Functional brain networks: great expectations,hard times and the big leap forward

Many physical and biological systems can be studied using complex network theory, a new statistical physics understanding of graph theory. The recent application of complex network theory to the study of functional brain networks has generated great enthusiasm as it allows addressing hitherto non-standard issues in the field, such as efficiency of brain functioning or vulnerability to damage. However, in spite of its high degree of generality, the theory was originally designed to describe systems profoundly different from the brain. We discuss some important caveats in the wholesale application of existing tools and concepts to a field they were not originally designed to describe. At the same time, we argue that complex network theory has not yet been taken full advantage of, as many of its important aspects are yet to make their appearance in the neuroscience literature. Finally, we propose that, rather than simply borrowing from an existing theory, functional neural networks can inspire a fundamental reformulation of complex network theory, to account for its exquisitely complex functioning mode.     

Origins of life: A comparison of theories and application to Mars

The field of study that deals with the origins of life does not have a consensus for a theory of life's origin. An analysis of the range of theories offered shows that they share some common features that may be reliable predictors when considering the possible origins of life on another planet. The fundamental datum dealing with the origins of life is that life appeared early in the history of the Earth, probably before 3.5 Ga and possibly before 3.8 Ga. What might be called the standard theory (the Oparin-Haldane theory) posits the production of organic molecules on the early Earth followed by chemical reactions that produced increased organic complexity leading eventually to organic life capable of reproduction, mutation, and selection using organic material as nutrients. A distinct class of other theories (panspermia theories) suggests that life was carried to Earth from elsewhere — these theories receive some support from recent work on planetary impact processes. Other alternatives to the standard model suggest that life arose as an inorganic (clay) form and/or that the initial energy source was not organic material but chemical energy or sunlight. We find that the entire range of current theories suggests that liquid water is the quintessential environmental criterion for both the origin and sustenance of life. It is therefore of interest that during the time that life appeared on Earth we have evidence for liquid water present on the surface of Mars.     

An Interpretive Review of the Origin of Life Research

Life appears to be a natural property of matter, but the problem of its origin only arose after early scientists refuted continuous spontaneous generation. There is no chance of life arising ‘all at once’, we need the standard scientific incremental explanation with large numbers of small steps, an approach used in both physical and evolutionary sciences. The necessity for considering both theoretical and experimental approaches is emphasized. After describing basic principles that are available (including the Darwin-Eigen cycle), the search for origins is considered under four main themes. These are the RNA-world hypothesis; potential intermediates between an RNA-world and a modern world via the evolution of protein synthesis and then of DNA; possible alternatives to an RNA-world; and finally the earliest stages from the simple prebiotic systems to RNA. The triplicase/proto-ribosome theory for the origin of the ribosome is discussed where triples of nucleotides are added to a replicating RNA, with the origin of a triplet code well-before protein synthesis begins. The length of the code is suggested to arise from the early development of a ratchet mechanism that overcomes the problem of continued processivity of an RNA-based RNA-polymerase. It is probable that there were precursor stages to RNA with simpler sugars, or just two nucleotides, but we do not yet know of any better alternatives to RNA that were likely to arise naturally. For prebiotic stages (before RNA) a flow-reactor model is suggested to solve metabolism, energy gradients, and compartmentation simultaneously – thus the intense interest in some form of flow reactor. If an autocatalytic cycle could arise in such a system we would be major steps ahead. The most likely physical conditions for the origin of life require further clarification and it is still unclear whether the origin of life is more of an entropy (information) problem (and therefore high temperatures would be detrimental), rather than a kinetic problem (where high temperatures may be advantageous).     

Engineered toxins "zymoxins" are activated by the HCV NS3 protease by removal of an inhibitory protein domain

The synthesis of inactive enzyme precursors, also known as "zymogens," serves as a mechanism for regulating the execution of selected catalytic activities in a desirable time and/or site. Zymogens are usually activated by proteolytic cleavage. Many viruses encode proteases that execute key proteolytic steps of the viral life cycle. Here, we describe a proof of concept for a therapeutic approach to fighting viral infections through eradication of virally infected cells exclusively, thus limiting virus production and spread. Using the hepatitis C virus (HCV) as a model, we designed two HCV NS3 protease-activated "zymogenized" chimeric toxins (which we denote "zymoxins"). In these recombinant constructs, the bacterial and plant toxins diphtheria toxin A (DTA) and Ricin A chain (RTA), respectively, were fused to rationally designed inhibitor peptides/domains via an HCV NS3 protease-cleavable linker. The above toxins were then fused to the binding and translocation domains of Pseudomonas exotoxin A in order to enable translocation into the mammalian cells cytoplasm. We show that these toxins exhibit NS3 cleavage dependent increase in enzymatic activity upon NS3 protease cleavage in vitro. Moreover, a higher level of cytotoxicity was observed when zymoxins were applied to NS3 expressing cells or to HCV infected cells, demonstrating a potential therapeutic window. The increase in toxin activity correlated with NS3 protease activity in the treated cells, thus the therapeutic window was larger in cells expressing recombinant NS3 than in HCV infected cells. This suggests that the "zymoxin" approach may be most appropriate for application to life-threatening acute infections where much higher levels of the activating protease would be expected.     

