Eugene Gladyshev

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.

Genomic evidence for ameiotic evolution in the bdelloid rotifer Adineta vaga

Loss of sexual reproduction is considered an evolutionary dead end for metazoans, but bdelloid rotifers challenge this view as they appear to have persisted asexually for millions of years. Neither male sex organs nor meiosis have ever been observed in these microscopic animals: oocytes are formed through mitotic divisions, with no reduction of chromosome number and no indication of chromosome pairing. However, current evidence does not exclude that they may engage in sex on rare, cryptic occasions. Here we report the genome of a bdelloid rotifer, Adineta vaga (Davis, 1873), and show that its structure is incompatible with conventional meiosis. At gene scale, the genome of A. vaga is tetraploid and comprises both anciently duplicated segments and less divergent allelic regions. However, in contrast to sexual species, the allelic regions are rearranged and sometimes even found on the same chromosome. Such structure does not allow meiotic pairing; instead, we find abundant evidence of gene conversion, which may limit the accumulation of deleterious mutations in the absence of meiosis. Gene families involved in resistance to oxidation, carbohydrate metabolism and defence against transposons are significantly expanded, which may explain why transposable elements cover only 3% of the assembled sequence. Furthermore, 8% of the genes are likely to be of non-metazoan origin and were probably acquired horizontally. This apparent convergence between bdelloids and prokaryotes sheds new light on the evolutionary significance of sex.

Telomere-associated endonuclease-deficient Penelope-like retroelements in diverse eukaryotes

The evolutionary origin of telomerases, enzymes that maintain the ends of linear chromosomes in most eukaryotes, is a subject of debate. Penelope-like elements (PLEs) are a recently described class of eukaryotic retroelements characterized by a GIY-YIG endonuclease domain and by a reverse transcriptase domain with similarity to telomerases and group II introns. Here we report that a subset of PLEs found in bdelloid rotifers, basidiomycete fungi, stramenopiles, and plants, representing four different eukaryotic kingdoms, lack the endonuclease domain and are located at telomeres. The 5′ truncated ends of these elements are telomere-oriented and typically capped by species-specific telomeric repeats. Most of them also carry several shorter stretches of telomeric repeats at or near their 3′ ends, which could facilitate utilization of the telomeric G-rich 3′ overhangs to prime reverse transcription. Many of these telomere-associated PLEs occupy a basal phylogenetic position close to the point of divergence from the telomerase-PLE common ancestor and may descend from the missing link between early eukaryotic retroelements and present-day telomerases.

A widespread class of reverse transcriptase-related cellular genes

Reverse transcriptases (RTs) polymerize DNA on RNA templates. They fall into several structurally related but distinct classes and form an assemblage of RT-like enzymes that, in addition to RTs, also includes certain viral RNA-dependent RNA polymerases (RdRP) synthesizing RNA on RNA templates. It is generally believed that most RT-like enzymes originate from retrotransposons or viruses and have no specific function in the host cell, with telomerases being the only notable exception. Here we report on the discovery and properties of a unique class of RT-related cellular genes collectively named rvt. We present evidence that rvts are not components of retrotransposons or viruses, but single-copy genes with a characteristic domain structure that may contain introns in evolutionarily conserved positions, occur in syntenic regions, and evolve under purifying selection. These genes can be found in all major taxonomic groups including protists, fungi, animals, plants, and even bacteria, although they exhibit patchy phylogenetic distribution in each kingdom. We also show that the RVT protein purified from one of its natural hosts, Neurospora crassa, exists in a multimeric form and has the ability to polymerize NTPs as well as dNTPs in vitro, with a strong preference for NTPs, using Mn2+ as a cofactor. The existence of a previously unknown class of single-copy RT-related genes calls for reevaluation of the current views on evolution and functional roles of RNA-dependent polymerases in living cells.

Genome Structure of Bdelloid Rotifers: Shaped by Asexuality or Desiccation?

Bdelloid rotifers are microscopic invertebrate animals best known for their ancient asexuality and the ability to survive desiccation at any life stage. Both factors are expected to have a profound influence on their genome structure. Recent molecular studies demonstrated that, although the gene-rich regions of bdelloid genomes are organized as colinear pairs of closely related sequences and depleted in repetitive DNA, subtelomeric regions harbor diverse transposable elements and horizontally acquired genes of foreign origin. Although asexuality is expected to result in depletion of deleterious transposons, only desiccation appears to have the power to produce all the uncovered genomic peculiarities. Repair of desiccation-induced DNA damage would require the presence of a homologous template, maintaining colinear pairs in gene-rich regions and selecting against insertion of repetitive DNA that might cause chromosomal rearrangements. Desiccation may also induce a transient state of competence in recovering animals, allowing them to acquire environmental DNA. Even if bdelloids engage in rare or obscure forms of sexual reproduction, all these features could still be present. The relative contribution of asexuality and desiccation to genome organization may be clarified by analyzing whole-genome sequences and comparing foreign gene and transposon content in species which lost the ability to survive desiccation.

