Growth regulation inCoprinus radiatus

Some of the morphological and physiological parameters of stipe growth or elongation inCoprinus radiatus were investigated. During the development of the fruit body the number of cells in a row in the growing portion of the stipe doubled during the development of the button, and again during the phase of rapid stipe elongation. Also during the stage of rapid elongation the cells in the upper 2/3 of the stipe increased 6–8 fold in length. The existence of a growth regulator synthesized in the cap and exerting control over the stipe was demonstrated through decapitation experiments. The cap appears to be required for normal stipe development until the stipe reaches about 1/4 of its final length. Through decapitation and cap-stipe exchanges it was found that the cap produced growth regulator up to the time of autodigestion; however, the stipe responded to the regulator only during a brief period at the onset of elongation.     

Behavior of chromosomes after meiosis inCoprinus cinereus

The movement was investigated of a specific chromosome in the F₁ progeny of the basidiomyceteCoprinus cinereus. We focussed our attention on the smallest chromosome of the 5302 strain. We first constructed a chromosome-smallest library and screened it with a chromosome-specific clone, pRC 1. The pRC 1 probe hybridized only with the smallest chromosome of the 5302 strain, and it detected one band of different mobility in two parental strains. In the F₁ progeny, the probe hybridized with one to three chromosomes. Most of the hybridized chromosomes in the F₁ progeny were positioned in terms of mobility between the hybridized chromosomes of the two parental strains. Therefore, they were probably generated by meiotic recombination between homologous chromosomes of different sizes.     

Inheritance of chromosome length polymorphisms inCoprinus cinereus

The sexually compatible strains ofCoprinus cinereus 5302 and Dd 13 revealed chromosome length polymorphisms in their electrophoretic karyotypes. The dikaryon derived from two monokaryons contained a mixture of the two electrophoretic patterns. F₁ progenies were isolated by crossingC. cinereus 5302 and Dd 13 strains and it showed unique karyotypes. Chromosome length polymorphisms of both parental strains were inherited at random in the F₁ progenies. As a result, several novel electrophoretic karyotypes which had not been observed in either parental strains were found in the F₁ progeny. The rDNA probe hybridized with one chromosome in both parental strains, with two chromosomes in the hybridization pattern of both parental strains in the dikaryon, and with one to two chromosomes in the F₁ progenies. The relation between mating type and hybridization pattern has thus not been made clear in the case of F₁ progeny.     

Partial contig map of the smallest chromosome inCoprinus cinereus

We have constructed a chromosome-specific cosmid library from electrophoretically separated chromosomes of the basidiomyceteCoprinus cinereus and performed contig mapping and analysis of chromosome length polymorphisms (CLPs) for the smallest chromosome of the 5302 strain. A contig map of about 300 kb indicated that the novel size chromosomes in the F₁ progeny were apparently recombinants containing physical markers derived from both ends and central regions in this map. This may be the first case in which the formation of CLPs in the F₁ generation has been explained using the contig map. The results obtained were consistent with the hypothesis that novel CLPs were produced by meiotic recombination between the parental homologous chromosomes of unequal sizes.     

Darkness-induced factor affecting basidiocarp maturation in Coprinus macrorhizus

Graftings were made between young basidiocarps of Coprinus macrorhizusgrown under different light programs. Results showed that adiffusible factor(s) affecting basidiocarp maturation was inducedby the darkness of phase 3. (Received July 3, 1979; )     

Photo-induced karyogamy in a Basidiomycete, Coprinus macrorhizus

In Coprinus macrorhizus, the fruit-body matured when two 2-hrlight periods were given separately after the formation of theprimordium. The karyogamy may be triggered by the exposure tothe second light, because fusion of two nuclei in the basidiumwas always observed 12 hr after exposure to the second light,irrespective of the time when given. (Received August 15, 1973; )     

Cytological studies on sporeless mutant in the basidiomyceteCoprinus macrorhizus

Meiotic events in the sporeless mutant, NG107, and in the wild-type dikaryon of the basidiomyceteCoprinus macrorhizus were studied comparatively with light and electron microscopes in order to determine precisely which meiotic process was blocked in the mutant. In most basidia of the mutant, when incubated at 28 C, meiosis stopped at meta-anaphase I although the kinetic apparatus was formed. Then, the basidia became mononucleate, electron dense, and finally disintegrated. This mutant, when incubated at 20 C, produced basidiospores approximately one third of the wild-type. Basidia with irregular number of spores were observed in higher frequency than in the wild-type. Experimental results of temperature shifts from 28 C to 20 C and the reciprocal shift suggest that meta-anaphase I might be thermosensitive in the mutant.     

Effects of light on basidiocarp maturation in Coprinus macrorhizus

The effect of white light on basidiocarp maturation in Coprinusmacrorhizus was examined. The results obtained showed that basidiocarpmaturation was separable into five successive phases in regardto photosensitivity: phase 1 in which light triggered maturation;phase 2 in which light delayed maturation; phase 3 in whichlight stopped maturation; phase 4 in which light acceleratedmaturation; and phase 5 in which light had no effect upon theprogress of maturation. The relation between the light effect and the events duringspore formation including nuclear fusion, meiosis and sterigmaformation was examined with the following results: the lightof phase 1 initiated nuclear fusion; the light given in phase2 delayed the initiation of nuclear fusion and the later progressup to prophase 1; without the darkness of phase 3, meiosis neverproceeded beyond prophase 1; and the light given in phase 4accelerated the progress from the initiation of nuclear fusionto prophase 1. (Received August 15, 1977; )     

Effect of cyclic AMP on glycogen phosphorylase in Coprinus macrorhizus.