Life as physics and chemistry: A system view of biology

Cellular life can be viewed as one of many physical natural systems that extract free energy from their environments in the most efficient way, according to fundamental physical laws, and grow until limited by inherent physical constraints. Thus, it can be inferred that it is the efficiency of this process that natural selection acts upon. The consequent emphasis on metabolism, rather than replication, points to a metabolism-first origin of life with the adoption of DNA template replication as a second stage development. This order of events implies a cellular regulatory system that pre-dates the involvement of DNA and might, therefore, be based on the information acquired as peptides fold into proteins, rather than on genetic regulatory networks. Such an epigenetic cell regulatory model, the independent attractor model, has already been proposed to explain the phenomenon of radiation induced genomic instability. Here it is extended to provide an epigenetic basis for the morphological and functional diversity that evolution has yielded, based on natural selection of the most efficient free energy transduction. Empirical evidence which challenges the current genetic basis of cell and molecular biology and which supports the above proposal is discussed.     

Bootstrapping model of the origin of life

The origin of life is analyzed in terms of the self-facilitating aspect of evolution. According to the self-facilitation (or bootstrap) principle the structure of biological systems becomes increasingly suited to effective evolutionary search through the process of evolution. The principle may be extended to primitive collections of polymers with catalytic properties. The origin of the code may be based on a form of bootstrapping evolution. Once a primitive code appeared it could become more sophisticated through a multi-coding mechanism together with classical Darwinian mechanisms. The bootstrapping principle is also formulated in terms of the fluctuation-instability framework of Prigogine, Nicolis, and Babloyantz. Primitive collections of polymers accumulate evolution-enhancing redundancies which tend to reduce the extent to which they change when destabilized by fluctuation, but which increase the chances that these changes will lead to new predominant regimes. We show that the bootstrapping of evolutionary amenability is accompanied by the accumulation of a thermodynamic load, that is, a free energy cost which reduces the mechanistic efficiency. The bootstrapping effect suggests that the origin of life is most fruitfully approached as a long process during which the capacity to evolve facilitates itself in a step by step fashion rather than as a series of low probability events.     

Tests of continuum theories as models of ion channels. I. Poisson-Boltzmann theory versus Brownian dynamics

Continuum theories of electrolytes are widely used to describe physical processes in various biological systems. Although these are well-established theories in macroscopic situations, it is not clear from the outset that they should work in small systems whose dimensions are comparable to or smaller than the Debye length. Here, we test the validity of the mean-field approximation in Poisson-Boltzmann theory by comparing its predictions with those of Brownian dynamics simulations. For this purpose we use spherical and cylindrical boundaries and a catenary shape similar to that of the acetylcholine receptor channel. The interior region filled with electrolyte is assumed to have a high dielectric constant, and the exterior region representing protein a low one. Comparisons of the force on a test ion obtained with the two methods show that the shielding effect due to counterions is overestimated in Poisson-Boltzmann theory when the ion is within a Debye length of the boundary. As the ion gets closer to the boundary, the discrepancy in force grows rapidly. The implication for membrane channels, whose radii are typically smaller than the Debye length, is that Poisson-Boltzmann theory cannot be used to obtain reliable estimates of the electrostatic potential energy and force on an ion in the channel environment.     

Biocomplexity in a highly migratory pelagic marine fish, Atlantic herring

The existence of biologically differentiated populations has been credited with a major role in conferring sustainability and in buffering overall productivity of anadromous fish population complexes where evidence for spatial structure is uncontroversial. Here, we describe evidence of correlated genetic and life history (spawning season linked to spawning location) differentiation in an abundant and highly migratory pelagic fish, Atlantic herring, Clupea harengus, in the North Sea (NS) and adjacent areas. The existence of genetically and phenotypically diverse stocks in this region despite intense seasonal mixing strongly implicates natal homing in this species. Based on information from genetic markers and otolith morphology, we estimate the proportional contribution by NS, Skagerrak (SKG) and Kattegat and western Baltic (WBS) fish to mixed aggregations targeted by the NS fishery. We use these estimates to identify spatial and temporal differences in life history (migratory behaviour) and habitat use among genetically differentiated migratory populations that mix seasonally. Our study suggests the existence of more complex patterns of intraspecific diversity than was previously recognized. Sustainability may be compromised if such complex patterns are reduced through generalized management (e.g. area closures) that overlooks population differences in spatial use throughout the life cycle.     