Direct recognition of homology between double helices of DNA in Neurospora crassa

Chromosomal regions of identical or nearly identical DNA sequence can preferentially associate with one another in the apparent absence of DNA breakage. Molecular mechanism(s) underlying such homology-dependent pairing phenomena remain(s) unknown. Using Neurospora crassa repeat-induced point mutation (RIP) as a model system, we show that a pair of DNA segments can be recognized as homologous if they share triplets of base pairs arrayed with the matching periodicity of 11 or 12 base pairs. This pattern suggests direct interactions between slightly underwound co-aligned DNA duplexes engaging once per turn and over many consecutive turns. The process occurs in the absence of MEI3, the only RAD51/DMC1 protein in N. crassa, demonstrating independence from the canonical homology recognition pathway. A new perspective is thus provided for further analysis of the breakage-independent recognition of homology that underlies RIP and, potentially, other processes where sequence-specific pairing of intact chromosomes is involved.

Potential genetic consequences of a recent bottleneck in the Amur tiger of

The Amur tiger, Panthera tigris altaica, is a highly endangered felid whose range and population size has been severely reduced in recent times. At present, the wild population is estimated at 490 individuals, having rebounded from the 20–30 tigers remaining following a severe bottleneck in the 1940s. The current study presents preliminary data on the patterns and levels of genetic variation in the mitochondrial DNA control region using DNA extracted from non-invasively sampled faecal material, collected throughout the entire range of P. t. altaica in the Russian Far East. Analysis of 82 scat samples representing at least 27 individuals revealed extremely low levels of CR haplotype diversity, characterized by a single widespread haplotype (96.4%) and two rare variants, each differing by a single step within the hypervariable I (2.4%) and central conserved regions (1.2%), respectively. A comparison with previous data on cytochrome bvariation in 14 captive individuals revealed a potentially greater amount of genetic variation represented in captivity relative to that found in the wild population. The extremely low levels of mitochondrial DNA variation in the wild population is discussed in light of the demographic processes that might have shaped these patterns as well as the potential bias introduced through analysis of fecal samples. These results highlight the continuing need to assess levels of genetic variation even in recovering populations that are increasing in number and underscore the important role that captive breeding programs may play in preserving remnant genetic diversity of endangered species.

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.

Rotifer rDNA-specific R9 retrotransposable elements generate an exceptionally long target site duplication upon insertion.

Ribosomal DNA genes in many eukaryotes contain insertions of non-LTR retrotransposable elements belonging to the R2 clade. These elements persist in the host genomes by inserting site-specifically into multicopy target sites, thereby avoiding random disruption of single-copy host genes. Here we describe R9 retrotransposons from the R2 clade in the 28S RNA genes of bdelloid rotifers, small freshwater invertebrate animals best known for their long-term asexuality and for their ability to survive repeated cycles of desiccation and rehydration. While the structural organization of R9 elements is highly similar to that of other members of the R2 clade, they are characterized by two distinct features: site-specific insertion into a previously unreported target sequence within the 28S gene, and an unusually long target site duplication of 126 bp. We discuss the implications of these findings in the context of bdelloid genome organization and the mechanisms of target-primed reverse transcription.

A single-copy IS5-like transposon in the genome of a bdelloid rotifer

Abstract In the course of sequencing telomeric chromosomal regions of the bdelloid rotifer Adineta vaga, we encountered an unusual DNA transposon. Unlike other bdelloid and, more generally, eukaryotic transposable elements (TEs), it exhibits similarity to prokaryotic insertion sequences (ISs). Phylogenetic analysis indicates that this transposon, named IS5_Av, is related to the ISL2 group of the IS5 family of bacterial IS elements. Despite the apparent intactness of the single open reading frame coding for a DDE transposase and the perfect identity of its 213-bp terminal inverted repeats (TIRs), the element is present in only one copy per diploid genome. It does not exhibit any detectable levels of transcription, so that its transposase gene appears to be silent in the bdelloid host. Although horizontal transfers of TEs between kingdoms are not known to happen in nature, it appears likely that IS5_Av underwent integration into the A. vaga genome relatively recently, but was not successful in adapting to the new host and failed to increase in copy number. Alternatively, it might be the only known member of a novel eukaryotic DNA TE superfamily which is so rare that its other members, if any, have not yet been identified in eukaryotic genomes sequenced to date.