Karl Esser

The mitochondrial plasmid of Podospora anserina: A mobile intron of a mitochondrial gene

SummaryIn the ascomycete Podospora anserina strain ageing (senescence) is caused by a mitochondrial plasmid. In juvenile mycelia it is an integral part of the mtDNA and becomes liberated during ageing. The nucleotide sequence of this plasmid and of its flanking regions was determined. It consists of 2,539 by and contains an un identified reading frame (URF) originating in the adjacent mtDNA upstream of excision point 1. Within the URF a putative 48 by autonomously replicating sequence (ars) was identified. At both excision sites of the plasmid there are two short nonidentical interrupted palindromes and a few base pairs apart from these palindromes, both upstream and downstream, two short inverted repeats are localised. The experimental data make it evident that the mt plasmid is an intron of the cytochrome c oxidase gene (subunit I) which may be excised at the DNA level and thus become the mobile infective agent causing senescence. The concept of this mobile intron and current hypotheses concerning the relationship between introns and transposons are stressed.

Linear plasmids among eukaryotes: fundamentals and application.

Introduction. Plasmids of eukaryotes have long been a neglected subject. Because they were initially found only in Saccharomyces cerevisiae (Sinclair et al. 1967), they were considered to be genetic elements of limited distribution with a cryptic function. Subsequently plasmids were detected in a plant, Zea mays (Pring et al. 1977), and in a filamentous fungus, Podospora anserina (Stahl et al. 1978). The fungal plasmid turned out to be a circular molecule, as expected in accordance with the generally accepted plasmid configuration (ccc = covalently closed circular). The corn plasmid, however, was the first linear plasmid to be described, though it was to prove antecedent to many others. Circular plasmids of eukaryotes have been reviewed extensively (see Esser ct al. 1986). Linear plasmids, many of which have been detected and analysed more recently, have yet to be reviewed in respect of their common properties and their potential application for research.

Evidence for plasmid like DNA in a filamentous fungus, the ascomycete Podospora anserina.

SummaryThe existece of plasmid like DNA was demonstrated in senescent mycelia of Podospora anserina (strain s) by biophysical and electronmicroscopic methods. According to their contour length of about 1.4 and 2.7 μm respectively the molecular weight for the monomer is in the range of 3·106.

Plasmid-like DNA is part of mitochondrial DNA in Podospora anserina

SummaryAs previously reported, a ccc DNA with a contour length of 0.75 µm and molecular weight of 2.4 kb (termed plasmid-like, p1DNA) is the causative agent of senescence in the fungus Podospora anserina. Its postulated location in mtDNA was proved correct by the following experiments:1.Restriction analysis of mtDNA resulted in different molecular weights for both, juvenile (95 kb) and senescent (30 kb) mtDNA. The construction of a detailed restriction map made evident the fact that senescent mtDNA comprises only a part of its juvenile counterpart.2.Hybridization experiments (Southern blots) between 3H-labelled plDNA and mtDNA cleaved by restriction juvenile mtDNA are homologous to plDNA.3.Fine mapping experiments (construction of restriction maps and heteroduplex experiments) between plDNA integrated into a bacterial vector and its postulated equivalent, derived from juvenile mtDNA and also integrated into a bacterial vector, allowed a precise determination of the site of plDNA insertion within the juvenile mtDNA. All of these data fit into a previously published model in which, during aging, plDNA is excised from mtDNA and becomes autonomous for replication and function.

The onset of senescence is affected by DNA rearrangements of a discontinuous mitochondrial gene in Podospora anserina.

SummaryMapping and transcription studies have revealed that in Podospora anserina the causative agent of senescence, a mitochondrial plasmid (p1DNA), is identical with intronl of the discontinuous gene for cytochrome-c-oxidase subunit 1 (COI), which is 2 kpb from the discontinuous gene for cytochrome b (Cytb). A mitochondrial mutant (ex1) devoid of the COI, but not of the Cytb gene provides longevity. A molecular model for the onset of senescence is presented.

Chromosomal and extrachromosomal control of senescence in the ascomycete Podospora anserina.

SummaryIn Podospora anserina senescence leading to cellular death occurs regularly after prolonged vegetative propagation. However, the life span of this ascomycete may be extended by various means:I. Mutations in at least 8 morphogenetic genes belonging to 4 linkage groups postpone drastically or even prevent in certain pairwise combinations (e.g. i viv) the onset of senescence. 2. Inhibitors of mt DNA and of mitochondrial protein synthesis show a life prolonging effect when added in low concentrations to the growth medium. 3. A similar effect was found when mycelia were fed exclusively on non repressive carbon sources.Whereas the anti-aging effect of specific mutated genes is rather permanent, the life prolonging action of the inhibitors and carbon sources is restricted and temporary. These substances have no long lasting effect, since after their removal from the medium aging proceeds.Physiological experiments have further shown the existence of three phases in the life span of Podospora anserina. During the juvenile phase aging is prevented by all of these compounds; during the presenescent phase aging is prevented by inhibitors of mt DNA only, and during the senescent phase aging is irreversible.Senescence may be induced in juvenile protoplasts by DNA extracted from senescent mycelia. This, together with the well known fact that senescence is extrachromosomically inherited, points to extrachromosomal DNA as the causative agent of senescence. This kind of DNA may be connected with or perhaps located in the mitochondria.Collectively, the data are consistent in showing that the syndrome of senescence in Podospora anserina is controlled by a chromosomal-extrachromosomal is controlled by a chromosomal-extrachromosomal interaction. In this system, extrachromosomal DNA, perhaps a mt DNA, is identical with the infectious principle initiating the decay of the cell, and nuclear genes supervise its expression.

