Computational universality of fungal sandpile automata

Hyphae within the mycelia of the ascomycetous fungi are compartmentalised by septa. Each septum has a pore that allows for inter-compartmental and inter-hyphal streaming of cytosol and even organelles. The compartments, however, have special organelles, Woronin bodies, that can plug the pores. When the pores are blocked, no flow of cytoplasm takes place. Inspired by the controllable compartmentalisation within the mycelium of the ascomycetous fungi we designed two-dimensional fungal automata. A fungal automaton is a cellular automaton where communication between neighbouring cells can be blocked on demand. We demonstrate computational universality of the fungal automata by implementing sandpile cellular automata circuits there. We reduce the Monotone Circuit Value Problem to the Fungal Automaton Prediction Problem. We construct families of wires, cross-overs and gates to prove that the fungal automata are P-complete

Logics in fungal Mycelium networks

The living mycelium networks are capable of efficient sensorial fusion over very large areas and distributed decision making. The information processing in the mycelium networks is implemented via propagation of electrical and chemical signals en pair with morphological changes in the mycelium structure. These information processing mechanisms are manifested in experimental laboratory findings that show that the mycelium networks exhibit rich dynamics of neuron-like spiking behaviour and a wide range of non-linear electrical properties. On an example of a single real colony of Aspergillus niger, we demonstrate that the non-linear transformation of electrical signals and trains of extracellular voltage spikes can be used to implement logical gates and circuits. The approaches adopted include numerical modelling of excitation propagation on the mycelium network, representation of the mycelium network as a resistive and capacitive network and an experimental laboratory study on mining logical circuits in mycelium bound composites

What if plants compute?

The unexpended cognitive capacities of plants suggest the possibility of combining them with advances in computation. It is important to explore such a new field of research despite the incompleteness of the empirical support for it

Mining logical circuits in fungi

Living substrates are capable for nontrivial mappings of electrical signals due to the substrate nonlinear electrical characteristics. This property can be used to realise Boolean functions. Input logical values are represented by amplitude or frequency of electrical stimuli. Output logical values are decoded from electrical responses of living substrates. We demonstrate how logical circuits can be implemented in mycelium bound composites. The mycelium bound composites (fungal materials) are getting growing recognition as building, packaging, decoration and clothing materials. Presently the fungal materials are passive. To make the fungal materials adaptive, i.e. sensing and computing, we should embed logical circuits into them. We demonstrate experimental laboratory prototypes of many-input Boolean functions implemented in fungal materials from oyster fungi P. ostreatus. We characterise complexity of the functions discovered via complexity of the space-time configurations of one-dimensional cellular automata governed by the functions. We show that the mycelium bound composites can implement representative functions from all classes of cellular automata complexity including the computationally universal. The results presented will make an impact in the field of unconventional computing, experimental demonstration of purposeful computing with fungi, and in the field of intelligent materials, as the prototypes of computing mycelium bound composites

Responsive fungal insoles for pressure detection

Mycelium bound composites are promising materials for a diverse range of applications including wearables and building elements. Their functionality surpasses some of the capabilities of traditionally passive materials, such as synthetic fibres, reconstituted cellulose fibres and natural fibres. Thereby, creating novel propositions including augmented functionality (sensory) and aesthetic (personal fashion). Biomaterials can offer multiple modal sensing capability such as mechanical loading (compressive and tensile) and moisture content. To assess the sensing potential of fungal insoles we undertook laboratory experiments on electrical response of bespoke insoles made from capillary matting colonised with oyster fungi Pleurotus ostreatus to compressive stress which mimics human loading when standing and walking. We have shown changes in electrical activity with compressive loading. The results advance the development of intelligent sensing insoles which are a building block towards more generic reactive fungal wearables. Using FitzHugh-Nagumo model we numerically illustrated how excitation wave-fronts behave in a mycelium network colonising an insole and shown that it may be possible to discern pressure points from the mycelium electrical activity

Fungal Automata

We study a cellular automaton (CA) model of information dynamics on a single hypha of a fungal mycelium. Such a filament is divided in compartments (here also called cells) by septa. These septa are invaginations of the cell wall and their pores allow for flow of cytoplasm between compartments and hyphae. The septal pores of the fungal phylum of the Ascomycota can be closed by organelles called Woronin bodies. Septal closure is increased when the septa become older and when exposed to stress conditions. Thus, Woronin bodies act as informational flow valves. The one dimensional fungal automata is a binary state ternary neighbourhood CA, where every compartment follows one of the elementary cellular automata (ECA) rules if its pores are open and either remains in state `0' (first species of fungal automata) or its previous state (second species of fungal automata) if its pores are closed. The Woronin bodies closing the pores are also governed by ECA rules. We analyse a structure of the composition space of cell-state transition and pore-state transitions rules, complexity of fungal automata with just few Woronin bodies, and exemplify several important local events in the automaton dynamics

