Extreme anti-oxidant protection against ionizing radiation in bdelloid rotifers.

Bdelloid rotifers, a class of freshwater invertebrates, are extraordinarily resistant to ionizing radiation (IR). Their radioresistance is not caused by reduced susceptibility to DNA double-strand breakage for IR makes double-strand breaks (DSBs) in bdelloids with essentially the same efficiency as in other species, regardless of radiosensitivity. Instead, we find that the bdelloid Adineta vaga is far more resistant to IR-induced protein carbonylation than is the much more radiosensitive nematode Caenorhabditis elegans. In both species, the dose-response for protein carbonylation parallels that for fecundity reduction, manifested as embryonic death. We conclude that the great radioresistance of bdelloid rotifers is a consequence of an unusually effective system of anti-oxidant protection of cellular constituents, including those required for DSB repair, allowing bdelloids to recover and continue reproducing after doses of IR causing hundreds of DSBs per nucleus. Bdelloid rotifers therefore offer an advantageous system for investigation of enhanced anti-oxidant protection and its consequences in animal systems.

Retroelements containing introns in diverse invertebrate taxa.

We report that two structurally similar transposable elements containing reverse transcriptase (RT), Penelope in Drosophila virilis and Athena in bdelloid rotifers, have proliferated as copies containing introns. The ability of Penelope-like elements (PLEs) to retain introns, their separate phylogenetic placement and their peculiar structural features make them a novel class of eukaryotic retroelements.

Phylogenomics of unusual histone H2A Variants in Bdelloid rotifers.

Rotifers of Class Bdelloidea are remarkable in having evolved for millions of years, apparently without males and meiosis. In addition, they are unusually resistant to desiccation and ionizing radiation and are able to repair hundreds of radiation-induced DNA double-strand breaks per genome with little effect on viability or reproduction. Because specific histone H2A variants are involved in DSB repair and certain meiotic processes in other eukaryotes, we investigated the histone H2A genes and proteins of two bdelloid species. Genomic libraries were built and probed to identify histone H2A genes in Adineta vaga and Philodina roseola, species representing two different bdelloid families. The expressed H2A proteins were visualized on SDS-PAGE gels and identified by tandem mass spectrometry. We find that neither the core histone H2A, present in nearly all other eukaryotes, nor the H2AX variant, a ubiquitous component of the eukaryotic DSB repair machinery, are present in bdelloid rotifers. Instead, they are replaced by unusual histone H2A variants of higher mass. In contrast, a species of rotifer belonging to the facultatively sexual, desiccation- and radiation-intolerant sister class of bdelloid rotifers, the monogononts, contains a canonical core histone H2A and appears to lack the bdelloid H2A variant genes. Applying phylogenetic tools, we demonstrate that the bdelloid-specific H2A variants arose as distinct lineages from canonical H2A separate from those leading to the H2AX and H2AZ variants. The replacement of core H2A and H2AX in bdelloid rotifers by previously uncharacterized H2A variants with extended carboxy-terminal tails is further evidence for evolutionary diversity within this class of histone H2A genes and may represent adaptation to unusual features specific to bdelloid rotifers.

Genomic signature of sexual reproduction in the bdelloid rotifer Macrotrachella quadricornifera.

Bdelloid rotifers, common freshwater invertebrates of ancient origin and worldwide distribution have long been thought to be entirely asexual, being the principal exception to the view that in eukaryotes the loss of sex leads to early extinction. That bdelloids are facultatively sexual is shown by a study of allele sharing within a group of closely related bdelloids of the species Macrotrachella quadricornifera, supporting the view that sexual reproduction is essential for long-term success in all eukaryotes.

Extreme resistance of bdelloid rotifers to ionizing radiation.

Rotifers of class Bdelloidea are common invertebrate animals with highly unusual characteristics, including apparently obligate asexuality, the ability to resume reproduction after desiccation at any life stage, and a paucity of transposable genetic elements of types not prone to horizontal transmission. We find that bdelloids are also extraordinarily resistant to ionizing radiation (IR). Reproduction of the bdelloids Adineta vaga and Philodina roseola is much more resistant to IR than that of Euchlanis dilatata, a rotifer belonging to the desiccation-intolerant and facultatively sexual class Monogononta, and all other animals for which we have found relevant data. By analogy with the desiccation- and radiation-resistant bacterium Deinococcus radiodurans, we suggest that the extraordinary radiation resistance of bdelloid rotifers is a consequence of their evolutionary adaptation to survive episodes of desiccation encountered in their characteristic habitats and that the damage incurred in such episodes includes DNA breakage that is repaired upon rehydration. Such breakage and repair may have maintained bdelloid chromosomes as colinear pairs and kept the load of transposable genetic elements low and may also have contributed to the success of bdelloid rotifers in avoiding the early extinction suffered by most asexuals.

