The sensitivity of the commonly used progressive multiple sequence alignment method has been greatly improved for the alignment of divergent protein sequences. Firstly, individual weights are assigned to each sequence in a partial alignment in order to down-weight near-duplicate sequences and up-weight the most divergent ones. Secondly, amino acid substitution matrices are varied at different alignment stages according to the divergence of the sequences to be aligned. Thirdly, residue-specific gap penalties and locally reduced gap penalties in hydrophilic regions encourage new gaps in potential loop regions rather than regular secondary structure. Fourthly, positions in early alignments where gaps have been opened receive locally reduced gap penalties to encourage the opening up of new gaps at these positions. These modifications are incorporated into a new program, CLUSTAL W which is freely available.

/pdf/clustal-w-improving-the-sensitivity-of-progressive-multiple-2na2s9sq2h.pdf

Clustal w: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice

Extrachromosomal rDNA circles--a cause of aging in yeast.

https://www.rand.org/content/dam/rand/www/external/labor/aging/rsi/rsi_papers/2009/benz5.pdf

Trends in oxidative aging theories

In nearly all eukaryotes, at least some individuals inherit mitochondrial and chloroplast genes from only one parent. There is no single mechanism of uniparental inheritance: organelle gene inheritance is blocked by a variety of mechanisms and at different stages of reproduction in different species. Frequent changes in the pattern of organelle gene inheritance during evolution suggest that it is subject to varying selective pressures. Organelle genes often fail to recombine even when inherited biparentally; consequently, their inheritance is asexual. Sexual reproduction is apparently less important for genes in organelles than for nuclear genes, probably because there are fewer of them. As a result organelle sex can be lost because of selection for special reproductive features such as oogamy or because uniparental inheritance reduces the spread of cytoplasmic parasites and selfish organelle DNA.

https://www.pnas.org/content/pnas/92/25/11331.full.pdf

Uniparental inheritance of mitochondrial and chloroplast genes: mechanisms and evolution.

Mitochondrial mutations may increase oxidative stress: implications for carcinogenesis and aging?

Senescence in podospora anserina: Amplification of a mitochondrial DNA sequence

The non-nuclear DNA of a number of independent senescent cultures of Podospora anserina was extracted and studied. In all cases, a specific repetitive DNA (SEN-DNA) arranged in multimeric sets of circular molecules, was identified. Depending on the senescent culture, the SEN-DNA was found either in a band of about same density as the mitochondrial DNA from young mycelia (1.694 g/cm3) or in a band of higher density (1.699 g/cm3). Electron microscopy, restriction enzyme analysis and Southern hybridization experiments allowed us to establish that: (1) SEN-DNAs obtained from independent senescent cultures, both from the same strain and from different strains, can differ in the size of their monomer unit (from 2.5 to 6.3 kb). (2) All SEN-DNAs hybridize with mitochondrial DNA of a young culture and not with nuclear DNA. (3) These SEN-DNAs belong to two classes which hybridize with two non-overlapping regions of the mitochondrial chromosome.

Mitochondrial DNA amplification in senescent cultures of Podospora anserina : Variability between the retained, amplified sequences

In Podospora anserina, a strict aerobic fungus, senescence1 (arrest of vegetative growth of the mycelium) is correlated with severe modifications of the mitochondrial DNA similar to those observed in the rho− mutants of yeast2. Senescent cultures yield a specific DNA (SEN-DNA) consisting of small circular molecules arranged in multimeric series of sizes3,4 and which results from the amplification of either one of two non-overlapping segments of the mitochondrial DNA5,6. One region 0.89 µm long [2.6 kilobases (kb)] is most frequently amplified, in different strains of P. anserina and also in subcultures of a strain reaching senescence independently6. In ‘young’ cultures of the mating type-minus wild-type strain s (s mt−), the 2.6 kb region exists both as an integral part of the mitochondrial chromosome and as rare free sequences6. The results presented here show that (1) the free form is absent from two strains, s mt+ and s mt− capr1, which both display increased longevity and which differ respectively from the s mt− strain by a nuclear allele at the mt locus and by a mitochondrial mutation conferring resistance to chloramphenicol; and (2) in the mutant mex1, which develops beyond the edge of arrest of a senescent mycelium, that is, escapes senescence, the mitochondrial DNA precisely lacks the 2.6kb region. The results suggest that excision and/or amplification of the 2.6 kb sequence is the cause of senescence.

A sequence of mitochondrial DNA is associated with the onset of senescence in a fungus.

Several lines of evidence have implicated reactive oxygen species (ROS) in the pathogenesis of various degenerative diseases and in organismal ageing Furthermore, it has been shown recently that the alternative pathway respiration present in plants lowers ROS mitochondrial production An alternative oxidase (AOXp) also occurs in the filamentous fungus Podospora anserina We show here that overexpression of this oxidase does not decrease ROS production and has no effect on longevity, mitochondrial stability or ageing in this fungus In the same way, inactivation of the gene has no effect on these parameters In contrast, overexpression of the alternative oxidase in the long-lived cox5::BLE mutant, deficient in cytochrome c oxidase, considerably increases ROS production of the mutant It rescues slow growth rate and female sterility, indicating an improved energy level This overexpression also restores senescence and mitochondrial DNA instability, demonstrating that these parameters are controlled by the energy level and not by the expression level of the alternative oxidase We also suggest that expression of this oxidase in organisms naturally devoid of it could rescue respiratory defects resulting from cytochrome pathway dysfunctions

Overexpression of the alternative oxidase restores senescence and fertility in a long-lived respiration-deficient mutant of Podospora anserina.

In the filamentous fungus Podospora anserina, the association of two nuclear genes inevitably leads to a "premature death" phenotype consisting of an early end of vegetative growth a few days after ascospore germination. Mycelia showing this phenotype contain a mitochondrial chromosome that always bears the same deletion. One of the break points is exactly at the 5' splice site of a particular mitochondrial intron, suggesting that the deletion event could result from molecular mechanisms also involved in intron mobility. One of the nuclear genes involved in triggering this site-specific event belongs to the mating-type minus haplotype; the other is a mutant allele of a gene encoding a cytosolic ribosomal protein.

Odile Begel

Papers

Senescence in podospora anserina: Amplification of a mitochondrial DNA sequence

Mitochondrial DNA amplification in senescent cultures of Podospora anserina : Variability between the retained, amplified sequences

A sequence of mitochondrial DNA is associated with the onset of senescence in a fungus.

Overexpression of the alternative oxidase restores senescence and fertility in a long-lived respiration-deficient mutant of Podospora anserina.

A site-specific deletion in mitochondrial DNA of Podospora is under the control of nuclear genes.