RNA-like Polymers Research Articles

Stochastic parabolic growth promotes coexistence and a relaxed error threshold in RNA-like replicator populations

The RNA world hypothesis proposes that during the early evolution of life, primordial genomes of the first self-propagating evolutionary units existed in the form of RNA-like polymers. Autonomous, non-enzymatic, and sustained replication of such information carriers presents a problem, because product formation and hybridization between template and copy strands reduces replication speed. Kinetics of growth is then parabolic with the benefit of entailing competitive coexistence, thereby maintaining diversity. Here, we test the information-maintaining ability of parabolic growth in stochastic multispecies population models under the constraints of constant total population size and chemostat conditions. We find that large population sizes and small differences in the replication rates favor the stable coexistence of the vast majority of replicator species (‘genes’), while the error threshold problem is alleviated relative to exponential amplification. In addition, sequence properties (GC content) and the strength of resource competition mediated by the rate of resource inflow determine the number of coexisting variants, suggesting that fluctuations in building block availability favored repeated cycles of exploration and exploitation. Stochastic parabolic growth could thus have played a pivotal role in preserving viable sequences generated by random abiotic synthesis and providing diverse genetic raw material to the early evolution of functional ribozymes.

Stochastic parabolic growth promotes coexistence and a relaxed error threshold in RNA-like replicator populations.

The RNA world hypothesis proposes that during the early evolution of life, primordial genomes of the first self-propagating evolutionary units existed in the form of RNA-like polymers. Autonomous, non-enzymatic, and sustained replication of such information carriers presents a problem, because product formation and hybridization between template and copy strands reduces replication speed. Kinetics of growth is then parabolic with the benefit of entailing competitive coexistence, thereby maintaining diversity. Here, we test the information-maintaining ability of parabolic growth in stochastic multispecies population models under the constraints of constant total population size and chemostat conditions. We find that large population sizes and small differences in the replication rates favor the stable coexistence of the vast majority of replicator species ('genes'), while the error threshold problem is alleviated relative to exponential amplification. In addition, sequence properties (GC content) and the strength of resource competition mediated by the rate of resource inflow determine the number of coexisting variants, suggesting that fluctuations in building block availability favored repeated cycles of exploration and exploitation. Stochastic parabolic growth could thus have played a pivotal role in preserving viable sequences generated by random abiotic synthesis and providing diverse genetic raw material to the early evolution of functional ribozymes.

Visualizing ribonuclease digestion of RNA-like polymers produced by hot wet-dry cycles

Polymerization of nucleotides under prebiotic conditions simulating the early Earth has been extensively studied. Several independent methods have been used to verify that RNA-like polymers can be produced by hot wet-dry cycling of nucleotides. However, it has not been shown that these RNA-like polymers are similar to biological RNA with 3′-5′ phosphodiester bonds. In the results described here, RNA-like polymers were generated from 5′-monophosphate nucleosides AMP and UMP. To confirm that the polymers resemble biological RNA, ribonuclease A should catalyze hydrolysis of the 3′-5′ phosphodiester bonds between pyrimidine nucleotides to each other or to purine nucleotides, but not purine-purine nucleotide bonds. Here we show AFM images of specific polymers produced by hot wet-dry cycling of AMP, UMP and AMP/UMP (1:1) solutions on mica surfaces, before and after exposure to ribonuclease A. AMP polymers were unaffected by ribonuclease A but UMP polymers disappeared. This indicates that a major fraction of the bonds in the UMP polymers is indeed 3′-5′ phosphodiester bonds. Some of the polymers generated from the AMP/UMP mixture also showed clear signs of cleavage. Because ribonuclease A recognizes the ester bonds in the polymers, we show for the first time that these prebiotically produced polymers are in fact similar to biological RNA but are likely to be linked by a mixture of 3′-5′ and 2′-5′ phosphodiester bonds.

Molecular take on phase separation of FUS.

