Genome-Wide Characterization of Endogenous Retroviruses in the Bat Myotis lucifugus Reveals Recent and Diverse Infections

Bats are increasingly recognized as reservoir species for a variety of zoonotic viruses that pose severe threats to human health. While many RNA viruses have been identified in bats, little is known about bat retroviruses. Endogenous retroviruses (ERVs) represent genomic fossils of past retroviral infections and, thus, can inform us on the diversity and history of retroviruses that have infected a species lineage. Here, we took advantage of the availability of a high-quality genome assembly for the little brown bat, Myotis lucifugus, to systematically identify and analyze ERVs in this species. We mined an initial set of 362 potentially complete proviruses from the three main classes of ERVs, which were further resolved into 13 major families and 86 subfamilies by phylogenetic analysis. Consensus or representative sequences for each of the 86 subfamilies were then merged to the Repbase collection of known ERV/long terminal repeat (LTR) elements to annotate the retroviral complement of the bat genome. The results show that nearly 5% of the genome assembly is occupied by ERV-derived sequences, a quantity comparable to findings for other eutherian mammals. About one-fourth of these sequences belong to subfamilies newly identified in this study. Using two independent methods, intraelement LTR divergence and analysis of orthologous loci in two other bat species, we found that the vast majority of the potentially complete proviruses identified in M. lucifugus were integrated in the last ∼25 million years. All three major ERV classes include recently integrated proviruses, suggesting that a wide diversity of retroviruses is still circulating in Myotis bats.     

One-third rules with equality: Second-order evolutionary stability conditions in finite populations

The one-third law of evolutionary dynamics [Nowak et al. 2004. Emergence of cooperation and evolutionary stability in finite populations. Nature 428, 246-650] describes a robustness criterion for evolution in a finite population: If at an A-frequency of 1/3, the fitness of an A player is greater (smaller) than the fitness of a B player, then a single A mutant that appears in a population of otherwise all B has a fixation probability greater (smaller) than the neutral threshold 1/N, the inverse population size. We examine the case where at an A-frequency of 1/3, the fitness of an A player is exactly equal to the fitness of a B player. We find that in this case the relative magnitude of the cross payoffs matters: If the payoff of A against B is larger (smaller) than the payoff of B against A, then a single A mutant has a fixation probability larger (smaller) than 1/N. If the cross payoffs coincide, we are in the special case of a partnership game, where the deviation cost from an inefficient equilibrium is exactly balanced by the potential gain of switching to the payoff dominant equilibrium. We show that in this case the fixation probability of A is lower than 1/N. Finally, we illustrate our findings by a language game with differentiated costs of signals.     

Replication of nonautonomous retroelements in soybean appears to be both recent and common

Retrotransposons and their remnants often constitute more than 50% of higher plant genomes. Although extensively studied in monocot crops such as maize (Zea mays) and rice (Oryza sativa), the impact of retrotransposons on dicot crop genomes is not well documented. Here, we present an analysis of retrotransposons in soybean (Glycine max). Analysis of approximately 3.7 megabases (Mb) of genomic sequence, including 0.87 Mb of pericentromeric sequence, uncovered 45 intact long terminal repeat (LTR)-retrotransposons. The ratio of intact elements to solo LTRs was 8:1, one of the highest reported to date in plants, suggesting that removal of retrotransposons by homologous recombination between LTRs is occurring more slowly in soybean than in previously characterized plant species. Analysis of paired LTR sequences uncovered a low frequency of deletions relative to base substitutions, indicating that removal of retrotransposon sequences by illegitimate recombination is also operating more slowly. Significantly, we identified three subfamilies of nonautonomous elements that have replicated in the recent past, suggesting that retrotransposition can be catalyzed in trans by autonomous elements elsewhere in the genome. Analysis of 1.6 Mb of sequence from Glycine tomentella, a wild perennial relative of soybean, uncovered 23 intact retroelements, two of which had accumulated no mutations in their LTRs, indicating very recent insertion. A similar pattern was found in 0.94 Mb of sequence from Phaseolus vulgaris (common bean). Thus, autonomous and nonautonomous retrotransposons appear to be both abundant and active in Glycine and Phaseolus. The impact of nonautonomous retrotransposon replication on genome size appears to be much greater than previously appreciated.     

