Programmed ascospore death in the homothallic ascomycete Coniochaeta tetraspora

Immature asci of Coniochaeta tetraspora originally contain eight uninucleate ascospores. Two ascospore pairs in each ascus survive and mature, and two die and degenerate. Arrangement of the two ascospore types in individual linear asci is what would be expected if death is controlled by a chromosomal gene segregating at the second meiotic division in about 50% of asci. Cultures originating from single homokaryotic ascospores or from single uninucleate conidia are self-fertile, again producing eight-spored asci in which four spores disintegrate, generation after generation. These observations indicate that differentiation of two nuclear types occurs de novo in each sexual generation, that it involves alteration of a specific chromosome locus, and that the change occurs early in the sexual phase. One, and only one, of the two haploid nuclei entering each functional zygote must carry the altered element, which is segregated into two of the four meiotic products and is eliminated when ascospores that contain it disintegrate. Fusion of nuclei cannot be random-a recognition mechanism must exist. More study will be needed to determine whether the change that is responsible for ascospore death is genetic or epigenetic, whether it occurs just before the formation of each ascus or originates only once in the ascogonium prior to proliferation of ascogenous hyphae, and whether it reflects developmentally triggered alteration at a locus other than mating type or the activation of a silent mating-type gene that has pleiotropic effects. Similar considerations apply to species such as Sclerotinia trifoliorum and Chromocrea spinulosa, in which all ascospores survive but half the spores in each ascus are small and self-sterile. Unlike C. tetraspora, another four-spored species, Coniochaetidium savoryi, is pseudohomothallic, with ascus development resembling that of Podospora anserina.     

Suppressed recombination and a pairing anomaly on the mating-type chromosome of Neurospora tetrasperma

Neurospora crassa and related heterothallic ascomycetes produce eight homokaryotic self-sterile ascospores per ascus. In contrast, asci of N. tetrasperma contain four self-fertile ascospores each with nuclei of both mating types (matA and mata). The self-fertile ascospores of N. tetrasperma result from first-division segregation of mating type and nuclear spindle overlap at the second meiotic division and at a subsequent mitotic division. Recently, Merino et al. presented population-genetic evidence that crossing over is suppressed on the mating-type chromosome of N. tetrasperma, thereby preventing second-division segregation of mating type and the formation of self-sterile ascospores. The present study experimentally confirmed suppressed crossing over for a large segment of the mating-type chromosome by examining segregation of markers in crosses of wild strains. Surprisingly, our study also revealed a region on the far left arm where recombination is obligatory. In cytological studies, we demonstrated that suppressed recombination correlates with an extensive unpaired region at pachytene. Taken together, these results suggest an unpaired region adjacent to one or more paired regions, analogous to the nonpairing and pseudoautosomal regions of animal sex chromosomes. The observed pairing and obligate crossover likely reflect mechanisms to ensure chromosome disjunction.     

Meiotic silencing in the homothallic fungus Gibberella zeae

The homothallic ascomycete fungus Gibberella zeae is an important pathogen on major cereal crops. The objective of this study was to determine whether meiotic silencing occurs in G. zeae. Cytological studies demonstrated that GFP and RFP-fusion proteins were not detected during meiosis, both in heterozygous outcrosses and homozygous selfings. The deletion of rsp-1, a homologue used for studies on meiotic silencing of Neurospora crassa, triggered abnormal ascospores from selfing, but outcrosses between the mutant and wild-type strain resulted in some ascospores with mutant phenotype (low occurrence of ascus dominance). When the ectopic mutants that carried an additional copy of rsp-1 were selfed, they primarily produced ascospores with normal shape but a few ascospores (0.23 %) were abnormal, in which both endogenous and ectopically integrated genes contained numerous point mutations. The ectopic mutants showed low occurrence of ascus dominance in outcrosses with strains that carried the wild-type allele. Approximately 10 % of ascospores were abnormal but all of the single-ascospore isolates produced normal-shaped ascospores from selfing. However, no ascus dominance was observed when the mutants were outcrossed with a sad-1 deletion mutant, which lacks the putative RNA-dependent RNA polymerase essential for meiotic silencing in N. crassa. All results were consistent with those generated from an additional gene, roa, required for ascospore morphogenesis. This study demonstrated that G. zeae possesses a functional meiotic silencing mechanism which is triggered by unpaired DNA, as in N. crassa.     

Hypoxylon cohaerens: Cytology of the ascus

The haploid chromosome number ofHypoxylon cohaerens apparently is 5, based on counts made at meiotic prophase and meiotic and mitotic metaphases. Newly formed ascospores are at first uninucleate, becoming binucleate following mitosis in the ascospore. Subsequently, one of the two nuclei disappears. Maturing ascospores are uninucleate.Scientific Paper No. 3732 Washington State University. College of Agriculture, Project 1767. This study was supported in part by National Science Foundation Grant GB-19924.     

