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Physarum polycephalum is a species of slime mold. Physarium is one of the three main groups of slime molds. Physarum takes the form of a syncytium, a giant single cell with thousands of nuclei called a plasmodium. The plasmodium form is sometimes referred to as a giant amoeba. P. polycephalum is also called an acellular type of slime mold, as opposed to cellular slime molds which are made up of many individual cells. The plasmodium is covered in a layer of glycoprotein gel, which gives it a slimy appearance.
Plasmodium moves by extension and retraction of multiple tubular extensions called pseudopods, a feature for which it got the Latin name polycephalum meaning multi-headed. P. polycephalum are useful for research on the movement of cell contents, called cytoplasmic streaming because the process can be viewed under relatively low magnification. This slime mold is a model system used by biochemists, cell biologists, chemists, developmental biologists, geneticists and physicists.
Slime molds are found in moist forest environments, likely feeding on microbes that grow in leaf litter, bark, mushrooms and other substrates. Physarum and it’s related species are found throughout the world with highest concentrations in Europe, North America and Japan. In their natural environment Physarum can grow up to several feet in diameter.
Slime molds superficially resemble fungi and were once classified as such in Myxomycophyta but they are now known to be unrelated to fungi. P. polycephalum was first described by von Schweinitz in 1822 and have a long-standing tradition in research in Japan. The Japanese emperor Hirohito who reigned from 1926-1989 was known to be fascinated by slime molds, and wrote a book devoted to them under an alias.
Before advanced techniques in cell culture and tissue engineering, P. polycephalum was a useful model organism that could be grown axenically (i.e. in pure culture) of up to 100 liters. The organism could provide a large amount of material for biochemical analysis. Cancer researchers tested antitumor compounds on P. polycephalum. By the 1970s and 1980s, progress in mammalian and bacterial cell culture exceeded that of P. polycephalum.
P. polycephalum belongs to Amoebozoa, the sister group to Opisthokonts which include fungi and animals. Together these two branches form the supergroup Amorphea.
Slime molds are neither plants, animals or fungi but share traits in common with each of these groups. For example, P. polycephalum forms a dormant state under non-favorable conditions, called a sclerotium, which contains cellulose in the walls, a molecule also found in plants. The sporangia of the slime mold resemble the fruiting bodies of fungi and similar to fungi, P. polycephalum feed on decomposing plants. In the plasmodial form, Physarum ingests food using phagocytosis, a process by which cells take in large particles into membrane-bound vesicles. Physarum also secretes enzymes to break down materials which are absorbed by a similar process called pinocytosis.
The physiology of P. polycephalum’s photosensing shares aspects with both plants and animals. Common to animals, P. polycephalum displays light-induced sporulation pathways and negative phototactic motility responses.
The pigments which give P. polycephalum its yellow color are believed to be unique to the organism. Several pigments have been identified and some of them serve as photoreceptors.
P. polycephalum is a member of the myxogastria class of slime molds which are characterized by the formation of the syncytial plasmodia. Myxogastria have common ancestry to cellular slime molds such as Dictyostelia discoideium. The main tool of classification in myxogastria is the fruiting body characteristics and evolutionary reconstruction is based on DNA sequence data. According to molecular evidence, the genus Physarum is not uniform and Badhamia utricularis is a close relative of P. pohycephalum.
Figure from Schaap P. et al. (2016) Genome Biol. Evol. 8(1):109-125
P. polycephalum has both a heterothallic and apogamic life cycle as represented by the outer and inner circle, respectively in the figure above. Mononucleate amoebae have four developmental options: To propagate by open mitosis or differentiate into a flagellate, cyst or multinucleate plasmodium. In the heterothallic cycle two amoebae of different mating type mate with each other to form a mononucleate diploid zygote and the zygote grows without cell division developing into a diploid multinucleate plasmodium where nuclei divide synchronously with their nuclear envelopes intact, also known as closed mitosis. The plasmodium continues to grow as long as environmental conditions are favorable. A mature multinucleate plasmodium can develop into a sclerotium to survive drought. A starved plasmodium can be induced by visible light or heat shock to begin sporulation and form fruiting bodies. Each sporangium has hundreds of haploid, mononucleate spores that have been formed by meiosis.
In the apogamic cycle, mononucleate amoeba that carry a gadAh mutant allele may develop into a multinucleate, haploid plasmodium without mating. Since apogamic development can be suppressed experimentally by elevated temperature, amoebal clones can be propagated. A diploid plasmodium can be made by mating two amoebae of different mating types.
The plasmodium forms as tubular network and as it moves the network reshapes and adapts to environmental stimuli. Cytoplasm streams through the networks and change direction periodically. P. polycephalum plasmonia can move up to a speed of 5 cm/h. This slime mold can grow to cover an area of up to 900 cm2. While feeding, P. polycephalum plasmodia extrude calcium and indigestible residue which act as foraging stimuli that are attractive to other plasmodium cells. Chemotactic orientation is induced by calcium. Two plasmodia can fuse if they carry identical genotypes.