Poly(ADP-ribose) (PAR) is an RNA-like polymer that regulates an increasing number of biological processes. Dysregulation of PAR is implicated in neurodegenerative diseases characterized by abnormal protein aggregation, including amyotrophic lateral sclerosis (ALS). PAR forms condensates with FUS, an RNA-binding protein linked with ALS, through an unknown mechanism. Here, we demonstrate that a strikingly low concentration of PAR (1 nM) is sufficient to trigger condensation of FUS near its physiological concentration (1 μM), which is three orders of magnitude lower than the concentration at which RNA induces condensation (1 μM). Unlike RNA, which associates with FUS stably, PAR interacts with FUS transiently, triggering FUS to oligomerize into condensates. Moreover, inhibition of a major PAR-synthesizing enzyme, PARP5a, diminishes FUS condensation in cells. Despite their structural similarity, PAR and RNA co-condense with FUS, driven by disparate modes of interaction with FUS. Thus, we uncover a mechanism by which PAR potently seeds FUS condensation. I will also present polymer property of PAR which is distinct from DNA and RNA.

Visualizing RNA polymers produced by hot wet-dry cycling

It is possible that the transition from abiotic systems to life relied on RNA polymers that served as ribozyme-like catalysts and for storing genetic information. The source of such polymers is uncertain, but previous investigations reported that wet–dry cycles simulating prebiotic hot springs provide sufficient energy to drive condensation reactions of mononucleotides to form oligomers and polymers. The aim of the study reported here was to verify this claim and visualize the products prepared from solutions composed of single mononucleotides and 1:1 mixture of two mononucleotides. Therefore, we designed experiments that allowed comparisons of all such mixtures representing six combinations of the four mononucleotides of RNA. We observed irregular stringy patches and crystal strands when wet-dry cycling was performed at room temperature (20 °C). However, when the same solutions were exposed to wet–dry cycles at 80 °C, we observed what appeared to be true polymers. Their thickness was consistent with RNA-like products composed of covalently bonded monomers, while irregular strings and crystal segments of mononucleotides dried or cycled at room temperature were consistent with structures assembled and stabilized by weak hydrogen bonds. In a few instances we observed rings with short polymer attachments. These observations are consistent with previous claims of polymerization during wet–dry cycling. We conclude that RNA-like polymers and rings could have been synthesized non-enzymatically in freshwater hot springs on the prebiotic Earth with sizes sufficient to fold into ribozymes and genetic molecules required for life to begin.

Poly(ADP-ribose) drives condensation of FUS via a transient interaction

Poly(ADP-ribose) (PAR) is an RNA-like polymer that regulates an increasing number of biological processes. Dysregulation of PAR is implicated in neurodegenerative diseases characterized by abnormal protein aggregation, including amyotrophic lateral sclerosis (ALS). PAR forms condensates with FUS, an RNA-binding protein linked with ALS, through an unknown mechanism. Here, we demonstrate that a strikingly low concentration of PAR (1nM) is sufficient to trigger condensation of FUS near its physiological concentration (1μM), which is three orders of magnitude lower than the concentration at which RNA induces condensation (1μM). Unlike RNA, which associates with FUS stably, PAR interacts with FUS transiently, triggering FUS to oligomerize into condensates. Moreover, inhibition of a major PAR-synthesizing enzyme, PARP5a, diminishes FUS condensation in cells. Despite their structural similarity, PAR and RNA co-condense with FUS, driven by disparate modes of interaction with FUS. Thus, we uncover a mechanism by which PAR potently seeds FUS condensation.

Poly(ADP-ribose) drives condensation of FUS via a transient interaction

Poly(ADP-ribose) (PAR) is an RNA-like polymer that regulates an increasing number of biological processes. Dysregulation of PAR is implicated in neurodegenerative diseases characterized by abnormal protein aggregation, including Amyotrophic Lateral Sclerosis (ALS). PAR forms condensates with FUS, an RNA-binding protein linked with ALS, through an unknown mechanism. Here, we demonstrate that a strikingly low concentration of PAR (1 nM) is sufficient to trigger condensation of FUS near its physiological concentration (1 μM), which is three orders of magnitude lower than the concentration at which RNA induces condensation (1 μM).

