Eighty million years without sex: the role of genome structure in bdelloid rotifers

Bdelloid (pronounced with a silent 'b') rotifers are abundant, harmless, microscopic animals, most of which live in temporary freshwater pools or damp habitats. They have two remarkable characteristics: 1) They have survived without sex for perhaps 80 million years. No males have ever been found, and molecular evidence is consistent with a long evolutionary history of asexual reproduction. Although other asexual organisms are known, they have arisen recently in evolutionary history and are usually thought to be evolutionary dead ends which will rapidly become extinct. As a result, bdelloids were called 'an evolutionary scandal' by the late Prof John Maynard Smith, a prominent evolutionist. 2) They can survive almost complete water loss. Living creatures are mostly water and need to retain this water to stay alive. However, bdelloids are one group of organisms which can dry but not die; they are 'desiccation tolerant'. As their surroundings dry out, bdelloids enter a state of suspended animation in which life processes become undetectable, but when rehydrated, they come back to life as if nothing had happened. Even more remarkably, they can remain in the dry state for many years without apparent ill effects and, while dry, are highly resistant to extremes of temperature and pressure. In this proposal, potential links between these characteristics will be explored relating to how bdelloids use gene copies. In diploid organisms, all 'single copy' genes are in fact present in two copies, each on a homologous chromosome. The available evidence suggests that bdelloids resemble diploids, in that they have two copies of most genes analysed. However, for some genes, perhaps 15%, they have four copies. It has therefore been suggested that bdelloids are degenerate tetraploids, descending from an ancestor with four copies of all genes, but that many of these have been lost during evolution. Sexual organisms undergo a process of genetic exchange during the generation of germ cells (eggs and sperm) which leads to homologous gene copies ('alleles') in a population being very similar in sequence. Because bdelloids reproduce asexually, this does not happen and therefore corresponding gene copies ('former alleles') will accumulate changes ('mutations') over time. So former alleles will have substantially different DNA sequences: this has been called the Meselson effect after the scientist who first discovered it. We have shown recently in bdelloids that such sequence divergence between one pair of gene copies can also result in functional differences, and that this can potentially be exploited by bdelloids to increase their ability to survive desiccation. Evolution of gene function in this way can't happen in sexual organisms, and it is therefore something that only asexuals can exploit. There is another proposal for how bdelloids use gene copies, however. Meselson and colleagues have suggested that they are used for repairing damaged genes. One likely outcome of drying out is that DNA becomes broken, and Meselson proposes that bdelloids employ intact gene copies to replace those which are damaged. This process should work in the opposite direction to the sequence divergence noted above, and thereby eliminate any functional divergence of related gene copies, and with it any evolutionary advantage it confers. This proposal will therefore test these two opposing ideas and attempt to discover whether bdelloid rotifers use gene copies to diversify the function of their genes, or to maintain gene function.

Bdelloid rotifers have survived for tens of millions of years without sexual reproduction: males have never been observed, and genetic evidence is consistent with the absence of meiotic recombination. One consequence of this is the Meselson effect, i.e. sequence divergence of pairs of genes which were formerly alleles in the sexual ancestor of bdelloids. We demonstrated recently that sequence divergence was also accompanied by structural and functional divergence in a pair of bdelloid genes encoding LEA proteins, which are associated with desiccation tolerance in plants, animals and micro-organisms. The two LEA proteins have either a protein protection, or a membrane protection, function during desiccation; the development of such complementary activities is likely to be adaptive in the drought-prone habitats frequented by bdelloid rotifers. We therefore propose to investigate whether this phenomenon - the functional divergence of former alleles - is more widespread, and in particular whether it occurs in other genes connected with desiccation tolerance. A library of ~70 dehydration-induced cDNAs has been identified and suitable candidates, together with control genes not thought to be involved in desiccation tolerance, will be analysed at the gene and protein level for divergent function. Analyses will be carried out in the species Adineta ricciae, and comparisons of some gene pairs made in other bdelloid species with varying degrees of desiccation tolerance. A contrasting hypothesis has been proposed suggesting that bdelloids use gene copies as a template to repair DNA damage likely to result from desiccation. We will therefore also test this hypothesis by first assessing whether significant double strand breakage occurs during desiccation, and then looking for evidence of break repair by gene conversion. We will also attempt to show evidence of novel gene conversion events subsequent to desiccation or irradiation.