Evidence for degenerate tetraploidy in bdelloid rotifers.

Rotifers of class Bdelloidea have evolved for millions of years apparently without sexual reproduction. We have sequenced 45- to 70-kb regions surrounding the four copies of the hsp82 gene of the bdelloid rotifer Philodina roseola, each of which is on a separate chromosome. The four regions comprise two colinear gene-rich pairs with gene content, order, and orientation conserved within each pair. Only a minority of genes are common to both pairs, also in the same orientation and order, but separated by gene-rich segments present in only one or the other pair. The pattern is consistent with degenerate tetraploidy with numerous segmental deletions, some in one pair of colinear chromosomes and some in the other. Divergence in 1,000-bp windows varies along an alignment of a colinear pair, from zero to as much as 20% in a pattern consistent with gene conversion associated with recombinational repair of DNA double-strand breaks. Although pairs of colinear chromosomes are a characteristic of sexually reproducing diploids and polyploids, a quite different explanation for their presence in bdelloids is suggested by the recent finding that bdelloid rotifers can recover and resume reproduction after suffering hundreds of radiation-induced DNA double-strand breaks per oocyte nucleus. Because bdelloid primary oocytes are in G(1) and therefore lack sister chromatids, we propose that bdelloid colinear chromosome pairs are maintained as templates for the repair of DNA double-strand breaks caused by the frequent desiccation and rehydration characteristic of bdelloid habitats.

Degenerate tetraploidy was established before bdelloid rotifer families diverged.

Rotifers of Class Bdelloidea are abundant freshwater invertebrates known for their remarkable ability to survive desiccation and their lack of males and meiosis. Sequencing and annotation of approximately 50-kb regions containing the four hsp82 heat shock genes of the bdelloid Philodina roseola, each located on a separate chromosome, have suggested that its genome is that of a degenerate tetraploid. In order to determine whether a similar structure exists in a bdelloid distantly related to P. roseola and if degenerate tetraploidy was established before the two species separated, we sequenced regions containing the hsp82 genes of a bdelloid belonging to a different family, Adineta vaga, and the histone gene clusters of P. roseola and A. vaga. Our findings are entirely consistent with degenerate tetraploidy and show that it was established before the two bdelloid families diverged and therefore probably before the bdelloid radiation.

A deep-branching clade of retrovirus-like retrotransposons in bdelloid rotifers.

Rotifers of class Bdelloidea, a group of aquatic invertebrates in which males and meiosis have never been documented, are also unusual in their lack of multicopy LINE-like and gypsy-like retrotransposons, groups inhabiting the genomes of nearly all other metazoans. Bdelloids do contain numerous DNA transposons, both intact and decayed, and domesticated Penelope-like retroelements Athena, concentrated at telomeric regions. Here we describe two LTR retrotransposons, each found at low copy number in a different bdelloid species, which define a clade different from previously known clades of LTR retrotransposons. Like bdelloid DNA transposons and Athena, these elements are found preferentially in telomeric regions. Unlike bdelloid DNA transposons, many of which are decayed, the newly described elements, named Vesta and Juno, inhabiting the genomes of Philodina roseola and Adineta vaga, respectively, appear to be intact and represent recent insertions, possibly from an exogenous source. We describe the retrovirus-like structure of the new elements, containing gag, pol, and env-like open reading frames, and discuss their possible origins, transmission, and behavior in bdelloid genomes.

Evidence for meiotic sex in bdelloid rotifers.

In their study of genetic exchange in the bdelloid rotifer Adineta vaga, Debortoli et al. [1] conclude that the patchwork pattern of allele sharing among three individuals in the genomic regions they examined is "…unlikely to arise in cases of PTH (Oenothera-like) meiosis since haplotypes are transferred as entire blocks…" and therefore that "Genetic exchange among bdelloid rotifers is more likely due to horizontal gene transfer than to meiotic sex." This assumes without justification that horizontal gene transfer (HGT) in bdelloids precludes the sexual transmission of entire haplotypes, for which we have reported evidence in the bdelloid Macrotrachela quadricornifera[2]. And it does not consider the contribution to such a patchwork pattern that would result from conversion and subsequent outcrossing, even in Oenothera-like systems.

Allele Sharing and Evidence for Sexuality in a Mitochondrial Clade of Bdelloid Rotifers.