From CoA ester supply to a yeast communication toolkit in Saccharomyces cerevisiae

Saccharomyces cerevisiae is the most widely used eukaryotic chassis in synthetic biology, as hu-manity and yeast share a long and fruitful history. For synthetic biology applications, S. cerevisiae was extensively used for metabolic engineering as well as for the construction of artificial net-works. To contribute to the metabolic engineering achievements conducted in S. cerevisiae, we extended its metabolic capacities by providing non-native short-chain acyl-coenzyme A esters as metabolic precursors. In order to advance the construction of artificial networks to multicel-lular systems we provided a comprehensive yeast communication toolkit (YCTK), and demon-strated its usability for the rapid assembly of synthetic cell-cell communication systems. Engineered production of short-chain acyl-coenzyme A esters in Saccharomyces cerevisiae Globally, S. cerevisiae is one of the most commonly used chassis organisms in modern biotech-nology and constitutes a high economic value to the growing bioecomomy. With the objective to produce novel natural products in S. cerevisiae a bottleneck of the chassis was uncovered. Short-chain acyl-coenzyme A esters serve as intermediate compounds in fatty acid biosynthesis, and are building blocks for the production of polyketides, biopolymers, and other value-added chemicals. However, S. cerevisiae’s limited repertoire of short-chain acyl-CoAs effectively pre-vents its application as a production host for a plethora of natural products. To address and re-solve this limitation, we introduced metabolic pathways to five different acyl-CoA esters into S. cerevisiae. We engineered plasmid-based yeast strains that provide propionyl-CoA, methylmalonyl-CoA, n-butyryl-CoA, isovaleryl-CoA, and n-hexanoyl-CoA. For the production of propionyl-CoA and methylmalonyl-CoA, we reestablished a published feeding-dependent pro-duction route using the PrpE and Pcc enzymes to serve as benchmark for our feeding-independent production pathways that provided in our study comparable product concentra-tions. To ensure efficient extraction of the produced metabolites we established a yeast-specific metabolite extraction protocol to determine the intracellular acyl-CoA concentrations in the engineered strains. For the production of isovaleryl-CoA, we tested two different pathways but only obtained product formation from the alternative isovaleryl-CoA biosynthetic (AIB) pathway originating from Myxococcus xanthus and obtained 5.5±1.2 µM isovaleryl-CoA. To our knowledge, this is the first reported functional heterologous expression of this pathway in S. cerevisiae. For the production of n-butyryl-CoA and n-hexanoyl-CoA, we adapted the butanol production pathway for our purposes and measured approximately 6 µM intracellular concen-tration of butyryl-CoA and hexanoyl-CoA. For the feeding-dependent pathway towards propio-nyl-CoA we obtained intracellular concentrations of 5.3 ± 2.4 µM while the feeding independ-ent 3-hydroxypropionate (3HP) pathway produced 8.5 ± 3.7 µM. The extension of both propio-nyl-CoA pathways to produce methylmalonyl-CoA resulted only into production of 0.5 ± 0.1 µM and 0.3 ± 0.3 µM methylmalonyl-CoA. Not only but particularly for the production of methylmalonyl-CoA further optimization is required. To allow rapid pathway prototyping, op-timization and testing of alternative enzymes, we established a short-chain acyl-CoA Golden Gate collection. This collection enables together with the well-known Dueber yeast toolkit YTK collection the examination of different enzymes variants and to investigate optimized expres-sion of the corresponding genes. We conclude that the acyl-CoAs produced here, that are common building blocks of secondary metabolites, prepared the ground for prospective engineered production of a variety of natural products in S. cerevisiae. These acyl-CoA producing strains together with the short-chain acyl-CoA collection lay the foundation to further explore S. cerevisiae as a heterologous production host for high-value secondary metabolite production. Yeast communication toolkit The construction of multicellular networks was a proposed aim already early on in synthetic biology. Today, they still hold many promises like the division of labor or the performance of more complex tasks. Most of the systems so far were implemented in bacterial chassis and only a few examples exist for the eukaryotic chassis S. cerevisiae. Especially for gram-negative bacterial chassis, the quorum sensing system provides a large diversity of ready to use communication systems. Also, yeast species evolved a communication system using peptide-based pheromones to interact with the opposite mating type. Here, we employed the natural diversity of the pep-tide α-factor pheromones, the corresponding GPCR receptors, as well as of barrier proteases, that function similarly to quorum quenching enzymes. With the establishment of the Golden Gate yeast communication toolkit (YCTK) we provide a standardized collection of parts that al-low the rapid construction of multicellular networks in the model organism S. cerevisiae. The feasible designs are limitless as well as the number of envisioned applications. The YCTK collec-tion consists of responder (pheromone-responsive promoters), sender (mfα1 genes – α-factors), receiver (Ste2 receptors) and barrier (Bar1 proteases) parts. We characterized the dynamics of the pheromone-inducible promoters in the different mating-type strain backgrounds and de-termined the dose-response to the α-factor as well as their temporal response. The different promoters exhibited a range of different dynamics and properties that enable the implementa-tion of different prospective network design motives. The characterization results of the Ste2 receptors indicated that our collection is comprised of receptors with high α-factor promiscuity and of receptors with high substrate specificity for their cognate α-factor. Further we found that different Ste2 receptors exhibit different sensitivities towards the cognate as well as to non-cognate α-factors. The promiscuity of the Ste2 receptors did not correlate with the α-factor se-quences. Our likelihood analysis of the Ste2 receptors indicated that the ones closer related to S. cerevisiae tend to be stimulated by the α-factors of related species. Our likelihood analysis of the Ste2 receptors coincided with the phylogenetic relationships of the species. Interesting is also the finding that α-factors of species for which the receptor exhibited high α-factor promis-cuity stimulated only a few receptors. Even though only five of the selected barrier proteases were functionally expressed the characterization of the protease promiscuity was to our knowledge the most comprehensive study of its kind so far. Similar to the receptors we identi-fied promiscuous and substrate specific barrier proteases. The proposed model of a coevolution between the receptor and barrier proteases to recognize similar sequence motives of the α-factor was partly validated, however, the model is not universally applicable according to our results. The extended knowledge of the pheromone-inducible promoters, the crosstalk be-tween α-factors, receptors and barrier proteases, and an initial tunability test enabled proof of principle construction of multicellular systems using the YTCK collection. We engineered mul-ticellular logic gate-like population networks that allow the receiver cells to conditionally re-spond to the population composition. While the α-factor signaling motif is functional and was used to successfully establish OR and AND gate-like systems, signal disruption by a barrier pro-tease of a self-stimulating or a signaling motif requires further optimization. Overall, the reali-zation of multicellular networks using the YCTK was proven to be successful. To summarize, with the YCTK we provide a set of comprehensively characterized sender, re-ceiver, and barrier parts to facilitate the implementation of cell-cell and thus multicellular communication networks in S. cerevisiae