In the laboratory smaller models of P. polycephalum are often needed for microscopy investigations and biochemical experiments. Puncturing a vein results in the release of endoplasm and formation of protoplasmic drops which rapidly create a new plasma membrane and the formation of invaginations and vacuoles. P. polycephalum can be maintained in constantly agitated axenic cultures whereby shear forces create microplasmodia. When microplasmodia are plated on solid agar, they fuse and form a connected network, with a shape that depends on environmental parameters such as substrate softness, nutrient and repellent concentrations. P. polycephalum’s genome is comprised of 188 million nucleotides, encoding 34,000 genes, 50 percent more genes than in the human genome.
Analysis of P. polycephalum genome revealed that tyrosine kinase-mediated signaling emerged early in evolution and was lost in the genomes of fungi and other amoebozoa such as D. discoideium. The evolution of tyrosine kinase-mediated signaling was once thought to be a pivotal step in the evolution of multicellularity in animals since it plays such an important role in embryogenesis and physiology. The presence of tyrosine kinase in P. polycephalum supports the likelihood that tyrosine kinases were already present in the last common ancestor to eukaryotes.
Genomic analysis suggest P. polycephalum is a prototypical eukaryote with features attributed to the last common ancestor of Amorphea. Sequencing of P. polycephalum shows it to have higher molecular complexity than other Amoebozoa and also displays many features of animal cells related to the cross-talk of signaling molecules and resulting dynamic behavior.
P. polycephalum has been used to study molecules that drive growth and development. Sporulation and transition from amoeba to plasmodia is form of cell differentiation. Cell cycle regulation and DNA replication are also areas of research that use P. polycephalum as a model organism. The large size facilitates research on the cytoskeleton and locomotion, environmental sensing and response (photo-chemotaxis).
P. polycephalum maintains its numerous nuclei with high genetic stability with a synchronous mitotic cycle. A diploid nucleus in Physarum contains about 200 linear minichromosomes, each about 60 kbp in length. It is possible to hybridize different strains of P. polycephalum to facilitate the generation and analysis of mutants. As of 2017, transformation of plasmid DNA into P. polycephalum as not been realized but it has become possible in D. discoideum.
P. polycephalum is studied to understand the mechanisms of long-range fluid flow in organisms, which is essential for spreading resources and signals to different areas. Internal movements in P. polycephalum are visible even to the naked eye. Shuttle streaming also known as cytoplasmic streaming is the movement of endoplasm (fluid cytoplasm) through veins and the conversion from endo- to ectoplasm (gel-like rigid cytoplasm) in certain zones. When Physarum forages the forefront of the organism takes on a fanlike configuration. The cell contents stream back and forth at approximately 60-second intervals through a network of vein-like tubes. The tubes can be up to 1 mm in diameter. As the organism moves the network of tubes is reorganized.
Shuttle-streaming and responses to the environmental stimuli are coupled. The problem-solving abilities of P. polycephalum are driven by a coupled-oscillator based sensorimotor system present in its membrane. Contractions are at the organismal level such that the protoplasm flows rhythmically back and forth throughout the cell. In response to quality of the local environment and the contraction intensities of the neighbouring regions, individual contractile regions change their contraction intensity. When P. polycephalum encounters a food source, intensity increases and when it encounters a repulsive stimuli such as bright light, it decreases. This results in a change in the pattern of contractions throughout the cells and movement either away or towards the stimuli.
In shuttle streaming there are contraction-relaxation cycles driven by F-actin filaments and the motor protein myosin. Contraction of the actomyosin network creates hydraulic pressure gradients. Other proteins called actin-binding proteins play a role in the functioning and reorganization of the actomyosin network. What triggers the oscillations and drives coordinated contractions is not known.
Actomyosin threads can be generated by squeezing out the endoplasm into water, enabling in vitro studies of actin and myosin over several contraction and relaxation cycles. This method was useful for analysing the molecular events such as the role of ATP in cell motility.
Physarum makes behavioral decisions that are more complex than the localized decisions of its parts. Information processing in single celled organisms such as P. polycephalum have been suggested to represent a simple precursor of brain-dependent higher functions. P. polycepahlum is described as displaying behavioral intelligence.
P. polycephalum does not have brain but it is considered to have an internal and ‘external memory’, the latter refers to the ability of the slime mold to recognize its own trail. During foraging, when the slime mold encounters slime deposits, it avoids those already covered areas. P. polycephalum has been shown to solve mazes and geometric puzzles. P. polycephalum can perform risk-management tasks and chooses an optimal choices, which hints at a rudimentary form of intelligence.
