Identification of the telomeric sequence of the acellular slime molds Didymium iridis and Physarum polycephalum.

We have determined the telomeric DNA sequence of the acellular slime molds Didymium iridis and Physarum polycephalum. In both organisms the telomeres consist of tandem repeats of the hexamer 5'(TTAGGG)3'. This sequence was determined by cloning and sequencing the telomeric fragment of the linear extrachromosomal ribosomal DNA from Didymium, as well as direct end labeling and sequencing the rDNA from both organisms. Interestingly, this sequence is identical to the telomeric DNA sequence of the flagellated protozoan Trypanosoma brucei, and suggests that despite the diversity of telomeric sequences previously determined in lower eukaryotes, the necessity to create functional telomeres has led to constraints on these sequences.     

Insertional Editing of Mitochondrial tRNAs of Physarum polycephalum and Didymium nigripes

tRNAs encoded on the mitochondrial DNA of Physarum polycephalum and Didymium nigripes require insertional editing for their maturation. Editing consists of the specific insertion of a single cytidine or uridine relative to the mitochondrial DNA sequence encoding the tRNA. Editing sites are at 14 different locations in nine tRNAs. Cytidine insertion sites can be located in any of the four stems of the tRNA cloverleaf and usually create a G·C base pair. Uridine insertions have been identified in the T loop of tRNA^Lys from Didymium and tRNA^Glu from Physarum. In both tRNAs, the insertion creates the GUUC sequence, which is converted to GTΨC (Ψ = pseudouridine) in most tRNAs. This type of tRNA editing is different from other, previously described types of tRNA editing and resembles the mRNA and rRNA editing in Physarum and Didymium. Analogous tRNAs in Physarum and Didymium have editing sites at different locations, indicating that editing sites have been lost, gained, or both since the divergence of Physarum and Didymium. Although cDNAs derived from single tRNAs are generally fully edited, cDNAs derived from unprocessed polycistronic tRNA precursors often lack some of the editing site insertions. This enrichment of partially edited sequences in unprocessed tRNAs may indicate that editing is required for tRNA processing or at least that RNA editing occurs as an early event in tRNA synthesis.     

Apogamic induction of haploid plasmodia in a myxomycete,Didymium iridis

Interisolate crosses between haploid (mean DNA = 0.32) CR 5-5 (A²) myxamoebae and polyploid (mean DNA = 1.80) CR 2–25 (A⁵) myxamoebae of the myxomycete Didymium iridis result in plasmodia that have the haploid (mean DNA = 0.32) DNA content rather than the predicted polyploid value. F₁ clones possess the mating type allele of the CR 5-5 clone only, and they also have the same mean DNA content as CR 5-5 myxamoebae. Crosses between these F₁ clones and CR 2–25 myxamoebae again resulted in the production of haploid plasmodia. Hence, the polyploid CR 2–25 clone appears to induce the CR 5-5 clone to produce plasmodia without involving itself in nuclear fusion.     

Patterns of mitochondrial inheritance in the myxogastrid Didymium iridis

Seven strains of the Central American A1 mating series of Didymium iridis were crossed in all possible combinations. Individual plasmodia were isolated and grown to a stage where total DNA could be isolated for DNA-DNA hybridization with cloned mitochondrial DNA probes to determine the pattern of mitochondrial inheritance. Random, biased, and dominant patterns of uniparental mitochondrial inheritance were observed, as well as rare cases of biparental inheritance, depending on the particular parental strains mated. The diverse patterns suggest that the factors controlling mitochondrial inheritance in D. iridis are complex. Differences between trials of the same matings suggest that non-genetic factors may also influence mitochondrial inheritance.     

Mutagenesis and mutant selection in Physarum polycephalum

Summary Physarum polycephalum myxamoebae were exposed to ultraviolet irradiation and plated in the absence and presence of caffeine. Caffeine reduces the shoulder on the UV¹ dose-survival curve, thereby increasing the UV-sensitivity for survival. Caffeine alone is a moderate mutagen. Used in conjunction with UV a strong mutagenic action is observed. Active growth is required for both of these mutagenic actions.Populations of Physarum myxamoebae mutagenized with NMG or EMS could be enriched for two classes of mutants by incubating at high temperature (30°C) with 5-bromodeoxyuridine-substituted bacteria followed by irradiation with long wave UV light and recovery at low temperature (23°C). One class of mutants was obtained in high yields after repeated cycles of light inactivation. These are not heat sensitive. Rather they are defective in utilization of DNA precursors provided by the bacteria. The other mutant class, obtained in low yields after limited selection, are heat sensitive. Three independent mutants of this kind, all eaky, were obtained. Reconstruction experiments show that all are selectants.     

Control of chemotaxis in Physarum polycephalum

Plasmodia migrate towards those situations which increase the frequency of their alternations in streaming, and away from those which decrease the frequency. Therefore peristalsis-like waves in Physarum move in the direction opposite from the net movement of the organism. The mechanism is fundamentally related to other known types of chemotaxis.     

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.     

Ultrastructural events and kinetics of microcyst germination in the myxomycete Didymium iridis

Microcysts of the myxomycete Didymium iridis were induced to excyst by transfer to 5mM potassium phosphate buffer. After 1 h in suspension, 90% of the microcysts had germinated into myxamoebae distinguishable by phase contrast microscopy and staining with Lugol's iodine. Both pH and osmolarity affected the kinetics of excystment. The rate and extent of excystment were decreased by cycloheximide but remained unaffected by actinomycin D, suggesting a requirement for protein synthesis but not RNA synthesis. Initially, the outer wall layers separated from the inner layer, which gradually expanded and loosened. The protoplast rehydrated and reverted to a vegetative morphology. Excysting cells were characterized by nucleolar inclusions, changes in the nuclear envelope and plasma membrane, appearance of ringed cisternal elements and microbodies in the cytoplasm, and formation of a densely fibrous zone adjacent to the site of emergence. Excysting populations have been classified into characteristic stages: mature, initiated, swollen, and pre-emergent microcysts.Florida Agricultural Experiment Station Journal Series No. 2407