Organohalide respiration in pristine environments: implications for the natural halogen cycle

Halogenated organic compounds, also termed organohalogens, were initially considered to be of almost exclusively anthropogenic origin. However, over 5000 naturally synthesized organohalogens are known today. This has also fuelled the hypothesis that the natural and ancient origin of organohalogens could have primed development of metabolic machineries for their degradation, especially in microorganisms. Among these, a special group of anaerobic microorganisms was discovered that could conserve energy by reducing organohalogens as terminal electron acceptor in a process termed organohalide respiration. Originally discovered in a quest for biodegradation of anthropogenic organohalogens, these organohalide‐respiring bacteria (OHRB) were soon found to reside in pristine environments, such as the deep subseafloor and Arctic tundra soil with limited/no connections to anthropogenic activities. As such, accumulating evidence suggests an important role of OHRB in local natural halogen cycles, presumably taking advantage of natural organohalogens. In this minireview, we integrate current knowledge regarding the natural origin and occurrence of industrially important organohalogens and the evolution and spread of OHRB, and describe potential implications for natural halogen and carbon cycles.     

The formalism of fractal aggregation phenomena of colloidal drug delivery systems

Classical Newtonian Physics and Euclidean Geometry are currently used to describe biological phenomena and the processes of drug formulation, which are characterized by homogeneity and linearity. On the other hand, at the mesoscopic level, the principles and the laws of physics are quite different from the Classical Newtonian Physics and Euclidean approach especially at nanoscale dimension. The investigation of the aggregation process of liposomes is of paramount importance due to their applications in pharmaceutical nanotechnology as drug delivery systems and as membrane models, in biosciences. The physical stability and the aggregation process of colloidal systems as well as the surface phenomena are described using the Derjaguin-Landau-Verwey-Overbeek (DLVO) theory. The elucidation of the dimensionality of liposome aggregates obeys the fractal approach because the aggregation phenomena are irreversible. This approach can be correlated with the extended DLVO theory, which includes the hydration energy, too.     

Insights into the product release mechanism of dengue virus NS3 helicase

The non-structural protein 3 helicase (NS3h) is a multifunctional protein that is critical in RNA replication and other stages in the flavivirus life cycle. NS3h uses energy from ATP hydrolysis to translocate along single stranded nucleic acid and to unwind double stranded RNA. Here we present a detailed mechanistic analysis of the product release stage in the catalytic cycle of the dengue virus (DENV) NS3h. This study is based on a combined experimental and computational approach of product-inhibition studies and free energy calculations. Our results support a model in which the catalytic cycle of ATP hydrolysis proceeds through an ordered sequential mechanism that includes a ternary complex intermediate (NS3h-Pi-ADP), which evolves releasing the first product, phosphate (Pi), and subsequently ADP. Our results indicate that in the product release stage of the DENV NS3h a novel open-loop conformation plays an important role that may be conserved in NS3 proteins of other flaviviruses as well.     

Thumb inhibitor binding eliminates functionally important dynamics in the hepatitis C virus RNA polymerase

Hepatitis C virus (HCV) has infected almost 200 million people worldwide, typically causing chronic liver damage and severe complications such as liver failure. Currently, there are few approved treatments for viral infection. Thus, the HCV RNA‐dependent RNA polymerase (gene product NS5B) has emerged as an important target for small molecule therapeutics. Potential therapeutic agents include allosteric inhibitors that bind distal to the enzyme active site. While their mechanism of action is not conclusively known, it has been suggested that certain inhibitors prevent a conformational change in NS5B that is crucial for RNA replication. To gain insight into the molecular origin of long‐range allosteric inhibition of NS5B, we employed molecular dynamics simulations of the enzyme with and without an inhibitor bound to the thumb domain. These studies indicate that the presence of an inhibitor in the thumb domain alters both the structure and internal motions of NS5B. Principal components analysis identified motions that are severely attenuated by inhibitor binding. These motions may have functional relevance by facilitating interactions between NS5B and RNA template or nascent RNA duplex, with presence of the ligand leading to enzyme conformations with narrower and thus less accessible RNA binding channels. This study provides the first evidence for a mechanistic basis of allosteric inhibition in NS5B. Moreover, we present evidence that allosteric inhibition of NS5B results from intrinsic features of the enzyme free energy landscape, suggesting a common mechanism for the action of diverse allosteric ligands. Proteins 2013. © 2012 Wiley Periodicals, Inc.     