Maze-solving of P. polycephalum was demonstrated by Japanese researcher Toshiyuki Nakagaki. After allowing the plasmodium to cover an extensive labyrinth, he placed oat flakes at the entry and exit. The slime mold quickly rearranged its veins until the most efficient path between the two food sources remained. P. polycephalum also can solve a maze without previously exploring the whole area as long as a chemo-attractant is placed at the destination site. In a later experiment, Nakagaki and coworkers made the arrangement of oat flakes match the geography of Tokyo and its surrounding cities and the plasmodium in growing to reach out and colonize each of the food sources formed a network of veins with a similar shape to the existing railway system. Many man-made structures and highways and wireless sensor networks have been simulated by P. polycephalum.
Photographs by Professor Toshiyuki Nakagaki published in Sun, Y. (2019) arXiv
The above figure shows the tubular structure of P. polycephalum (a). Intelligent behaviors such as following the shortest path in a maze (b) and building high-quality networks that connect multiple food points (c) are shown. Nakagaki won an Ig Nobel Prize for cognitive science in 2008 for his experiments where slime molds find the minimum-length solution between two points in a labyrinth.
Although it is known that P. polycephalum can communicate across its entire body, the nature of the communication is not known and elastic waves, flow of a molecular stimulus or electrical impulses have been proposed. Calcium is a likely candidate for a signaling molecule used to communicate information. It is known to regulate actin-myosin dynamics and these drive oscillations in the network of tubes. It has been shown using a mathematical model that the complex movement behavior of P. polycephalum can be coordinated using a simple feedback mechanism. In this model an external stimulus triggers the release of a signaling molecule that is transferred with the cytoplasmic flows. As the molecule moves, it increases local contraction amplitude and generates additional cytoplasmic flows to carry itself further into the network, where it increases local contraction.
P. polycephalum can display habituation, a decrease in aversive behavior to an iterative stimulus, for unpleasant stimuli such as quinine, caffeine and salt. The adaptive response to salt was shown to be transferred between slime molds by cytoplasm mixing when a habituated cell fused to a naïve individual.
P. polycephalum is able to anticipate the timing of periodic events. After a plasmodium was exposed to dry conditions at repeated intervals of time as it moved along a narrow lane, the slime mold demonstrated anticipation of the condition change. The plasmodium slowed down its movement at the time when the dry condition would have been applied even when it was not.
P. polycephalum has been used in sensing, computing and robotic devices. Biologically inspired algorithms can be extracted from observing formations built by the slime mold. A network of veins formed by a P. polycephalum plasmodium can be described as the outcome of natural biological computation in order to create a robust and efficient network.
The foraging behavior of the P. polycephalum plasmodium can be thought of as computation where data are represented by spatial configurations of attractants and repellents and the results are represented by the structure of the protoplasmic network of tubes. P. polycephalum is also considered a reaction-diffusion medium or an excitable medium encapsulated in a biological membrane that can be used in the design of sensing and computing devices.
Logic units that process information have been built with living slime molds. Research lead by Adrew Adamatzky and Theresa Schubert demonstrated the tube networks of P. polycephalum plasmodia can absorb and transport different colored dyes and mix them to make a third color which can serve as an “output”. P. polycephalum slime molds were incorporated into a “lab-on-a-chip” device that used dyes with magnetic nanoparticles and tiny fluorescent beads. The researcher’s slime mold network of tubes were demonstrated to carry out XOR or NOR Boolean operations. Chaining together arrays of logic gates could allow a slime mold computer to carry out binary operations.
A research team led by Dianella Howarth at St. John’s University, New York demonstrated a way to program computing behavior of P. polycephalum using hairy roots induced in Valeeriana officinalis plants by infection with Agrobacterium rihizogenes. As a plant cell culture system, the hairy roots can be cloned and propagated and elicit a positive chemotactic response from the slime mold to modulate its activity. The electrophysiological response to hairy root culture biomass can be measured by the plamodium’s electrical activity. An example of their experimental circuit is shown below.
From Riciglioano V. (2015) A circuit with plasmonium inoculated agar blobs on aluminum electrodes with a protoplasmic tube connecting them.
P. polycephalum was aboard the ‘Challenger’ and further experiments in space were performed by German astronaut Ulf Merbold to study its reaction to gravity (graviresponse).
P. polycephalum is in the Guinness Book of Records (1989) as the largest single cell ever grown. This 3 kg, 5.5 m2 specimen was grown in a W shape on oak flakes in honor of Karl-Ernst Wohlfarth-Bottermann, then Director of University of Bonn (Germany) Institute of Cytology and Microbiology.
P. polycephalum became an organism on display at the Paris zoo in 2019 and was named “the blob” after the 1958 science-fiction horror film about an alien life form that consumes everything in its path.