Numerical and experimental investigation of multi-species bacterial co-aggregation

This paper deals with the mathematical modeling of bacterial co-aggregation and its numerical implementation in a FEM framework. Since the concept of co-aggregation refers to the physical binding between cells of different microbial species, a system composed of two species is considered in the modeling framework. The extension of the model to an arbitrary number of species is straightforward. In addition to two-species (multi-species growth) dynamics, the transport of a nutritional substance and the extent of co-aggregation are introduced into the model as the third and fourth primary variables. A phase-field modeling approach is employed to describe the co-aggregation between the two species. The mathematical model is three-dimensional and fully based on the continuum description of the problem without any need for discrete agents which are the key elements of the individual-based modeling approach. It is shown that the use of a phase-field-based model is equivalent to a particular form of classical diffusion-reaction systems. Unlike the so-called mixture models, the evolution of each component of the multi-species system is captured thanks to the inherent capability of phase-field modeling in treating systems consisting of distinct multi-phases. The details of numerical implementation in a FEM framework are also presented. Indeed, a new multi-field user element is developed and implemented in ANSYS for this multiphysics problem. Predictions of the model are compared with the experimental observations. By that, the versatility and applicability of the model and the numerical tool are well established

Exploring the Dynamics of Fungal Cellular Automata

Cells in a fungal hyphae are separated by internal walls (septa). The septa have tiny pores that allow cytoplasm flowing between cells. Cells can close their septa blocking the flow if they are injured, preventing fluid loss from the rest of filament. This action is achieved by special organelles called Woronin bodies. Using the controllable pores as an inspiration we advance one and two-dimensional cellular automata into Elementary fungal cellular automata (EFCA) and Majority fungal automata (MFA) by adding a concept of Woronin bodies to the cell state transition rules. EFCA is a cellular automaton where the communications between neighboring cells can be blocked by the activation of the Woronin bodies (Wb), allowing or blocking the flow of information (represented by a cytoplasm and chemical elements it carries) between them. We explore a novel version of the fungal automata where the evolution of the system is only affected by the activation of the Wb. We explore two case studies: the Elementary Fungal Cellular Automata (EFCA), which is a direct application of this variant for elementary cellular automata rules, and the Majority Fungal Automata (MFA), which correspond to an application of the Wb to two dimensional automaton with majority rule with Von Neumann neighborhood. By studying the EFCA model, we analyze how the 256 elementary cellular automata rules are affected by the activation of Wb in different modes, increasing the complexity on applied rule in some cases. Also we explore how a consensus over MFA is affected when the continuous flow of information is interrupted due to the activation of Woronin bodies.Comment: 31 pages, 30 figure