Parvovirus initiator protein NS1 and RPA coordinate replication fork progression in a reconstituted DNA replication system

We show here that the DNA helicase activity of the parvoviral initiator protein NS1 is highly directional, binding to the single strand at a recessed 5' end and displacing the other strand while progressing in a 3'-to-5' direction on the bound strand. NS1 and a cellular site-specific DNA binding factor, PIF, also known as glucocorticoid modulating element binding protein, bind to the left-end minimal replication origin of minute virus of mice, forming a ternary complex. In this complex, NS1 is activated to nick one DNA strand, becoming covalently attached to the 5' end of the nick in the process and providing a 3' OH for priming DNA synthesis. In this situation, the helicase activity of NS1 did not displace the nicked strand, but the origin duplex was distorted by the NS1-PIF complex, as assayed by its sensitivity to KMnO(4) oxidation, and a stretch of about 14 nucleotides on both strands of the nicked origin underwent limited unwinding. Addition of Escherichia coli single-stranded DNA binding protein (SSB) did not lead to further unwinding. However, addition of recombinant human single-stranded DNA binding protein (RPA) to the initiation reaction catalyzed extensive unwinding of the nicked origin, suggesting that RPA may be required to form a functional replication fork. Accordingly, the unwinding mediated by NS1 and RPA promoted processive leading-strand synthesis catalyzed by recombinant human DNA polymerase delta, PCNA, and RFC, using the minimal left-end origin cloned in a plasmid as a template. The requirement for RPA, rather than SSB, in the unwinding reaction indicated that specific NS1-RPA protein interactions were formed. NS1 was tested by enzyme-linked immunosorbent assay for binding to two- or three-subunit RPA complexes expressed from recombinant baculoviruses. NS1 efficiently bound each of the baculovirus-expressed complexes, indicating that the small subunit of RPA is not involved in specific NS1 binding. No NS1 interactions were observed with E. coli SSB or other proteins included as controls.     

An open system framework for integrating critical zone structure and function

The ??critical zone?? includes the coupled earth surface systems of vegetation, regolith and groundwater that are essential to sustaining life on the planet. The function of this zone is the result of complex interactions among physical, chemical and biological processes and understanding these interactions remains a major challenge to earth system sciences. Here we develop an integrated framework based on thermodynamic theory to characterize the critical zone as a system open to energy and mass fluxes that are forced by radiant, geochemical, and elevational gradients. We derive a statement that demonstrates the relative importance of solar radiation, water, carbon, and physical/chemical denudation mass fluxes to the critical zone energy balance. Within this framework we use rates of effective energy and mass transfer [EEMT; W m^?2] to quantify the relevant flux-gradient relations. Synthesis of existing data demonstrates that variation in energetics associated with primary production and effective precipitation explains substantial variance in critical zone structure and function. Furthermore, we observe threshold behavior in systems that transition to primary production predominance of the energy flux term. The proposed framework provides a first order approximation of non-linearity in critical zone processes that may be coupled with physical and numerical models to constrain landscape evolution.     

Attenuation of equine influenza viruses through truncations of the NS1 protein

Equine influenza is a common disease of the horse, causing significant morbidity worldwide. Here we describe the establishment of a plasmid-based reverse genetics system for equine influenza virus. Utilizing this system, we generated three mutant viruses encoding carboxy-terminally truncated NS1 proteins. We have previously shown that a recombinant human influenza virus lacking the NS1 gene (delNS1) could only replicate in interferon (IFN)-incompetent systems, suggesting that the NS1 protein is responsible for IFN antagonist activity. Contrary to previous findings with human influenza virus, we found that in the case of equine influenza virus, the length of the NS1 protein did not correlate with the level of attenuation of that virus. With equine influenza virus, the mutant virus with the shortest NS1 protein turned out to be the least attenuated. We speculate that the basis for attenuation of the equine NS1 mutant viruses generated is related to their level of NS1 protein expression. Our findings show that the recombinant mutant viruses are impaired in their ability to inhibit IFN production in vitro and they do not replicate as efficiently as the parental recombinant strain in embryonated hen eggs, in MDCK cells, or in vivo in a mouse model. Therefore, these attenuated mutant NS1 viruses may have potential as candidates for a live equine influenza vaccine.     