A cinematographic study of meiosis in salamander spermatocytes in vitro

Summary Salamander spermatocytes were isolated in a modified Eagle's medium in Rose Chambers. The behavior of the spermatocytes during meiosis was recorded on a time lapse, phase contrast film. The two meiotic divisions progressed without visible irregularities in freshly isolated spermatocytes. Times required for the various meiotic events were obtained. Spermatocytes four days in vitro carried out the first meiotic division, but there were many abnormalities and the second meiotic division did not occur. At first meiotic metaphase, whole bivalent oscillations were accompanied by a relatively higher frequency oscillatory movement of the two homologous kinetochore regions. Oscillations of the kinetochore region were independently variable in magnitude and frequency. A system is proposed by which the metaphase bivalent movements are explained in terms of two pulling forces acting with variable intensity and frequency in opposite directions at the two homologous kinetochores. Meiosis in heavily compressed spermatocytes was blocked at the first meiotic metaphase, apparently because of the absence of a bipolar meiotic apparatus. In compressed spermatocytes, the centrosome divided but the two resulting centrosomes failed to reach their definitive polar positions. After about two hours of separation, the two centrosomes reversed their movement and fused to form a single centrosome from which a unipolar half-spindle radiated.This investigation was supported by grant GB-15 from the National Science Foundation and by Public Health Service Research Grant GB 12431-02 from the Division of General Medical Sciences.Deceased June 17, 1964.     

Theory of hemoglobin facilitated oxygen transport

The transport of oxygen in a hemoglobin-saturated medium is theoretically investigated using classical transport theory. It is found that all the chemical complexes can be expressed as a single function of oxygen pressure. A potential difference together with apH shift is predicted to occur across the medium. This research was supported by the United States Public Health Service Training Grant No. 5-T1-GM-833 from the National Institute of General Medical Sciences. This research was supported by a United States Public Health Service Research Career Program Award 5-K6-GM-18,420 from the National Institute of General Medical Sciences.     

Use of Neurospora spore killer strains to obtain centromere linkage data without dissecting asci

Use of a centromere-linked Spore killer gene Sk reduces manyfold the labor involved in obtaining tetrad data that would otherwise require ordered dissection of intact linear eight-spored asci. Heterozygous crosses are made for Spore killer (SkK X SkS) and for markers to be tested. In such crosses only SkK ascospores survive. The four viable (SkK) and four aborted (SkS) ascospores of each ascus are ejected from the perithecium as a physically disordered group. The four surviving SkK ascospores of individual asci are germinated and scored. SkK segregates from SkS at the first meiotic division. If both marker alleles are represented in the surviving products, they must therefore have segregated from one another at the second division. Four-spore (Fsp) genes have been used to eliminate one postmeiotic nuclear division, so that only two ascospores per ascus need to be scored. The Spore killer method has been useful for mapping closely linked genes in centromere regions, for identifying genes that are far out on chromosome arms, for obtaining information on meiotic crossing-over, and for comparing linkages in different species.     

The diffuse diplotene stage of meiotic prophase in Neurospora

The prophase stages of meiosis in Neurospora crassa are re-examined following McClintock (1945) and Singleton (1953). A diffuse chromosome stage occurring between pachynema and diakinesis is described. It is proposed that the diffuse stage does not necessarily represent a condition of intense gene activity in the sense of directing the metabolic activity of the ascus.Supported by U. S. Public Health Service grants AI-01462 and GM-14263.     

Morphology,development and cytology of Taphrina maculans Butler

Morphology, development and nuclear behavior of the ascogenous stroma and asci in the infection spots have been described inTaphrina maculans Butler. The fungus forms subcuticular and intercellular mycelium in the leaf tissues and the ascogenous layers originate through division of the subcuticular hyphal cells in the infection sites. Germination of ascogenous cells starts with their elongation in the uppermost layer forming asci and ascospores without formation of stalk cells. Meiosis of the fusion (diploid) nucleus occurs in the young ascus as in otherTaphrina species devoid of stalk cells. The haploid chromosome complement in this species consists of 3 chromosomes (n=3). All the cells in the stromatic layer are potential ascogenous cells and ascus formation continues, until all of them are exhausted in the infection spot. Eight ascospores are normally formed in each ascus, but multi-plication of ascospores may occurin situ later. Three morphologically distinct types of ascus opening are encountered, which are apparently not correlated with prevalent environment. Multiplication of ascospores after their discharge from mature asci occurs by budding proceded by a mitotic division of the spore nucleus. Blastospores (budded cells) germinate into short hyphae and binucleate condition of cells originates by mitotic division of the nucleus. Occurrence of giant cells containing 2 nuclei is often observed. Possible origin of Uredinales fromTaphrina-like ancestors has been indicated due to their close resemblance.     

