The emergence of synchronization behavior in Physarum polycephalum and its particle approximation

The regeneration process of contractile oscillation in the plasmodium of Physarum polycephalum is investigated experimentally and modelled computationally. When placed in a well, the Physarum cell restructures the body (fusion of small granule-like cells) and shows various complex oscillation patterns. After it completed the restructuring and regained synchronized oscillation within the body, the cell shows bilateral oscillation or rotating wave pattern. This regeneration process did not depend on the well size and all the cases tested here showed similar time course. Phase synchronization analysis based on Hilbert Transform also suggested that the cell can develop a fully synchronized oscillation within a fixed time no matter what the cell size is. A particle-based computational model was developed in order to model the emergence of oscillation patterns. Particles employing very simple and identical sensory and motor behaviors interacted with each other via the sensing and deposition of chemoattractants in a diffusive environment. From a random and almost homogeneous distribution, emergent domains of oscillatory activity emerged. By increasing the sensory radius the model simulated the regeneration process of the real plasmodium. In addition, the model replicated the rotating wave and bilateral oscillation pattern when the sensory radius was increased. The results suggest that complex emergent oscillatory behaviors (and thus the high-level systems which may utilize them, such as pumping and transport mechanisms) may be developed from simple materials inspired by Physarum slime mold.     

Characterization of the nucleolar protein, B-36, using monoclonal antibodies

A panel of nine monoclonal antibodies has been produced against a major nuclear protein, B-36, purified from the slime mold Physarum polycephalum. B-36, a 34 kD protein biochemically similar to the major structural proteins of mammalian hnRNP particles, was previously shown to be largely associated with the nucleolus. Eight of the monoclonal antibodies are specific for B-36 protein in Physarum and at least three different epitopes are represented among these eight. Using the monoclonal antibodies B-36 has been shown to be localized exclusively to the nucleolus in actively-growing Physarum cultures. The nucleolar localization of B-36 is dependent on the presence of intact RNA, but not DNA, supporting the hypothesis that B-36 is associated with nucleolar RNA, possibly in some analogous manner to the interaction of the related proteins within heterogeneous nuclear ribonucleoproteins (hnRNP) particles. B-36 is apparently a highly conserved nucleolar protein in eukaryotes as all eight of the monoclonal antibodies specific for B-36 in Physarum are also specific for a 34.5 kD nucleolar protein in rat liver. This indicates that a minimum of three distinct epitopes are conserved in B-36 protein from slime mold to rat.     

Chemotaxis toward carbohydrates and amino acids in Physarum polycephalum

A method is described for assaying chemotaxis in the acellular slime mold Physarum polycephalum. It consists of measuring the amount of plasmodium that moves on a strip of nitrocellulose membrane filter Millipore in response to a gradient of an attractant. Time course of chemotactic response of the slime mold is described. Different factors that affect chemotaxis in the slime mold such as: culture care and stage of growth of microplasmodia, substratum used for cell movement, nature of the gradient, effect of salts, pH and temperature are described. From concentration-response curves for different attractants several parameters of the chemotactic effect, such as threshold concentration, half maximal concentration, and maximal effective concentration can be determined. As a group, sugars are more effective chemotactic agents than amino acids. Glucose and galactose, which support the growth of the slime mold, are shown to have high positive chemotactic effect. 3-O-Methyl- -glucose and 2-deoxy- -glucose are two sugars that do not support growth but are very effective attractants. Conversely, fructose which supports slime mold growth is at best a weak attractant. The results support the view that the chemotactic effects of different sugars are not dependent on their growth-supporting value.     

Involvement of Glutathione in the Differentiation of the Slime Mold Physarum polycephalum

The effects of experimentally-altered glutathione concentration on differentiation of the slime mold, Physarum polycephalum were examined. Spherulation was induced by transfer of Physarum from growth medium to a salts-only starvation medium. As differentiation proceeded, superoxide dismutase (SOD) activity in control cultures increased by as much as 21-fold. This increase in SOD activity paralleled the rate of differentiation. Glutathione (GSH) concentration decreased during differentiation by more than 80% in all cultures, regardless of the initial concentration. The rate of differentiation was inversely related to the initial GSH concentration and directly proportional to the SOD activity. These observations suggest that a free radical mechanism may be involved in the differentiation of Physarum microplasmodia into spherules.     