Rotifers of Class Bdelloidea are common freshwater invertebrates of ancient origin whose apparent asexuality has posed a challenge to the view that sexual reproduction is essential for long-term evolutionary success in eukaryotes and to hypotheses for the advantage of sex. The possibility nevertheless exists that bdelloids reproduce sexually under unknown or inadequately investigated conditions. Although certain methods of population genetics offer definitive means for detecting infrequent or atypical sex, they have not previously been applied to bdelloid rotifers. We conducted such a test with bdelloids belonging to a mitochondrial clade of Macrotrachela quadricornifera. This revealed a striking pattern of allele sharing consistent with sexual reproduction and with meiosis of an atypical sort, in which segregation occurs without requiring homologous chromosome pairs.

The centenary of Janssens's chiasmatype theory.

The segregation and random assortment of characters observed by Mendel have their basis in the behavior of chromosomes in meiosis. But showing this actually to be the case requires a correct understanding of the meiotic behavior of chromosomes. This was achieved only gradually, over several decades, with much dispute and confusion along the way. One crucial step in the understanding of meiosis was provided in 1909 by Frans Alfons Janssens who published in La Cellule an article entitled "La théorie de la Chiasmatypie. Nouvelle interprétation des cinèses de maturation," which contains the first description of the chiasma structure. He observed that, of the four chromatids present at the connection sites (chiasmata sites) at diplotene or anaphase of the first meiotic division, two crossed each other and two did not. He therefore postulated that the maternal and paternal chromatids that crossed penetrated the other until they broke and rejoined in maternal and paternal segments new ways; the other two chromatids remained free and thus intact. This allowed him also to propose that the chromatids distributed in the four nuclei issued from the second meiotic division had various combinations of maternal and paternal segments of each chromosome. And conversely, permitted the appreciation that the laws of Mendelian segregation required breakage and joining (crossing over) between homologous non-sister chromatids. Although Janssens's article found a broad appreciative audience and had a large influence on the chromosomal theory at that time, his theory was resisted by both geneticists and cytologists for several decades. This Perspectives aims to highlight the novelty of Janssens's chiasmatype theory by examining the historical background and our actual understanding of meiotic recombination.

Massive horizontal gene transfer in bdelloid rotifers.

Horizontal gene transfer in metazoans has been documented in only a few species and is usually associated with endosymbiosis or parasitism. By contrast, in bdelloid rotifers we found many genes that appear to have originated in bacteria, fungi, and plants, concentrated in telomeric regions along with diverse mobile genetic elements. Bdelloid proximal gene-rich regions, however, appeared to lack foreign genes, thereby resembling those of model metazoan organisms. Some of the foreign genes were defective, whereas others were intact and transcribed; some of the latter contained functional spliceosomal introns. One such gene, apparently of bacterial origin, was overexpressed in Escherichia coli and yielded an active enzyme. The capture and functional assimilation of exogenous genes may represent an important force in bdelloid evolution.

Deleterious transposable elements and the extinction of asexuals.

The genomes of virtually all sexually reproducing species contain transposable elements. Although active elements generally transpose more rapidly than they are inactivated by mutation or excision, their number can be kept in check by purifying selection if its effectiveness becomes disproportionately greater as their copy number increases. In sexually reproducing species, such synergistic selection can result from ectopic crossing-over or from homologous recombination under negative epistasis. In addition, there may be controls on transposon activity that are associated with meiosis. Because a sexual lineage that abandons sex must lack such mechanisms, it may be driven to extinction by the unchecked proliferation of deleterious transposons inherited from its sexual progenitor. An important component of the evolutionary advantage of sex over asex may therefore lie in the ability of sex, despite facilitating the spread of deleterious elements within interbreeding populations, also to restrain their intragenomic proliferation.

Diverse DNA transposons in rotifers of the class Bdelloidea.

We surveyed the diversity, structural organization, and patterns of evolution of DNA transposons in rotifers of the class Bdelloidea, a group of basal triploblast animals that appears to have evolved for millions of years without sexual reproduction. Representatives of five superfamilies were identified: ITm (IS630/Tc/mariner), hAT, piggyBac, helitron, and foldback. Except for mariners, no fully intact copies were found. Mariners, both intact and decayed, are present in high copy number, and those described here may be grouped in several closely related lineages. Comparisons across lineages show strong evidence of purifying selection, whereas there is little or no evidence of such selection within lineages. This pattern could have resulted from repeated horizontal transfers from an exogenous source, followed by limited intragenomic proliferation, or, less plausibly, from within-host formation of new lineages under host- or element-based selection for function, in either case followed by eventual inactivation and decay. Unexpectedly, the flanking sequences surrounding the majority of mariners are very similar, indicating either insertion specificity or proliferation as part of larger DNA segments. Members of all superfamilies are present near chromosome ends, associated with the apparently domesticated retroelement Athena, in large clusters composed of diverse DNA transposons, often inserted into each other, whereas the examined gene-rich regions are nearly transposon-free.