Sedimentation Patterns of Rapidly Reversible Protein Interactions

The transport behavior of macromolecular mixtures with rapidly reversible complex formation is of great interest in the study of protein interactions by many different methods. Complicated transport patterns arise even for simple bimolecular reactions, when all species exhibit different migration velocities. Although partial differential equations are available to describe the spatial and temporal evolution of the interacting system given particular initial conditions, a general overview of the phase behavior of the systems in parameter space has not yet been reported. In the case of sedimentation of two-component mixtures, this study presents simple analytical solutions that solve the underlying equations in the diffusion-free limit previously subject to Gilbert-Jenkins theory. The new expressions describe, with high precision, the average sedimentation coefficients and composition of each boundary, which allow the examination of features of the whole parameter space at once, and may be used for experimental design and robust analysis of experimental boundary patterns to derive the stoichiometry and affinity of the complex. This study finds previously unrecognized features, including a phase transition between boundary patterns. The model reveals that the time-average velocities of all components in the reaction mixture must match—a condition that suggests an intuitive physical picture of an effective particle of the coupled cosedimentation of an interacting system. Adding to the existing numerical solutions of the relevant partial differential equations, the effective particle model provides physical insights into the relationships of the parameters that govern sedimentation patterns.     

Triadic conceptual structure of the maximum entropy approach to evolution

Many problems in evolutionary theory are cast in dyadic terms, such as the polar oppositions of organism and environment. We argue that a triadic conceptual structure offers an alternative perspective under which the information generating role of evolution as a physical process can be analyzed, and propose a new diagrammatic approach. Peirce's natural philosophy was deeply influenced by his reception of both Darwin's theory and thermodynamics. Thus, we elaborate on a new synthesis which puts together his theory of signs and modern Maximum Entropy approaches to evolution in a process discourse. Following recent contributions to the naturalization of Peircean semiosis, pointing towards ‘physiosemiosis’ or ‘pansemiosis’, we show that triadic structures involve the conjunction of three different kinds of causality, efficient, formal and final. In this, we accommodate the state-centered thermodynamic framework to a process approach. We apply this on Ulanowicz's analysis of autocatalytic cycles as primordial patterns of life. This paves the way for a semiotic view of thermodynamics which is built on the idea that Peircean interpretants are systems of physical inference devices evolving under natural selection. In this view, the principles of Maximum Entropy, Maximum Power, and Maximum Entropy Production work together to drive the emergence of information carrying structures, which at the same time maximize information capacity as well as the gradients of energy flows, such that ultimately, contrary to Schrödinger's seminal contribution, the evolutionary process is seen to be a physical expression of the Second Law.     

Modulation of the hepatitis C virus RNA-dependent RNA polymerase activity by the non-structural (NS) 3 helicase and the NS4B membrane protein

The hepatitis C virus (HCV) nonstructural protein 5B (NS5B) is believed to be the central catalytic enzyme responsible for HCV replication but there are many unanswered questions about how its activity is controlled. In this study we reveal that two other HCV proteins, NS3 (a protease/helicase) and NS4B (a hydrophobic protein of unknown function), physically and functionally interact with the NS5B polymerase. We describe a new procedure for generating highly pure NS4B, and use this protein in biochemical studies together with NS5B and NS3. To study the functional effects of the protein-protein interactions, we have developed an in vitro replication assay using the natural noncoding 3' regions of the respective positive ((+)-3'-untranslated region) and negative ((-)-3'-terminal region) RNA strands of the HCV genome. Our studies show that NS3 dramatically modulates template recognition by NS5B and changes the synthetic products generated by this enzyme. The use of an NTPase-deficient mutant form of NS3 demonstrates that the NTPase activity (and thus helicase activity) of this protein is specifically required for these effects. Moreover, NS4B is found to be a negative regulator of the NS3-NS5B replication complex. Overall, these results reveal that NS3, NS4B, and NS5B can interact to form a regulatory complex that could feature in the process of HCV replication.     

Recent Progress on the Analysis of Power-law Features in Complex Cellular Networks

Complex interactions between different kinds of bio-molecules and essential nutrients are responsible for cellular functions. Rapid advances in theoretical modeling and experimental analyses have shown that drastically different biological and non-biological networks share a common architecture. That is, the probability that a selected node in the network has exactly k edges decays as a power-law. This finding has definitely opened an intense research and debate on the origin and implications of this ubiquitous pattern. In this review, we describe the recent progress on the emergence of power-law distributions in cellular networks. We first show the internal characteristics of the observed complex networks uncovered using graph theory. We then briefly review some works that have significantly contributed to the theoretical analysis of cellular networks and systems, from metabolic and protein networks to gene expression profiles. This prevalent topology observed in so many diverse biological systems suggests the existence of generic laws and organizing principles behind the cellular networks.