Responses of the slime mold Physarum polycephalum to changing accelerations

Every cell is probably able to respond to gravity (g) via an unknown gravireceptor mechanism (supposed general gravisensitivity of cells). To investigate this mechanism a free-living ameboid cell, which uses gravity for its spatial orientation (geotaxis), was selected as a model system: the acellular slime mold Physarum polycephalum (Myxomycetes). In this paper results of 0 g-simulation experiments will be compared to results obtained in the Spacelab IML-1 Physarum experiment to stress the reliability of experiments performed on the fast-rotating clinostat.     

Identification of substrates for transglutaminase in Physarum polycephalum, an acellular slime mold, upon cellular mechanical damage

Transglutaminases are Ca(2+)-dependent enzymes that post-translationally modify proteins by crosslinking or polyamination at specific polypeptide-bound glutamine residues. Physarum polycephalum, an acellular slime mold, is the evolutionarily lowest organism expressing a transglutimase whose primary structure is similar to that of mammalian transglutimases. We observed transglutimase reaction products at injured sites in Physarum macroplasmodia upon mechanical damage. With use of a biotin-labeled primary amine, three major proteins constituting possible transglutimase substrates were affinity-purified from the damaged slime mold. The purified proteins were Physarum actin, a 40 kDa Ca(2+)-binding protein with four EF-hand motifs (CBP40), and a novel 33 kDa protein highly homologous to the eukaryotic adenine nucleotide translocator, which is expressed in mitochondria. Immunochemical analysis of extracts from the damaged macroplasmodia indicated that CBP40 is partly dimerized, whereas the other proteins migrated as monomers on SDS/PAGE. Of the three proteins, CBP40 accumulated most significantly around injured areas, as observed by immunofluoresence. These results suggested that transglutimase reactions function in the response to mechanical injury.     

Physarum vitronectin-like protein: an Arg-Gly-Asp-dependent cell-spreading protein with a distinct NH2-terminal sequence.

A 70-kDa protein cross-reacted with anti-bovine vitronectin was isolated from slime mold Physarum polycephalum. The NH2-terminal amino acid sequence of the protein, referred to as Physarum vitronectin-like protein, did not share any homology with those of animal vitronectins. It had cell-spreading activity, which was specifically inhibited by an Arg-Gly-Asp (RGD)-containing peptide.     

Sounds Synthesis with Slime Mould of Physarum Polycephalum

This paper introduces a novel application of bionic engineering: a bionic musical instrument using Physarum polycephalum. Physarum polycephalum is a huge single cell with thousands of nuclei, which behaves like a giant amoeba. During its foraging behavior this plasmodium produces electrical activity corresponding to different physiological states. We developed a method to render sounds from such electrical activity and thus represent spatio-temporal behavior of slime mould in a form apprehended auditorily. The electrical activity is captured by various electrodes placed on a Petri dish containing the cultured slime mold. Sounds are synthesized by a bank of parallel sinusoidal oscillators connected to the electrodes. Each electrode is responsible for one partial of the spectrum of the resulting sound. The behavior of the slime mould can be controlled to produce different timbres.     

On the Computing Potential of Intracellular Vesicles

Collision-based computing (CBC) is a form of unconventional computing in which travelling localisations represent data and conditional routing of signals determines the output state; collisions between localisations represent logical operations. We investigated patterns of Ca²⁺-containing vesicle distribution within a live organism, slime mould Physarum polycephalum, with confocal microscopy and observed them colliding regularly. Vesicles travel down cytoskeletal ‘circuitry’ and their collisions may result in reflection, fusion or annihilation. We demonstrate through experimental observations that naturally-occurring vesicle dynamics may be characterised as a computationally-universal set of Boolean logical operations and present a ‘vesicle modification’ of the archetypal CBC ‘billiard ball model’ of computation. We proceed to discuss the viability of intracellular vesicles as an unconventional computing substrate in which we delineate practical considerations for reliable vesicle ‘programming’ in both in vivo and in vitro vesicle computing architectures and present optimised designs for both single logical gates and combinatorial logic circuits based on cytoskeletal network conformations. The results presented here demonstrate the first characterisation of intracelluar phenomena as collision-based computing and hence the viability of biological substrates for computing.     

Expression of poriferasterol monoglucoside associated with differentiation of Physarum polycephalum

A glycolipid which was expressed during a differentiation from haploid myxoamoebae to diploid plasmodia of a true slime mold, Physarum polycephalum, has been examined. In the amoeboid stage, cells did not contain this glycolipid, but after conjugation of the haploid cells, this substance appeared and increased in its amount. From structural studies of the purified glycolipid, it has been identified as poriferasterol monoglucoside.     