Role of Stacking Interactions in the Stability of Primitive Genetics: A Quantum Chemical View.

The origin of genetic material on earth is an age-old, entangled mystery that lacks a unanimous explanation. Recent studies have suggested that noncanonical bases such as barbituric acid (BA), melamine (MM), cyanuric acid (CA), and 2,4,6-triaminopyrimidine (TAP) may have undergone molecular selection within the "prebiotic soup" to spontaneously form supramolecular assemblies, which then covalently assembled into an RNA-like polymer (preRNA). However, information on the role of intrinsic interactions of these candidate heterocycles in their molecular selection as the components of preRNA, and the subsequent transition from preRNA to RNA, is currently missing in the literature. To fill this gap in our knowledge on the origin and evolution of primitive genetics, the present work employs density functional theory (B3LYP-D3) to evaluate and compare the stacking propensities of dimers containing prebiotic noncanonical (BA, MM, CA, and TAP) and/or canonical RNA bases (A, C, G, and U). Our detailed analysis of the variation in stacking strength with respect to four characteristic geometrical parameters between the monomers [i.e., the vertical distance, the angle of rotation, and (two) displacements in the x and y directions] reveals that stacking between nonidentical bases is preferred over identical bases for both prebiotic-prebiotic and canonical-canonical dimers. This not only underscores the similarity between the fundamental chemical properties of preRNA and RNA constituents but also supports the likelihood of the evolution of modern (RNA) genetics from primitive (preRNA) genetics. Furthermore, greater average stacking stabilization of canonical dimers than that of dimers containing one canonical and one preRNA nucleobase (by ∼5 kJ mol-1) or dimers solely containing preRNA nucleobases (by ∼12 kJ mol-1) indicates that enhanced stacking is an important factor that may have spurred the evolution of preRNA to an intermediate informational polymer to RNA. More importantly, our study identifies the central roles of CA, BA, and TAP in stacking stabilization within the preRNA and of BA in stacking interactions within the intermediate polymers and suggests that these heterocycles may have played distinct roles in various stages during the evolution from preRNA to RNA. Overall, our results highlight the significance of stacking interactions in the selection of nucleobase components of preRNA.

Poly(Adp-Ribose) Drives Condensation of Fus Via a Transient Interaction

Poly(ADP-ribose) (PAR) is an RNA-like polymer that regulates an increasing number of biological processes. Dysregulation of PAR is implicated in neurodegenerative diseases characterized by abnormal protein aggregation, including Amyotrophic Lateral Sclerosis (ALS). PAR forms condensates with FUS, an RNA-binding protein linked with ALS, through an unknown mechanism. Here, we demonstrate that a strikingly low concentration of PAR (1 nM) is sufficient to trigger condensation of FUS near its physiological concentration (1 µM), which is three orders of magnitude lower than the concentration at which RNA induces condensation (1 µM). Unlike RNA, which associates with FUS stably, PAR interacts with FUS transiently, triggering FUS to oligomerize into condensates. Moreover, inhibition of a major PAR-synthesizing enzyme, PARP5a, diminishes FUS condensation in cells. Despite their structural similarity, PAR and RNA co-condense with FUS, driven by disparate modes of interaction with FUS. Thus, we uncover a mechanism by which PAR potently seeds FUS condensation.

