Integration of mitochondrial gene sequences within the nuclear genome during senescence in a fungus

Cellular senescence in the ascomycete fungus Podospora anserina is associated with the appearance of an altered mitochondrial genome. Discrete mitochondrial DNA sequences are excised and amplified and isolated as multimerically arranged, head-to-tail repetitions. We have referred to the most frequently observed excision/amplification product as alpha-event senDNA. It is a 2.6-kilobase pair (kbp) monomeric unit (see refs 1, 3, 7) and is often found in senescent mitochondria in conjunction with other excision products. At the final stage of senescence these plasmids constitute virtually all of the DNA present in senescent mitochondria; they have replicated to high copy number at the expense of the young native genome. Because P. anserina is characterized by race-specific timing of senescence (that is, a programme of senescence), we have begun to contrast rapidly and slowly senescing races in terms of senDNA. Here we present evidence that young mitochondria of the rapidly senescing race, A+, possess an extremely high copy number of alpha-event senDNA plasmid in contrast to the more slowly senescing races s+ or s-. Moreover, we observe that during senescence the alpha-event senDNA and the beta-event senDNA (a 9.8-kbp monomer) are transposed to the nucleus and integrated into nuclear DNA. These plasmids contain the coding information for subunits I and III (respectively) of the mitochondrial cytochrome c oxidase. This constitutes the first clear evidence for the active mobilization of genetic elements from the mitochondrion to the nucleus.     

Bioinformatic prediction of selenoprotein genes in the dolphin genome

Selenium (Se), an essential trace element in vivo, is present mainly as selenocystein (Sec) in various selenoproteins. The Sec residue is translated from an in-frame TGA codon, which traditionally functions as a stop codon. Prediction of selenoprotein genes is difficult due to the lack of an effective method for distinguishing the dual function of the TGA codon in the open reading frame of a selenoprotein gene. In this article a eukaryotic bioinformatic prediction system that we have developed was used to predict selenoprotein genes from the genome of the common bottlenose dolphin, Tursiops truncatus. Sixteen selenoprotein genes were predicted, including selenoprotein P and glutathione peroxidase. In particular, a type II iodothyronine deiodinase was found to have two Sec residues, while the type I iodothyronine deiodinase gene has two alternative splice forms. These results provide important information for the investigation of the relationship between a variety of selenoproteins and the evolution of the marine-living dolphin.     

Selfish genes, the phenotype paradigm and genome evolution

Natural selection operating within genomes will inevitably result in the appearance of DNAs with no phenotypic expression whose only 'function' is survival within genomes. Prokaryotic transposable elements and eukaryotic middle-repetitive sequences can be seen as such DNA's and thus no phenotypic or evolutionary function need be assigned to them.     

Proof and evolutionary analysis of ancient genome duplication in the yeast Saccharomyces cerevisiae

Whole-genome duplication followed by massive gene loss and specialization has long been postulated as a powerful mechanism of evolutionary innovation. Recently, it has become possible to test this notion by searching complete genome sequence for signs of ancient duplication. Here, we show that the yeast Saccharomyces cerevisiae arose from ancient whole-genome duplication, by sequencing and analysing Kluyveromyces waltii, a related yeast species that diverged before the duplication. The two genomes are related by a 1:2 mapping, with each region of K. waltii corresponding to two regions of S. cerevisiae, as expected for whole-genome duplication. This resolves the long-standing controversy on the ancestry of the yeast genome, and makes it possible to study the fate of duplicated genes directly. Strikingly, 95% of cases of accelerated evolution involve only one member of a gene pair, providing strong support for a specific model of evolution, and allowing us to distinguish ancestral and derived functions.     