Chemically induced changes in the morphology, dynamic activity and cytoskeletal organization of Physarum cell fragments

Spherical cell fragments derived from Physarum polycephalum by caffeine-treatment were used as an experimental system to investigate the influence of 15 externally applied substances on the general morphology, motile behavior and cytoskeletal organization of the acellular slime mold. In comparison to controls, the most obvious changes observed after chemical stimulation proved to be cytokinetic activities, ameboid-like movement phenomena, intense cell surface dynamics and formation of cytoplasmic actin fibrils. The results demonstrate the high adaptability of the microfilament system in Physarum even when subjected to extreme conditions in the external environment.     

Electron-cytochemical demonstration of acid phosphatase and thiolacetic acid-esterase activity in granular structures of the phagotrophic slime mold,Physarum confertum Macbr.

Summary The lysosomal marker enzyme, acid phosphatase, as well as non-specific esterase could be demonstrated cytochemically in cytoplasmic granules of the slime mold, Physarum confertum Macbr. The close association of enzyme-positive granules with food vacuoles is suggestive of their participation in the hydrolytic break-down of ingested food in a manner similar to that proposed for other phagotrophic cells and organisms.     

Actin co-purifies with RNA polymerase II.

RNA polymerase II, [EC2.7.7.6], from the slime mold Physarum polycephalum, purified over 4000-fold can contain a protein with an apparent molecular weight of 46,000. This protein is separated from the putative subunits of RNA polymerase II by polyacrylamide gel electrophoresis under non-denaturing conditions, and by chromatography on phosphocellulose. In this report we identify the protein as actin, and we point out that polypeptides of this apparent molecular weight which have been found associated with RNA polymerase II purified from other sources may also be actin from these organisms.     

Mitochondrial numbers increase during glucose deprivation in the slime mold Physarum polycephalum

Protoplasma - Glucose deprivation in the slime mold Physarum polycephalum leads to a specific morphotype, a highly motile mesoplasmodium. We investigated the ultrastructure of both mesoplasmodia...     

Slime mold inspired routing protocols for wireless sensor networks

Many biological systems are composed of unreliable components which self-organize effectively into systems that achieve a balance between efficiency and robustness. One such example is the true slime mold Physarum polycephalum which is an amoeba-like organism that seeks and connects food sources and efficiently distributes nutrients throughout its cell body. The distribution of nutrients is accomplished by a self-assembled resource distribution network of small tubes with varying diameter which can evolve with changing environmental conditions without any global control. In this paper, we exploit two different mechanisms of the slime mold??s tubular network formation process via laboratory experiments and mathematical behavior modeling to design two corresponding localized routing protocols for wireless sensor networks (WSNs) that take both efficiency and robustness into account. In the first mechanism of path growth, slime mold explores its immediate surroundings to discover and connect new food sources during its growth cycle. We adapt this mechanism for a path growth routing protocol by treating data sources and sinks as singular potentials to establish routes from the sinks to all the data sources. The second mechanism of path evolution is the temporal evolution of existing tubes through nonlinear feedback in order to distribute nutrients efficiently throughout the organism. Specifically, the diameters of tubes carrying large fluxes of nutrients grow to expand their capacities, and tubes that are not used decline and disappear entirely. We adapt the tube dynamics of the slime mold for a path evolution routing protocol. In our protocol, we identify one key adaptation parameter to adjust the tradeoff between efficiency and robustness of network routes. Through extensive realistic network simulations and ideal closed form or numerical computations, we validate the effectiveness of both protocols, as well as the efficiency and robustness of the resulting network connectivity.     

Isolation, identification, and characterization of histones from plasmodia of the true slime mold Physarum polycephalum using extraction with guanidine hydrochloride