Expansion and Accelerated Evolution of 9-Exon Odorant Receptors in Polistes Paper Wasps

Independent origins of sociality in bees and ants are associated with independent expansions of particular odorant receptor (OR) gene subfamilies. In ants, one clade within the OR gene family, the 9-exon subfamily, has dramatically expanded. These receptors detect cuticular hydrocarbons (CHCs), key social signaling molecules in insects. It is unclear to what extent 9-exon OR subfamily expansion is associated with the independent evolution of sociality across Hymenoptera, warranting studies of taxa with independently derived social behavior. Here, we describe OR gene family evolution in the northern paper wasp, Polistes fuscatus, and compare it to four additional paper wasp species spanning ∼40 million years of evolutionary divergence. We find 200 putatively functional OR genes in P. fuscatus, matching predictions from neuroanatomy, and more than half of these are in the 9-exon subfamily. Most OR gene expansions are tandemly arrayed at orthologous loci in Polistes genomes, and microsynteny analysis shows species-specific gain and loss of 9-exon ORs within tandem arrays. There is evidence of episodic positive diversifying selection shaping ORs in expanded subfamilies. Values of omega (d_N/d_S) are higher among 9-exon ORs compared to other OR subfamilies. Within the Polistes OR gene tree, branches in the 9-exon OR clade experience relaxed negative (relaxed purifying) selection relative to other branches in the tree. Patterns of OR evolution within Polistes are consistent with 9-exon OR function in CHC perception by combinatorial coding, with both natural selection and neutral drift contributing to interspecies differences in gene copy number and sequence.     

Retrotransposon evolution in diverse plant genomes

Retrotransposon or retrotransposon-like sequences have been reported to be conserved components of cereal centromeres. Here we show that the published sequences are derived from a single conventional Ty3-gypsy family or a nonautonomous derivative. Both autonomous and nonautonomous elements are likely to have colonized Poaceae centromeres at the time of a common ancestor but have been maintained since by active retrotransposition. The retrotransposon family is also present at a lower copy number in the Arabidopsis genome, where it shows less pronounced localization. The history of the family in the two types of genome provides an interesting contrast between "boom and bust" and persistent evolutionary patterns.     

Identification of putative nonautonomous transposable elements associated with several transposon families inCaenorhabditis elegans

Putative nonautonomous transposable elements related to the autonomous transposons Tc1, Tc2, Tc5, andmariner were identified in theC. elegans database by computational analysis. These elements are found throughout theC. elegans genome and are defined by terminal inverted repeats with regions of sequence similarity, or identity, to the autonomous transposons. Similarity between loci containing related nonautonomous elements ends at, or near, the boundaries of the terminal inverted repeats. In most cases the terminal inverted repeats of the putative nonautonomous transposable elements are flanked by potential target-site duplications consistent with the associated autonomous elements. The nonautonomous elements identified vary considerably in size (from 100 by to 1.5 kb in length) and copy number in the available database and are localized to introns and flanking regions of a wide variety ofC. elegans genes. Correspondence to: W. Belknap     

Context-dependent role of ATG4B as target for autophagy inhibition in prostate cancer therapy

ATG4B belongs to the autophagin family of cysteine proteases required for autophagy, an emerging target of cancer therapy. Developing pharmacological ATG4B inhibitors is a very active area of research. However, detailed studies on the role of ATG4B during anticancer therapy are lacking. By analyzing PC-3 and C4-2 prostate cancer cells overexpressing dominant negative ATG4B^C74Ain vitro and in vivo, we show that the effects of ATG4B^C74A are cell type, treatment, and context-dependent. ATG4B^C74A expression can either amplify the effects of cytotoxic therapies or contribute to treatment resistance. Thus, the successful clinical application of ATG4B inhibitors will depend on finding predictive markers of response.     

The Cassandra retrotransposon landscape in sugar beet (Beta vulgaris) and related Amaranthaceae: recombination and re-shuffling lead to a high structural variability

Background and AimsPlant genomes contain many retrotransposons and their derivatives, which are subject to rapid sequence turnover. As non-autonomous retrotransposons do not encode any proteins, they experience reduced selective constraints leading to their diversification into multiple families, usually limited to a few closely related species. In contrast, the non-coding Cassandra terminal repeat retrotransposons in miniature (TRIMs) are widespread in many plants. Their hallmark is a conserved 5S rDNA-derived promoter in their long terminal repeats (LTRs). As sugar beet (Beta vulgaris) has a well-described LTR retrotransposon landscape, we aim to characterize TRIMs in beet and related genomes.MethodsWe identified Cassandra retrotransposons in the sugar beet reference genome and characterized their structural relationships. Genomic organization, chromosomal localization, and distribution of Cassandra-TRIMs across the Amaranthaceae were verified by Southern and fluorescent in situ hybridization.Key resultsAll 638 Cassandra sequences in the sugar beet genome contain conserved LTRs and thus constitute a single family. Nevertheless, variable internal regions required a subdivision into two Cassandra subfamilies within B. vulgaris. The related Chenopodium quinoa harbours a third subfamily. These subfamilies vary in their distribution within Amaranthaceae genomes, their insertion times and the degree of silencing by small RNAs. Cassandra retrotransposons gave rise to many structural variants, such as solo LTRs or tandemly arranged Cassandra retrotransposons. These Cassandra derivatives point to an interplay of template switch and recombination processes – mechanisms that likely caused Cassandra’s subfamily formation and diversification.ConclusionsWe traced the evolution of Cassandra in the Amaranthaceae and detected a considerable variability within the short internal regions, whereas the LTRs are strongly conserved in sequence and length. Presumably these hallmarks make Cassandra a prime target for unequal recombination, resulting in the observed structural diversity, an example of the impact of LTR-mediated evolutionary mechanisms on the host genome.     

