The evolution of RNAi as a defence against viruses and transposable elements

RNA interference (RNAi) is an important defence against viruses and transposable elements (TEs). RNAi not only protects against viruses by degrading viral RNA, but hosts and viruses can also use RNAi to manipulate each other's gene expression, and hosts can encode microRNAs that target viral sequences. In response, viruses have evolved a myriad of adaptations to suppress and evade RNAi. RNAi can also protect cells against TEs, both by degrading TE transcripts and by preventing TE expression through heterochromatin formation. The aim of our review is to summarize and evaluate the current data on the evolution of these RNAi defence mechanisms. To this end, we also extend a previous analysis of the evolution of genes of the RNAi pathways. Strikingly, we find that antiviral RNAi genes, anti-TE RNAi genes and viral suppressors of RNAi all evolve rapidly, suggestive of an evolutionary arms race between hosts and parasites. Over longer time scales, key RNAi genes are repeatedly duplicated or lost across the metazoan phylogeny, with important implications for RNAi as an immune defence

Drosophila suzukii: the genetic footprint of a recent, world-wide invasion

Native to Asia, the soft-skinned fruit pest Drosophila suzukii has recently invaded the United States and Europe. The eastern United States represents the most recent expansion of their range, and presents an opportunity to test alternative models of colonization history. Here we investigate the genetic population structure of this invasive fruit fly, with a focus on the eastern United States. We sequenced six X-linked gene fragments from 246 individuals collected from a total of 12 populations. We examine patterns of genetic diversity within and between populations and explore alternative colonization scenarios using Approximate Bayesian Computation. Our results indicate high levels of nucleotide diversity in this species and suggest that the recent invasions of Europe and the continental United States are independent demographic events. More broadly speaking, our results highlight the importance of integrating population structure into demographic models, particularly when attempting to reconstruct invasion histories. Finally, our simulation results illustrate the general challenge of reconstructing invasion histories using genetic data and suggest that genome-level data are often required to distinguish among alternative demographic scenarios

Successive Increases in the Resistance of Drosophila to Viral Infection through a Transposon Insertion Followed by a Duplication

To understand the molecular basis of how hosts evolve resistance to their parasites, we have investigated the genes that cause variation in the susceptibility of Drosophila melanogaster to viral infection. Using a host-specific pathogen of D. melanogaster called the sigma virus (Rhabdoviridae), we mapped a major-effect polymorphism to a region containing two paralogous genes called CHKov1 and CHKov2. In a panel of inbred fly lines, we found that a transposable element insertion in the protein coding sequence of CHKov1 is associated with increased resistance to infection. Previous research has shown that this insertion results in a truncated messenger RNA that encodes a far shorter protein than the susceptible allele. This resistant allele has rapidly increased in frequency under directional selection and is now the commonest form of the gene in natural populations. Using genetic mapping and site-specific recombination, we identified a third genotype with considerably greater resistance that is currently rare in the wild. In these flies there have been two duplications, resulting in three copies of both the truncated allele of CHKov1 and CHKov2 (one of which is also truncated). Remarkably, the truncated allele of CHKov1 has previously been found to confer resistance to organophosphate insecticides. As estimates of the age of this allele predate the use of insecticides, it is likely that this allele initially functioned as a defence against viruses and fortuitously “pre-adapted” flies to insecticides. These results demonstrate that strong selection by parasites for increased host resistance can result in major genetic changes and rapid shifts in allele frequencies; and, contrary to the prevailing view that resistance to pathogens can be a costly trait to evolve, the pleiotropic effects of these changes can have unexpected benefits

Ancestral bias in the Hras1 gene and distal Chromosome 7 among inbred mice

Inbred strains of mice vary in their frequency of liver tumors initiated by a mutation in the Hras1 (H-ras) proto-oncogene. We sequenced 4.5 kb of the Hras1 gene on distal Chr 7 in a diverse set of 12 commonly used laboratory inbred strains of mice and detected no sequence variation to account for strain-specific differences in Hras1 mutation prevalence. Furthermore, the Hras1 sequence is essentially monoallelic for an ancestral gene derived from the M. m. domesticus species. To determine if the monoallelism and associated low rate of polymorphism are unique to Hras1 or representative of the general chromosomal locale, we extended the sequence analysis to 12 genes in the final 8 Mb of distal Chr 7. A region of at least 2.5 Mb that encompasses several genes, including Hras1 and the H19/Igf2 loci, demonstrates virtually no sequence variation. The 12 inbred strains share one dominant haplotype derived from the M. m. domesticus allele. Chromosomal regions flanking the monoallelic segment exhibit a significantly higher rate of variation and multiple haplotypes, a majority of which are attributed to M. m. domesticus or M. m. musculus ancestry

Ensuring center quality, proper patient selection and fair access to chimeric antigen receptor T-cell therapy: position statement of the Austrian CAR-T Cell Network