Histones from plasmodia of the true slime mold Physarum polycephalum have been prepared free of slime by an approach to histone isolation that uses extraction of nuclei with 40% guanidine hydrochloride and chromatography of the extract on Bio-Rex 70. This procedure followed by chromatography or electrophoresis has been used to obtain pure fractions of histones from Physarum microplasmodia. Physarum microplasmodia have five major histone fractions, and we show by amino acid analysis, apparent molecular weight on three gel systems containing sodium dodecyl sulfate, mobility on gels containing Triton X-100, and other characterizations that these fractions are analogous to mammalian histones H1, H2A, H2B, H3, and H4. Significant differences between Physarum and mammalian histones are noted, with histone H1 showing by far the greatest variation. Histones H1 and H4 from Physarum microplasmodia have similar, but not identical, products of partial chymotryptic digestion compared with those of calf thymus histones H1 and H4. Labeling experiments, in vivo, showed that histone H1 is the major phosphorylated histone and approximately 15 separate phosphopeptides are present in a tryptic digest of Physarum histone H1. The core histones from Physarum, histones H2A, H2B, H3, and H4, are rapidly acetylated; histone H4 shows five subfractions, analogous to the five subfractions of mammalian histone H4 (containing zero to four acetyllysine residues per molecule); histone H3 has a more complex pattern that we interpret as zero to four acetyllysine residues on each of two sequence variants of histone H3; histones H2A and H2B show less heterogeneity. Overall, the data show that Physarum microplasmodia have a set of histones that is closely analogous to mammalian histones.     

Amphibian oocyte maturation induced by extracts of Physarum polycephalum in mitosis

The orderly progression of eukaryotic cells from interphase to mitosis requires the close coordination of various nuclear and cytoplasmic events. Studies from our laboratory and others on animal cells indicate that two activities, one present mainly in mitotic cells and the other exclusively in G1-phase cells, play a pivotal role in the regulation of initiation and completion of mitosis, respectively. The purpose of this study was to investigate whether these activities are expressed in the slime mold Physarum polycephalum in which all the nuclei traverse the cell cycle in natural synchrony. Extracts were prepared from plasmodia in various phases of the cell cycle and tested for their ability to induce germinal vesicle breakdown and chromosome condensation after microinjection into Xenopus laevis oocytes. We found that extract of cells at 10-20 min before metaphase consistently induced germinal vesicle breakdown in oocytes. Preliminary characterization, including purification on a DNA-cellulose affinity column, indicated that the mitotic factors from Physarum were functionally very similar to HeLa mitotic factors. We also identified a number of mitosis-specific antigens in extracts from Physarum plasmodia, similar to those of HeLa cells, using the mitosis-specific monoclonal antibodies MPM-2 and MPM-7. Interestingly, we also observed an activity in Physarum at 45 min after metaphase (i.e., in early S phase since it has no G1) that is usually present in HeLa cells only during the G1 phase of the cell cycle. These are the first studies to show that maturation-promoting factor activity is present in Physarum during mitosis and is replaced by the G1 factor (or anti-maturation-promoting factor) activity in a postmitotic stage. A comparative study of these factors in this slime mold and in mammalian cells would be extremely valuable in further understanding their function in the regulation of eukaryotic cell cycle and their evolutionary relationship to one another.     

Calcium and malate are sporulation-promoting factors of Physarum polycephalum

Fruiting body formation (sporulation) is a distinctive, irreversible differentiation process in the life cycle of the slime mold Physarum polycephalum. The most important requirement for sporulation of Physarum is a period of starvation, and normally sporulation proceeds in the light. It is shown here that by omitting the liquid sporulation medium and elevating the temperature from 21 to 25 degrees C, sporulation can occur routinely in the dark. It is further shown that this autocrine signaling in the dark requires calcium ions and malate. A putative sporulation control factor was detected in conditioned media derived from plasmodia starved in the dark, which was then identified as polymalate. As an additional role for this previously detected polyanion, specific for the plasmodial state of Physarum, it is suggested that the secreted compound serves as a source for both malate and calcium ions and thus promotes sporulation without light signaling.     

Comparative study of histones from slime mold Physarum polycephalum and calf thymus

The histones from slime mold Physarum polycephalum and calf thymus were characterized in terms of some physico-chemical properties. The molecular weights of six principal histone fractions of Ph. polycephalum were found to be the following: P1--22 700, P3--15 700, P4a--15 000, P4b--14 300, P5--12 800 and P6--10 500. Electrophoretically homogenous histone fractions H1, H2b and H4 of calf thymus and histones P1, P3, P4b and P6 of slime mold were obtained by gel-filtration on Acrylex P-60. These findings suggest that fractions P1, P4a, P4b, P5 and P6 of slime mold Ph. polycephalum are homologus with respect to the histone fractions H1, H3, H2b, H2a and H4 of calf thymus. Only fraction P3 has no corresponding fraction in the calf thymus histones; a fraction corresponding to histone P3 of slime mold was absent.