Three highly divergent subfamilies of the impala transposable element coexist in the genome of the fungus Fusarium oxysporum

The transposable element impala is a member of the widespread superfamily of Tc1-mariner transposons, identified in the genome of the plant pathogenic fungus Fusarium oxysporum. This element is present in a low copy number and is actively transposed in the F.?oxysporum strain F24 that is pathogenic for melons. The structure of the impala family was investigated by cloning and sequencing all the genomic copies. The analysis revealed that this family is composed of full-length and truncated copies. Four copies contained a long open reading frame that could potentially encode a transposase of 340 amino acids. The presence of conserved functional domains (a nuclear localisation signal, a catalytic DDE domain and a DNA-binding domain) suggests that these four copies may be autonomous elements. Sequence comparisons and phylogenetic analysis of the impala copies defined three subfamilies, which differ by a high level of nucleotide polymorphism (around 20%). The coexistence of these divergent subfamilies in the same genome may indicate that the impala family is of ancient origin and/or that it arose by successive horizontal transmission events.     

Characterization of the intragenomic spread of the human endogenous retrovirus family HERV-W

This study examines the intragenomic spread of the human endogenous retrovirus family HERV-W from insertions present within the draft sequence of the human genome. Identification of shared diagnostic differences and phylogenetic analyses revealed the existence of three main subfamilies. The average divergence between sequences for each of the subfamilies suggests that most of the HERV-W elements were inserted within the genome during a short period of evolutionary time. Each one of the subfamilies consists of two types of insertions, the expected proviral sequences and other sequences resembling the structure of processed retrogenes. These HERV-W retrosequences extend from the R region of the 5' long-terminal repeat (LTR) to the R region of the 3' LTR (as viral genomic RNAs), end in poly(A) 3' tails, and are flanked by direct repeats longer than the proviral integrations. Furthermore, several of the HERV-W retrosequences are 5'-truncated at different sites. I suggest the involvement of the L1 machinery in these integrations and discuss the characteristic features of the evolutionary history of HERV-W, with emphasis on the putative impact of HERV-W retrosequence integrations on the mammalian genome.     

The secondary attachment site for bacteriophage lambda in the proA/B gene of Escherichia coli

We have determined the nucleotide sequence of a secondary λ attachment site in $\text{pro}\text{A}\text{B}$, a site that accounts for 3% of lysogens isolated from Escherichia coli strains deleted for the primary site. Direct sequence analysis of the transducing bacteriophages carrying the left and right att junctions, as well as the recombinant pro⁺ phage reveals that the $\text{pro}\text{A}\text{B}$ site shares an 11-nucleotide interrupted homology with the core sequence of the primary site. We have compared the $\text{pro}\text{A}\text{B}$att site with other secondary attachment sites to gain insights into the structural features important for λ integration.     

A Time-Calibrated Road Map of Brassicaceae Species Radiation and Evolutionary History

The Brassicaceae include several major crop plants and numerous important model species in comparative evolutionary research such as Arabidopsis, Brassica, Boechera, Thellungiella, and Arabis species. As any evolutionary hypothesis needs to be placed in a temporal context, reliably dated major splits within the evolution of Brassicaceae are essential. We present a comprehensive time-calibrated framework with important divergence time estimates based on whole-chloroplast sequence data for 29 Brassicaceae species. Diversification of the Brassicaceae crown group started at the Eocene-to-Oligocene transition. Subsequent major evolutionary splits are dated to ∼20 million years ago, coinciding with the Oligocene-to-Miocene transition, with increasing drought and aridity and transient glaciation events. The age of the Arabidopsis thaliana crown group is 6 million years ago, at the Miocene and Pliocene border. The overall species richness of the family is well explained by high levels of neopolyploidy (43% in total), but this trend is neither directly associated with an increase in genome size nor is there a general lineage-specific constraint. Our results highlight polyploidization as an important source for generating new evolutionary lineages adapted to changing environments. We conclude that species radiation, paralleled by high levels of neopolyploidization, follows genome size decrease, stabilization, and genetic diploidization.     