Heterogenic Incompatibility in Plants and Animals

Publisher Summary This chapter that in many cases the occurrence of heterogenic incompatibility has been unequivocally demonstrated by comprehensive genetic data. However, in a number of examples, the phenomenon is inferred from circumstantial evidence only. The main reason is that, effects properly attributed to heterogenic incompatibility either has been considered as having other causes or else have been merely mentioned as inexplicable secondary effects. However, in spite of these difficulties there are some general concepts, which need to be amplified in a more general manner: Distribution, nature of genetic determinants, and efficiency in sexual and vegetative phases, general action, and demarcation versus diffusible inhibitory substance, biochemical basis, and relations with histoincompatibility, consequences for taxonomy, and practical consequences. The chapter intends to acquaint biologists not only with the existence and versatile genetic basis of heterogenic incompatibility, but also with its general importance as a basic biological phenomenon.

Characterization and cloning of plasmid like DNA of the ascomycete Podospora anserina.

SummaryThe previously reported existence of plasmid-like (pl) DNA in senescent mycelia of Podospora anserina was confirmed using new methodology. Detailed analysis of bulk DNA has further shown a possible relationship between pl DNA and mt DNA.According to biophysical and electron microscopic experiments the pl DNA was found to consist of oligomeres having a basic unit with a contour length of 0.75 μm corresponding to 2.4 kb. To overcome the handicap that pl DNA is only produced in rather small amounts in the aging mycelia, this DNA was cloned in E. coli after insertion into a bacterial plasmid vector, pBR 322. It was possible to isolate a stable hybrid plasmid consisting of the vector and only one integrated monomere of pl DNA. The composition of this hybrid plasmid was confirmed by restriction endonuclease analysis and heteroduplex formation. A restriction map of the pl DNA is presented and its insertion site onto pBR 322 indicated.

The Phenoloxidases of the Ascomycete Podospora anserina

SummaryIn order to learn the internal conditions for the production of the various phenoloxidases produced by the Ascomycete Podospora anserina the wild strain has been grown under controlled conditions in a fermenter for a period of 34 days. Samples were withdrawn at regular intervals and assayed for mycelial yield and intra- and extracellular phenoloxidase production.Maximal yield was obtained at the following age of the culture: Mycelial production 9 d, tyrosinase 4 d, the high molecular weight laccase I between 9 and 19 d. The low molecular weight laccases II and III, initially present in medium concentrations, dropped to an early minimum after 4 days, followed by an increase with a maximum in the late autolytic phase.The changes in the phenoloxidase spectrum and the antiparallel production curve for the high molecular weight against the low molecular weight laccases are discussed in relation to the earlier observed genetical and physiological control of phenoloxidase synthesis and in relation to the possibility of laccase I being composed of active subunits of low molecular weight laccases.

Genetics of fruit body production in higher basidiomycetes

Summary1.In Polyporus ciliatus (Polyporaceae) dikaryotic fruiting is controlled by the tetrapolar mechanism of homogenic incompatibility.2.The occurrence of subunits of the mating type factors A and B, known in other Holobasidiomycetidae, could not be proved. If subunits are present, their distance must be less than 0.2 map units.3.Monokaryotic fruiting occurs in a strictly haploid phase. Neither karyogamy nor meiosis is required in the fruit bodies for the development of basidia having two spores.4.Genetic analysis has revealed that for the initiation of monokaryotic fruting a single gene (fi+/fi) is responsible. Additional genes control the shape of the fruit bodies (fb+/fb) and its fertility (mod+/mod).5.Monokaryotic fruiting is closely correlated with dikaryotic fruiting in two ways. On the one hand the B factors, part of the controlling system for dikaryotic fruiting, block monokaryotic fruiting when both partners are heterogenic for the B factor. On the other hand, despite the fact that monokaryotic fruiting is suppressed in a dikaryon, the fi gene controls the formation of dikaryotic fruit bodies. The allele fi+ enhances fruit body production whereas the allele fi may inhibit fruit body production completely.6.The action and interaction of the incompatibility factors, the genes responsible for monokaryotic fruiting and the genes causing heterogenic incompatibility is discussed with respect to evolution and concerted breeding of mushrooms.