental testing.

Molecular mechanisms of multiple toxin-antitoxin systems are coordinated to govern the persister phenotype.

Toxin-antitoxin systems are ubiquitous and have been implicated in persistence, the multidrug tolerance of bacteria, biofilms, and, by extension, most chronic infections. However, their purpose, apparent redundancy, and coordination remain topics of debate. Our model relates molecular mechanisms to population dynamics for a large class of toxin-antitoxin systems and suggests answers to several of the open questions.

A bistable hysteretic switch in an activator-repressor regulated restriction-modification system.

Restriction-modification (RM) systems are extremely widespread among bacteria and archaea, and are often specified by mobile genetic elements. In type II RM systems, where the restriction endonuclease (REase) and protective DNA methyltransferase (MTase) are separate proteins, a major regulatory challenge is delaying expression of the REase relative to the MTase after RM genes enter a new host cell. Basic understanding of this regulation is available for few RM systems, and detailed understanding for none.

Regulatory design governing progression of population growth phases in bacteria.

It has long been noted that batch cultures inoculated with resting bacteria exhibit a progression of growth phases traditionally labeled lag, exponential, pre-stationary and stationary. However, a detailed molecular description of the mechanisms controlling the transitions between these phases is lacking. A core circuit, formed by a subset of regulatory interactions involving five global transcription factors (FIS, HNS, IHF, RpoS and GadX), has been identified by correlating information from the well- established transcriptional regulatory network of Escherichia coli and genome-wide expression data from cultures in these different growth phases.

Design of the lac gene circuit revisited.

The lactose (lac) operon of Escherichia coli serves as the paradigm for gene regulation, not only for bacteria, but also for all biological systems from simple phage to humans. The details of the systems may differ, but the key conceptual framework remains, and the original system continues to reveal deeper insights with continued experimental and theoretical study. Nearly as long lasting in impact as the pivotal work of Jacob and Monod is the classic experiment of Novick and Weiner in which they demonstrated all-or-none gene expression in response to an artificial inducer.

Biomedical engineering strategies in system design space.

Modern systems biology and synthetic bioengineering face two major challenges in relating properties of the genetic components of a natural or engineered system to its integrated behavior. The first is the fundamental unsolved problem of relating the digital representation of the genotype to the analog representation of the parameters for the molecular components. For example, knowing the DNA sequence does not allow one to determine the kinetic parameters of an enzyme.

Phenotypic repertoire of the FNR regulatory network in Escherichia coli.

The FNR protein in Escherichia coli is an O(2) sensor that modifies global gene expression to adapt the cell to anaerobic growth. Regulation of FNR involves continuous cycling of the protein between its active and inactive states under aerobic conditions without apparent function. This raises the question of what benefit to the overall life cycle might compensate for the cost of cycling and reveals that the role of this process is poorly understood.

Relating mutant genotype to phenotype via quantitative behavior of the NADPH redox cycle in human erythrocytes.

Background: The NADPH redox cycle plays a key role in antioxidant protection of human erythrocytes. It consists of two enzymes: glucose-6-phosphate dehydrogenase (G6PD) and glutathione reductase. Over 160 G6PD variants have been characterized and associated with several distinct clinical manifestations.

Automated construction and analysis of the design space for biochemical systems.

Motivation: Our recent work introduced a generic method to construct the design space of biochemical systems: a representation of the relationships between system parameters, environmental variables and phenotypic behavior. In design space, the qualitatively distinct phenotypes of a biochemical system can be identified, counted, analyzed and compared. Boundaries in design space indicate a transition between phenotypic behaviors and can be used to measure a system's tolerance to large changes in parameters.

Regulation of aerobic-to-anaerobic transitions by the FNR cycle in Escherichia coli.

The FNR (fumarate nitrate reduction) protein plays a central role in the global oxygen response of a variety of bacteria. In Escherichia coli, FNR is the master transcriptional regulator of the transition between aerobic and anaerobic growth. Regulation of FNR is achieved by cycling the molecule between three states in a process dependent on oxygen.

Qualitatively distinct phenotypes in the design space of biochemical systems.

Although characterization of the genotype has undergone revolutionary advances as a result of the successful genome projects, the chasm between our understanding of a fully characterized gene sequence and the phenotypic repertoire of the organism is as broad and deep as it was in the pre-genomic era. There are two fundamental unsolved problems that provide the context for the challenges in relating genotype to phenotype. We address one of these and describe a generic method for constructing a system design space in which qualitatively distinct phenotypes can be identified and counted, their relative fitness analyzed and compared, and their tolerance to change measured.