Multiplex amplification of the mammoth mitochondrial genome and the evolution of Elephantidae

In studying the genomes of extinct species, two principal limitations are typically the small quantities of endogenous ancient DNA and its degraded condition, even though products of up to 1,600 base pairs (bp) have been amplified in rare cases. Using small overlapping polymerase chain reaction products, longer stretches of sequences or even whole mitochondrial genomes can be reconstructed, but this approach is limited by the number of amplifications that can be performed from rare samples. Thus, even from well-studied Pleistocene species such as mammoths, ground sloths and cave bears, no DNA sequences of more than about 1,000 bp have been reconstructed. Here we report the complete mitochondrial genome sequence of the Pleistocene woolly mammoth Mammuthus primigenius. We used about 200 mg of bone and a new approach that allows the simultaneous retrieval of multiple sequences from small amounts of degraded DNA. Our phylogenetic analyses show that the mammoth was more closely related to the Asian than to the African elephant. However, the divergence of mammoth, African and Asian elephants occurred over a short time, corresponding to only about 7% of the total length of the phylogenetic tree for the three evolutionary lineages.     

Large-scale analysis of the yeast genome by transposon tagging and gene disruption

Economical methods by which gene function may be analysed on a genomic scale are relatively scarce. To fill this need, we have developed a transposon-tagging strategy for the genome-wide analysis of disruption phenotypes, gene expression and protein localization, and have applied this method to the large-scale analysis of gene function in the budding yeast Saccharomyces cerevisiae. Here we present the largest collection of defined yeast mutants ever generated within a single genetic background--a collection of over 11,000 strains, each carrying a transposon inserted within a region of the genome expressed during vegetative growth and/or sporulation. These insertions affect nearly 2,000 annotated genes, representing about one-third of the 6,200 predicted genes in the yeast genome. We have used this collection to determine disruption phenotypes for nearly 8,000 strains using 20 different growth conditions; the resulting data sets were clustered to identify groups of functionally related genes. We have also identified over 300 previously non-annotated open reading frames and analysed by indirect immunofluorescence over 1,300 transposon-tagged proteins. In total, our study encompasses over 260,000 data points, constituting the largest functional analysis of the yeast genome ever undertaken.     

High mutation rate and predominance of insertions in the Caenorhabditis elegans nuclear genome

Mutations have pivotal functions in the onset of genetic diseases and are the fundamental substrate for evolution. However, present estimates of the spontaneous mutation rate and spectrum are derived from indirect and biased measurements. For instance, mutation rate estimates for Caenorhabditis elegans are extrapolated from observations on a few genetic loci with visible phenotypes and vary over an order of magnitude. Alternative approaches in mammals, relying on phylogenetic comparisons of pseudogene loci and fourfold degenerate codon positions, suffer from uncertainties in the actual number of generations separating the compared species and the inability to exclude biases associated with natural selection. Here we provide a direct and unbiased estimate of the nuclear mutation rate and its molecular spectrum with a set of C. elegans mutation-accumulation lines that reveal a mutation rate about tenfold higher than previous indirect estimates and an excess of insertions over deletions. Because deletions dominate patterns of C. elegans pseudogene variation, our observations indicate that natural selection might be significant in promoting small genome size, and challenge the prevalent assumption that pseudogene divergence accurately reflects the spontaneous mutation spectrum.     

Widespread horizontal transfer of mitochondrial genes in flowering plants

Horizontal gene transfer--the exchange of genes across mating barriers--is recognized as a major force in bacterial evolution. However, in eukaryotes it is prevalent only in certain phagotrophic protists and limited largely to the ancient acquisition of bacterial genes. Although the human genome was initially reported to contain over 100 genes acquired during vertebrate evolution from bacteria, this claim was immediately and repeatedly rebutted. Moreover, horizontal transfer is unknown within the evolution of animals, plants and fungi except in the special context of mobile genetic elements. Here we show, however, that standard mitochondrial genes, encoding ribosomal and respiratory proteins, are subject to evolutionarily frequent horizontal transfer between distantly related flowering plants. These transfers have created a variety of genomic outcomes, including gene duplication, recapture of genes lost through transfer to the nucleus, and chimaeric, half-monocot, half-dicot genes. These results imply the existence of mechanisms for the delivery of DNA between unrelated plants, indicate that horizontal transfer is also a force in plant nuclear genomes, and are discussed in the contexts of plant molecular phylogeny and genetically modified plants.