Glycogen phosphorylase (1,4-alpha-D-glucan:orthophosphate alpha-glucosyltransfase, EC 2.4.1.1) activity was found in mycelial extracts of Coprinus macrorhizus concurrently with decrease of glycogen content in mycelial cells. Incubation of the enzyme sample with cyclic AMP and ATP leads to a 3-fold activation of the glucogen phosphorylase activity. Activation of the enzyme partially purified through Sepharose 6B required a cellular fraction containing cyclic AMP-dependent protein kinase.     

Isolation and genetic analysis of resistant mutants to the benzimidazole fungicide benomyl inCoprinus cinereus

In total, 404 variants resistant to the antimicrotubule agent benomyl were isolated from UV-irradiated oidia of the basidiomyceteCoprinus cinereus. Part of the variants showed, in addition to benomyl resistance, heat sensitivity or heat dependence. Fifteen variants selected on the basis of different phenotypes were subjected to further analyses. All of the 15 variations were due to single-gene mutations, and the mutations comprised four groups (benA, benB, benC, andbenD) in terms of genetic linkage. Some of the 15 mutations affected nuclear migration in dikaryosis and/or fruiting processes.     

Sequential production of enzymes and basidiospore formation in fruiting bodies of Coprinus macrorhizus

The production of NADP-dependent glutamate dehydrogenase was initiated at the stage of first meiotic prophase in pileus cells but not in stipe cells of dikaryotic and monokaryotic fruiting bodies in Coprinus macrorhizus. The production of chitinase and glucanases assayed with laminarin and lichenan was observed after the completion of meiosis only in pileus cells. The light conditions that were effective for the delay or inhibition of cellular events in the pileus cells were also effective for the delay or inhibition of enzyme production. But all sporeless mutants tested, which were defective at the various stages of basidiospore formation, produced the normal levels of these enzymes. The results indicate that the sequential production of enzymes and cellular events leading to basidiospore formation in pileus cells are independent from each other.Abbreviation GDH_NADP NADP-dependent glutamate dehydrogenase     

Stipe elongation during basidiocarp maturation in Coprinus macrorhizus: Mechanical properties of stipe cell wall

Stipe elongation during basidiocarp maturation in the wild-type,#5026+5132, and the elongationless mutant, NG0398, of Coprinusmacrorhizus was studied, and the following results were obtained.

1. In the wild-type the middle zone of the stipe elongated 8.4times in 15 hr during maturation, while in the mutant it elongatedoaiy 2.2 times.
2. Component cells of the stipe elongated inparallel with thestipe elongation in both the wild-type andthe mutant. The widthof stipe cells was almost constant duringelongation in thewild-type, while it increased 2 times in themutant. Cell volumeincreased ca. 8 times in both stocks.
3. Theosmotic value of stipe cells was almost constant (0.45–0.50M) throughout elongation of both the wild-type and the elongationlessstipes.
4. Mechanical properties of the cell wall were examinedby measuringshrinkage, extensibility and minimum stress-relaxationtime(To) of the stipe during maturation. These parameters weredirectlyproportional to the elongation rate to follow.
5. Whenthe wild-type stipes were incubated in various concentrationsof mannitol solution and then in plain buffer solution, theextensibility of the stipe after the incubation in mannitolsolutions changed proportionally with the stipe length afterthe mannitol treatment, and To with the elongation capacityin plain buffer solution.

(Received March 3, 1977; )     

Relationship between deficiency of phosphoglucose isomerase in Coprinus macrorhizus and fruiting body formation.

A mutant (pgi) of Coprinus macrorhizus deficient in phosphoglucose isomerase did not grow on fructose and grew poorly on glucose. The pgi mutation inhibited the formation of monokaryotic and dikaryotic fruiting bodies.     

An efficient method for the isolation of mycelial protoplasts from Coprinus macrorhizus and other basidiomycetes

Summary A mixture of commercially available chitinase and cellulase released mycelial protoplasts of Coprinus macrorhizus in yields exceeding 10⁸/ml plasts from C. macrorhizus Fis^C regenerated hyphae and developed into normal fruiting bodies at frequencies of 20%–50%. The same method also released good yields of protoplasts from several other edible mushroom species.     

Timing of DNA replication during the meiotic process in monokaryotic basidiocarps of Coprinus macrorhizus

By applying the method of fluorescent microscopy to propidium iodide stained cells, change in the relative amount of DNA in a basidium was examined during the meiotic process in Coprinus macrorhizus. In the monokaryotic basidiocarp of the mutant strain Fis^c, mitotic DNA replication was first induced soon after a 3h-illumination period on the 10th day of culture, and subsequently meiotic DNA replication occurred after karyogamy.     

Intraspecific heterokaryon and fruit body formation in Coprinus macrorhizus by protoplast fusion of auxotrophic mutants

Summary Fusion of protoplasts of Coprinus macrorhizus mutants with different amino acid requirements resulted in the production of prototrophic clones at frequencies of 1–4% of the protoplasts surviving the fusion treatment. The frequencies were at least 200 times higher than those of the appearance of revertants. Few prototrophic colonies appeared also when the mutant protoplasts were individually subjected to fusion treatment, or when they were mixedly cultured without fusion treatment. It was thus concluded that intraspecific heterokaryons were formed by protoplast fusion.The auxotrophic mutants did not form fruit bodies when cultured singly or mixedly with each other. In contrast, the heterokaryons produced by protoplast fusion between the mutants of compatible mating types developed into fruit bodies with intermediate morphology of those of the strains from which the mutants were derived. Heterokaryons were also formed by fusion of mutant protoplasts with identical mating genotype, but they failed to form fruit bodies.Abbreviations PEG polyethyleneglycol - HEPES N-2-hydroxyethylpiperazine-N-2-ethane sulfonic acid