Ascospore aggregation and oxylipin distribution in the yeast Dipodascopsis tothii

Upon cultivation of the yeast Dipodascopsis tothii in its sexual stage, small ascospores are released individually from the ascus tip, which then assemble in sheathed cluster balls. In contrast to Dipodascopsis uninucleata, this yeast produced smooth bean shaped ascospores with sheath-like appendages that assemble in a disordered sheathed ball of ascospores outside the ascus. Strikingly, upon release, the ascus tip contained 3-hydroxy oxylipins, while the released ascospore clusters contained little or no 3-hydroxy oxylipins as indicated by immunofluorescence microscopy. In D. uninucleata, these oxylipins are concentrated on the spore surface and interspore matrix, but not on the ascus tip.     

Polarised complementation at the pyrimidine-3 locus of Neurospora

Summary A re-evaluation of complementation at the pyrimidine-3 locus ofNeurospora suggests the existence of polarity. This apparent polarity is supported by polar complementation of the majority of a number of mutants induced with the acridine ICR-170, and confirmed by the behavior of an allele which is shown by reversion studies to be a frameshift mutant. Supported by Public Health Service Grant AI-01462.     

A genetic analysis of linkage group I of Chlamydomonas reinhardi

Summary Crossing over in linkage group I ofChlamydomonas reinhardi was studied by means of the analysis of 1721 unordered tetrads. The data indicate that chiasma interference is positive, that chromatid interference is absent and that crossing over occurs at the 4-strand stage.

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Zusammenfassung Mit Hilfe der Analyse von 1721 ungeordneten Tetraden wurde das crossing-over in der Koppelungsgruppe I vonChlamydomonas reinhardi untersucht. Aus der Häufigkeit der verschiedenen Tetraden-Typen wird geschlossen, daß crossingover im Vierstrang-Stadium geschieht, und daß eine positive Chiasma-Interferenz existiert, während eine Chromatiden-Interferenz nicht vorkommt.

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This work was supported by the United States Public Health Service (Grant No. E 1421) and the National Science Foundation (Grant No. G 2812).

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A portion of this work was conducted while a Public Health Service Research Fellow of the National Cancer Institute.     

Genetic relationships between the multiple alcohol dehydrogenases of maize

There are three electrophoretically separable sets of alcohol dehydrogenase isozymes in maize. Previous work has shown that two of these isozymes (Sets I and II) share a subunit in common, since mutations in one of the Adh genes, Adh ₁, alter both isozymes. A mutation in the second Adh gene, Adh ₂, has now been induced and recovered. This mutant allele also alters two of the three isozymes—Sets III and II. Adh ₁ and Adh ₂ appear to segregate independently. Gel filtration data show that all ADH isozymes are indistinguishable in size. These findings support the hypothesis that the two Adh genes specify promoters which homo- and heterodimerize, yielding three types of ADH isozymes.This research was supported by National Science Foundation Grant GB 25594. M.F. is a recipient of Public Health Service Genetics Training Grant GM 82-12.     

Hemoglobin polymorphism in the deer mouse,Peromyscus maniculatus

Horizontal starch gel electrophoresis reveals three hemoglobin phenotypes in natural populations of the deer mouse, Peromyscus maniculatus,from northern Arizona. Phenotypic variation is controlled by a single locus. A sample of deer mice from Michigan was monomorphic for a multiple-banded phenotype. Hemoglobin polymorphism was also observed in Arizona samples of the species Peromyscus boylii.This investigation was supported by Public Health Service Research Grant R01-GM12190.     

Immunochemical cross-reactivity between intact purified myelin basic protein (MBP) and the synthetic encephalitogenic peptide S49

Three antisera to myelin basic protein—a rabbit antiserum pool against rat myelin, a rabbit antiserum pool against rat myelin basic protein (MBP), and a monkey antiserum against bovine MBP—were found to contain detectable levels of antibodies that would bind radiolabeled S49 (GSLPQKAQRPQDENG). Strongly encephalitogenic in Lewis rat, S49 is a synthetic peptide representing residues 69–84 of bovine MBP with a deletion of glycine-76 and histidine-77 to make it analogous to rat and guinea pig MBPs. The rabbit antimyelin antiserum and the monkey anti-MBP antiserum contained antibodies directed against a non-sequential determinant that required asparagine 84, the glycine-histidine deletion, and residues 69–71 for maximal activity. S49-reactive antibodies from the rabbit anti-MBP antiserum were directed solely against a sequential determinant comprising residues 69–71. S49-reactive antibodies from all three antisera reacted in liquid phase with purified intact rat, guinea pig, and bovine MBP showing that the determinant is exposed for B cell recognition even in bovine MBP and can serve both as immunogen and reactant.This work supported at Duke University Medical Center by Research Grant NS-10237 from the National Institutes of Health of the U.S. Public Health Service and the Medical Scientist Training Program Grant #5-T32-OMO-7171-08; at St. Luke's Hospital Center by NS-15322 from the National Institutes of Health of the U.S. Public Health Service; and at Northwestern University by Research Grant NS-06262 from the National Institutes of Health of the U.S. Public Health Service.     