A significant equilibrium distinguishing DNA/RNA-like phases in 2T SAW model on Husimi tetrahedron lattice

A modified two-tolerate (2T) SAW polymer model is carried on the Husimi tetrahedron lattice to describe the DNA/RNA-like biomacromolecules. With a simplified assemble of configurations to catch the zipping/unzipping feature of intra-pair-able polymer chains, the thermodynamics are exactly derived via the recursive property of Husimi lattice. The probability(density) of monomers in single and paired strands, and solvent are calculated from recursive partial partition functions to characterize the phase diagrams. The model is found to be a practical setup fitting both DNA- and RNA-like polymers, with rich phase diagrams presenting the DNA/RNA-like regions and the unwinding/polymerase transitions. An intriguing equilibrium is found to separate DNA- and RNA-like regions with a particular parameters setup, where the single and double strands co-exist in the system with constant densities regardless of temperature.

A Polyaddition Model for the Prebiotic Polymerization of RNA and RNA-Like Polymers.

Implicit in the RNA world hypothesis is that prebiotic RNA synthesis, despite occurring in an environment without biochemical catalysts, produced the long RNA polymers which are essential to the formation of life. In order to investigate the prebiotic formation of long RNA polymers, we consider a general solution of functionally identical monomer units that are capable of bonding to form linear polymers by a step-growth process. Under the assumptions that (1) the solution is well-mixed and (2) bonding/unbonding rates are independent of polymerization state, the concentration of each length of polymer follows the geometric Flory-Schulz distribution. We consider the rate dynamics that produce this equilibrium; connect the rate dynamics, Gibbs free energy of bond formation, and the bonding probability; solve the dynamics in closed form for the representative special case of a Flory-Schulz initial condition; and demonstrate the effects of imposing a maximum polymer length. Afterwards, we derive a lower bound on the error introduced by truncation and compare this lower bound to the actual error found in our simulation. Finally, we suggest methods to connect these theoretical predictions to experimental results.

Lipid-Assisted Polymerization of Nucleotides.

In addition to being one of the proponents of the “Lipid World hypothesis”, David Deamer, together with other colleagues, pioneered studies involving formation of RNA-like oligomers from their ‘non-activated’, prebiotically plausible monomeric moieties. In particular, the pioneering work in this regard was a publication from 2008 in Origins of Life and Evolution of Biospheres, The Journal of the International Astrobiology Society, wherein we described the formation of RNA-like oligomers from nucleoside 5’-monophosphates. In that study, we had simulated a terrestrial geothermal environment, a niche that is thought to have facilitated the prebiotic non-enzymatic synthesis of polynucleotides. We showed that a mixture of lipids and non-activated mononucleotides resulted in the formation of relatively long strands of RNA-like polymers when subjected to repeated cycles of dehydration and rehydration (DH-RH). Since 2008, terrestrial geothermal niches and DH-RH conditions have been explored in the context of several other prebiotic processes. In this article, we review the work that we and other researchers have carried out since then in this line of research, including the development of new apparatus to carry out the simulation of prebiotic terrestrial geothermal environments.

At the Interface of Three Nucleic Acids: The Role of RNA-Binding Proteins and Poly(ADP-ribose) in DNA Repair

RNA-binding proteins (RBPs) regulate RNA metabolism, from synthesis to decay. When bound to RNA, RBPs act as guardians of the genome integrity at different levels, from DNA damage prevention to the post-transcriptional regulation of gene expression. Recently, RBPs have been shown to participate in DNA repair. This fact is of special interest as DNA repair pathways do not generally involve RNA. DNA damage in higher organisms triggers the formation of the RNA-like polymer - poly(ADP-ribose) (PAR). Nucleic acid-like properties allow PAR to recruit DNA- and RNA-binding proteins to the site of DNA damage. It is suggested that poly(ADP-ribose) and RBPs not only modulate the activities of DNA repair factors, but that they also play an important role in the formation of transient repairosome complexes in the nucleus. Cytoplasmic biomolecules are subjected to similar sorting during the formation of RNA assemblages by functionally related mRNAs and promiscuous RBPs. The Y-box-binding protein 1 (YB-1) is the major component of cytoplasmic RNA granules. Although YB-1 is a classic RNA-binding protein, it is now regarded as a non-canonical factor of DNA repair.