Contrasting evolutionary patterns and target specificities among three Tourist-like MITE families in the maize genome

Miniature inverted-repeat transposable elements (MITEs) are short, non autonomous DNA elements that are widespread and abundant in plant genomes. The high sequence and size conservation observed in many MITE families suggest that they have spread recently throughout their respective host genomes. Here we present a maize genome wide analysis of three Tourist-like MITE families, mPIF, and two previously uncharacterized families, ZmV1 and Zead8. We undertook a bioinformatic analysis of MITE insertion sites, developed methyl-sensitive transposon display (M-STD) assays to estimate the associated level of CpG methylation at MITE flanking regions, and conducted a population genetics approach to investigate MITE patterns of expansion. Our results reveal that the three MITE families insert into genomic regions that present specific molecular features: they are preferentially AT rich, present low level of cytosine methylation as compared to the LTR retrotransposon Grande, and target site duplications are flanked by large and conserved palindromic sequences. Moreover, the analysis of MITE distances from predicted genes shows that 73% of 263 copies are inserted at less than 5 kb from the nearest predicted gene, and copies from Zead8 family are significantly more abundant upstream of genes. By employing a population genetic approach we identified contrasting patterns of expansion among the three MITE families. All elements seem to have inserted roughly 1 million years ago but ZmV1 and Zead8 families present evidences for activity of several master copies within the last 0.4 Mya.     

Electron transfer in reaction centers of Rhodopseudomonas sphaeroides. I. Determination of the charge recombination pathway of D+QAQ−B and free energy and kinetic relations between Q−AQB and QAQ−B

The electron-transfer reactions and thermodynamic equilibria involving the quinone acceptor complex in bacterial reaction centers from R. sphaeroides were investigated. The reactions are described by the scheme: We found that the charge recombination pathway of D⁺Q_AQ^?_B proceeds via the intermediate state D⁺Q^?_AQ_B, the direct pathway contributing less than approx. 5% to the observed recombination rate. The method used to obtain this result was based on a comparison of the kinetics predicted for the indirect pathway (given by the product k_AD-times the fraction of reaction centers in the Q^?_AQ_B state) with the observed recombination rate, k^obs_{D⁺ →D}. The kinetic measurements were used to obtain the pH dependence (6.1 ? pH ? 11.7) of the free energy difference between the states Q^?_AQ_B and Q_AQ^?_B. At low pH (less than 9) Q_AQ^?_B is stabilized relative to Q^?_AQ_B by 67 meV, whereas at high pH Q^?_AQ_B is energetically favored. Both Q^?_A and Q^?_B associate with a proton, with pK values of 9.8 and 11.3, respectively. The stronger interaction of the proton with Q^?_B provides the driving force for the forward electron transfer.     

Chromosome morphology of the czechoslovak species of the genusScorzonera

Karyotypic formulae of theScorzonera L. species are as follows:S. purpurea L.: K (2n)=14=8 A^m+6 B^sm and K (2n)=14+1=9 A^m+6 B^sm;S. austriaca Willd.: K (2n)=14=6 A^m+6 B^sm+2 C^st;S. humilis L.: K (2n)=14=12 A^m+2 B^sm;S. parviflora Jacq.: K (2n)=14=10 A^m+4 B^sm;S. hispanica L.: K (2n)=12 A^m+2 B^sm. The results of the study of the karyotypes of the generaScorzonera L.,Tragopogon L. andPodospermum DC. are summarized.     

Structure, divergence, and distribution of the CRR centromeric retrotransposon family in rice

The centromeric retrotransposon (CR) family in the grass species is one of few Ty3-gypsy groups of retroelements that preferentially transpose into highly specialized chromosomal domains. It has been demonstrated in both rice and maize that CRR (CR of rice) and CRM (CR of maize) elements are intermingled with centromeric satellite DNA and are highly concentrated within cytologically defined centromeres. We collected all of the CRR elements from rice chromosomes 1, 4, 8, and 10 that have been sequenced to high quality. Phylogenetic analysis revealed that the CRR elements are structurally diverged into four subfamilies, including two autonomous subfamilies (CRR1 and CRR2) and two nonautonomous subfamilies (noaCRR1 and noaCRR2). The CRR1/CRR2 elements contain all characteristic protein domains required for retrotransposition. In contrast, the noaCRR elements have different structures, containing only a gag or gag-pro domain or no open reading frames. The CRR and noaCRR elements share substantial sequence similarity in regions required for DNA replication and for recognition by integrase during retrotransposition. These data, coupled with the presence of young noaCRR elements in the rice genome and similar chromosomal distribution patterns between noaCRR1 and CRR1/CRR2 elements, suggest that the noaCRR elements were likely mobilized through the retrotransposition machinery from the autonomous CRR elements. Mechanisms of the targeting specificity of the CRR elements, as well as their role in centromere function, are discussed.