The isolation and analysis of one functional type ofpyr-3 mutant inNeurospora

Twenty-seven pyrimidine-requiring mutants were isolated as suppressors of anarg-3 mutant. All 27 are deficient for ATCase activity and show linkage to thecol-4 marker located on linkage group IV. Analyses of prototroph frequencies resulting from crosses of the new mutants to previously mappedpyr-3 mutants indicate that this functional type ofpyr-3 mutant is restricted to one region of the genetic map. Complementation studies with 11 of the new mutants further extend and subdivide the complementation map of thepyr-3 locus.This work was supported in part by National Institutes of Health, Public Health Service Grant GM 15137-01 and by National Science Foundation Grant GB5998.     

Stimulatory effect of selected surface-active agents on the growth of pathogenic human fungi

Summary The growth ofCryptococcus neoformans andCandida albicans in Sabouraud's dextrose broth with added surface-active agents was measured by nephelometric methods. The surface-active agents was measured by nephelometric methods. The surface-active agents utilized were Polyethylene glycol 400 monolaurate, Pluronic L 64, and Mulsor 224. These three compounds definitely stimulated the growth of the above organisms as indicated in the accompanying tables. The nephelometric runs were corroborated with plate counts taken at suitable intervals.This project was supported by Research Grant E-1045(R) of the National Institute of Health, United States Public Health Service.     

Genetic analysis of lon mutants of strain K-12 of Escherichia coli

Summary Following UV irradiation of AB1157 31 mucoid ultraviolet light UV sensitive mutants were isolated. These were all induced to form filaments by UV irradiation, i.e. they had all the phenotypic properties of Lon mutants. These lon mutants fell into two phenotypic classes based on their sensitivity to UV. The gene determining UV sensitivity and mucoidy in all mutants of both Class A and Class B was cotransducible with proC. Intra-class crosses by Pl transduction yielded no UV resistant recombinants. Inter-class crosses yielded UV resistant nonmucoid recombinants, the frequency depending on the direction of the cross. The data imply two adjacent blocks in the lon region of E. coli and the order of markers in this region is probably proC tsx lon Class A lon purE Class B.This work was carried out under Public Health Service Grant CA 05687-08 from the National Cancer Institute.Recipient of a Public Health Service Career Development Award.     

Isozyme variability in species of the genus Drosophila. I. A multiple allelic isozyme system in Drosophila busckii: Inheritance and general considerations

Starch gel electrophoretic analysis of a triallelic leucine aminopeptidase polymorphism in a laboratory population of Drosophila busckii is described. The three alleles involved are expressed without dominance. A series of single-pair matings revealed an excess of heterozygous types in most segregating families, suggesting selection against the homozygous genotypes. A few cases of heterogeneity among progeny extracted from a single family were the result of matings that produced Mendelian ratios. These few cases had no clear genetic explanation, but there is a suggestion of two isoalleles for the electrophoretically intermediate enzyme.The research reported here was begun at the University of Hawaii and completed at the University of Texas and was supported (in part) by Public Health Service Research Grant No. GM 11609 to W. S. Stone and M. R. Wheeler and by Training Grant No. 2 T1-GM-337-06 and GM 00337-07 to R. P. Wagner et al., from the National Institutes of Health.     

Electron microscopy of the skin of the teleost,Hippoglossoides elassodon

Summary The normal skin of the pleuronectid fish, Hippoglossoides elassodon, is described by light and electron microscopy. The epidermis consists of 5 to 9 layers of cells, the majority of which are squamous cells and the minority mucous cells. The squamous cells are characterized by numerous desmosomes and associated cytoplasmic filaments. The mucous cells accumulate mucous droplets in vacuoles of Golgi origin and are observed apparently in the process of releasing their content at the free surface. The dermis consists of alternating lamellae composed of typical collagen fibers. Pigment cells are of three types: melanophores, iridophores (guanophores), and lipophores.This work was supported by Public Health Service Research Grant CA-08158 from the National Cancer Institute.