Mobility of a Mononucleotide within a Lipid Matrix: A Neutron Scattering Study.

An essential question in studies on the origins of life is how nucleic acids were first synthesized and then incorporated into compartments about 4 billion years ago. A recent discovery is that guided polymerization within organizing matrices could promote a non-enzymatic condensation reaction allowing the formation of RNA-like polymers, followed by encapsulation in lipid membranes. Here, we used neutron scattering and deuterium labelling to investigate 5′-adenosine monophosphate (AMP) molecules captured in a multilamellar phospholipid matrix. The aim of the research was to determine and compare how mononucleotides are captured and differently organized within matrices and multilamellar phospholipid structures and to explore the role of water in organizing the system to determine at which level the system becomes sufficiently anhydrous to lock the AMP molecules into an organized structure and initiate ester bond synthesis. Elastic incoherent neutron scattering experiments were thus employed to investigate the changes of the dynamic properties of AMP induced by embedding the molecules within the lipid matrix. The influence of AMP addition to the lipid membrane organization was determined through diffraction measurement, which also helped us to define the best working Q range for dynamical data analysis with respect to specific hydration. The use of different complementary instruments allowed coverage of a wide time-scale domain, from ns to ps, of atomic mean square fluctuations, providing evidence of a well-defined dependence of the AMP dynamics on the hydration level.

Dry/Wet Cycling and the Thermodynamics and Kinetics of Prebiotic Polymer Synthesis.

The endoergic nature of protein and nucleic acid assembly in aqueous media presents two questions that are fundamental to the understanding of life’s origins: (i) how did the polymers arise in an aqueous prebiotic world; and (ii) once formed in some manner, how were they sufficiently persistent to engage in further chemistry. We propose here a quantitative resolution of these issues that evolved from recent accounts in which RNA-like polymers were produced in evaporation/rehydration cycles. The equilibrium Nm + Nn ↔ Nm+n + H2O is endoergic by about 3.3 kcal/mol for polynucleotide formation, and the system thus lies far to the left in the starting solutions. Kinetic simulations of the evaporation showed that simple Le Châtelier’s principle shifts were insufficient, but the introduction of oligomer-stabilizing factors of 5–10 kcal/mol both moved the process to the right and respectively boosted and retarded the elongation and hydrolysis rates. Molecular crowding and excluded volume effects in present-day cells yield stabilizing factors of that order, and we argue here that the crowded conditions in the evaporites generate similar effects. Oligomer formation is thus energetically preferred in those settings, but the process is thwarted in each evaporation step as diffusion becomes rate limiting. Rehydration dissipates disordered oligomer clusters in the evaporites, however, and subsequent dry/wet cycling accordingly “ratchets up” the system to an ultimate population of kinetically trappedthermodynamically preferred biopolymers.

Salt-promoted synthesis of RNA-like molecules in simulated hydrothermal conditions.

A fundamental problem in origins of life research is how the first polymers with the properties of nucleic acids were synthesized and incorporated into living systems on the prebiotic Earth. Here, we show that RNA-like polymers can be synthesized non-enzymatically from 5'-phosphate mononucleosides in salty environments. The polymers were identified and analyzed by gel electrophoresis, nanopore analysis, UV spectra, and action of RNases. The synthesis of phosphodiester bonds is driven by the chemical potential made available in the fluctuating hydrated and anhydrous conditions of hydrothermal fields associated with volcanic land masses.

Phenotype/genotype sequence complementarity and prebiotic replicator coexistence in the metabolically coupled replicator system.

BackgroundRNA or RNA-like polymers are the most likely candidates for having played the lead roles on the stage of the origin of life. RNA is known to feature two of the three essential functions of living entities (metabolism, heredity and membrane): it is capable of unlimited heredity and it has a proven capacity for catalysing very different chemical reactions which may form simple metabolic networks. The Metabolically Coupled Replicator System is a class of simulation models built on these two functions to show that an RNA World scenario for the origin of life is ecologically feasible, provided that it is played on mineral surfaces. The fact that RNA templates and their copies are of complementary base sequences has an obvious dynamical relevance: complementary strains may have very different structures and, consequently, functions – one may specialize for increasing enzymatic activity while the other takes the role of the gene of the enzyme.ResultsIncorporating the functional divergence of template and copy into the Metabolically Coupled Replicator System model framework we show that sequence complementarity 1) does not ruin the coexistence of a set of metabolically cooperating replicators; 2) the replicator system remains resistant to, but also tolerant with its parasites; 3) opens the way to the evolutionary differentiation of phenotype and genotype through a primitive version of phenotype amplification.ConclusionsThe functional asymmetry of complementary RNA strains results in a shift of phenotype/genotype (enzyme/gene) proportions in MCRS, favouring a slight genotype dominance. This asymmetry is expected to reverse due to the evolved trade-off of high “gene” replicability and high catalytic activity of the corresponding “enzyme” in expense of its replicability. This trade-off is the first evolutionary step towards the “division of labour” among enzymes and genes, which has concluded in the extreme form of phenotype amplification characteristic of our recent DNA-RNA-protein World.Electronic supplementary materialThe online version of this article (doi:10.1186/s12862-014-0234-8) contains supplementary material, which is available to authorized users.

Adenosine Monophosphate Forms Ordered Arrays in Multilamellar Lipid Matrices: Insights into Assembly of Nucleic Acid for Primitive Life

A fundamental question of biology is how nucleic acids first assembled and then were incorporated into the earliest forms of cellular life 4 billion years ago. The polymerization of nucleotides is a condensation reaction in which phosphodiester bonds are formed. This reaction cannot occur in aqueous solutions, but guided polymerization in an anhydrous lipid environment could promote a non-enzymatic condensation reaction in which oligomers of single stranded nucleic acids are synthesized. We used X-ray scattering to investigate 5′-adenosine monophosphate (AMP) molecules captured in a multilamellar phospholipid matrix composed of dimyristoylphosphatidylcholine. Bragg peaks corresponding to the lateral organization of the confined AMP molecules were observed. Instead of forming a random array, the AMP molecules are highly entangled, with the phosphate and ribose groups in close proximity. This structure may facilitate polymerization of the nucleotides into RNA-like polymers.

Efficient Self-Assembly in Water of Long Noncovalent Polymers by Nucleobase Analogues

Molecular self-assembly is widely appreciated to result from a delicate balance between several noncovalent interactions and solvation effects. However, current design approaches for achieving self-assembly in water with small, synthetic molecules do not consider all aspects of the hydrophobic effect, in particular the requirement of surface areas greater than 1 nm(2) for an appreciable free energy of hydration. With the concept of a minimum hydrophobic surface area in mind, we designed a system that achieves highly cooperative self-assembly in water. Two weakly interacting low-molecular-weight monomers (cyanuric acid and a modified triaminopyrimidine) are shown to form extremely long supramolecular polymer assemblies that retain water solubility. The complete absence of intermediate assemblies means that the observed equilibrium is between free monomers and supramolecular assemblies. These observations are in excellent agreement with literature values for the free energy of nucleic acid base interactions as well as the calculated free energy penalty for the exposure of hydrophobic structures in water. The results of our study have implications for the design of new self-assembling structures and hydrogel-forming molecules and may provide insights into the origin of the first RNA-like polymers.

Enhanced Nonenzymatic Ligation of Homopurine Miniduplexes: Support for Greater Base Stacking in a Pre‐RNA World

The ancestors of RNA? There is a long-standing proposal that contemporary nucleic acids might have evolved from RNA-like polymers that utilized only purine-purine base pairs. Here we demonstrate the great advantage that increased nucleobase stacking area provides for nonenzymatic ligation.