Chimeric antigen receptor T cells (CAR-T cells) are a novel form of cellular immunotherapy for patients with hematologic and oncologic malignancies. Known side effects of these approved cellular immunotherapies are cytokine release syndrome, immune-cell associated neurotoxicity syndrome, cytopenias, infections and long-lasting B cell aplasia. Safe administration of CAR-T cell therapy requires thorough patient selection and patient care in qualified CAR-T cell centers

Adaptive Evolution of a Stress Response Protein

Some cancers are mediated by an interplay between tissue damage, pathogens and localised innate immune responses, but the mechanisms that underlie these linkages are only beginning to be unravelled.Here we identify a strong signature of adaptive evolution on the DNA sequence of the mammalian stress response gene SEP53, a member of the epidermal differentiation complex fused-gene family known for its role in suppressing cancers. The SEP53 gene appears to have been subject to adaptive evolution of a type that is commonly (though not exclusively) associated with coevolutionary arms races. A similar pattern of molecular evolution was not evident in the p53 cancer-suppressing gene.Our data thus raises the possibility that SEP53 is a component of the mucosal/epithelial innate immune response engaged in an ongoing interaction with a pathogen. Although the pathogenic stress mediating adaptive evolution of SEP53 is not known, there are a number of well-known candidates, in particular viruses with established links to carcinoma

High Hemocyte Load Is Associated with Increased Resistance against Parasitoids in Drosophila suzukii, a Relative of D. melanogaster

Among the most common parasites of Drosophila in nature are parasitoid wasps, which lay their eggs in fly larvae and pupae. D. melanogaster larvae can mount a cellular immune response against wasp eggs, but female wasps inject venom along with their eggs to block this immune response. Genetic variation in flies for immune resistance against wasps and genetic variation in wasps for virulence against flies largely determines the outcome of any fly-wasp interaction. Interestingly, up to 90% of the variation in fly resistance against wasp parasitism has been linked to a very simple mechanism: flies with increased constitutive blood cell (hemocyte) production are more resistant. However, this relationship has not been tested for Drosophila hosts outside of the melanogaster subgroup, nor has it been tested across a diversity of parasitoid wasp species and strains. We compared hemocyte levels in two fly species from different subgroups, D. melanogaster and D. suzukii, and found that D. suzukii constitutively produces up to five times more hemocytes than D. melanogaster. Using a panel of 24 parasitoid wasp strains representing fifteen species, four families, and multiple virulence strategies, we found that D. suzukii was significantly more resistant to wasp parasitism than D. melanogaster. Thus, our data suggest that the relationship between hemocyte production and wasp resistance is general. However, at least one sympatric wasp species was a highly successful infector of D. suzukii, suggesting specialists can overcome the general resistance afforded to hosts by excessive hemocyte production. Given that D. suzukii is an emerging agricultural pest, identification of the few parasitoid wasps that successfully infect D. suzukii may have value for biocontrol

Patterns of Selection in Anti-Malarial Immune Genes in Malaria Vectors: Evidence for Adaptive Evolution in LRIM1 in Anopheles arabiensis

Co-evolution between Plasmodium species and its vectors may result in adaptive changes in genes that are crucial components of the vector's defense against the pathogen. By analyzing which genes show evidence of positive selection in malaria vectors, but not in closely related non-vectors, we can identify genes that are crucial for the mosquito's resistance against Plasmodium.We investigated genetic variation of three anti-malarial genes; CEC1, GNBP-B1 and LRIM1, in both vector and non-vector species of the Anopheles gambiae complex. Whereas little protein differentiation was observed between species in CEC1 and GNBP-B1, McDonald-Kreitman and maximum likelihood tests of positive selection show that LRIM1 underwent adaptive evolution in a primary malaria vector; An. arabiensis. In particular, two adjacent codons show clear signs of adaptation by having accumulated three out of four replacement substitutions. Furthermore, our data indicate that this LRIM1 allele has introgressed from An. arabiensis into the other main malaria vector An. gambiae.Although no evidence exists to link the adaptation of LRIM1 to P. falciparum infection, an adaptive response of a known anti-malarial gene in a primary malaria vector is intriguing, and may suggest that this gene could play a role in Plasmodium resistance in An. arabiensis. If so, our data also predicts that LRIM1 alleles in An. gambiae vary in their level of resistance against P. falciparum

A cis-regulatory sequence driving metabolic insecticide resistance in mosquitoes: Functional characterisation and signatures of selection

Although cytochrome P450 (CYP450) enzymes are frequently up-regulated in mosquitoes resistant to insecticides, no regulatory motifs driving these expression differences with relevance to wild populations have been identified. Transposable elements (TEs) are often enriched upstream of those CYP450s involved in insecticide resistance, leading to the assumption that they contribute regulatory motifs that directly underlie the resistance phenotype. A partial CuRE1 (Culex Repetitive Element 1) transposable element is found directly upstream of CYP9M10, a cytochrome P450 implicated previously in larval resistance to permethrin in the ISOP450 strain of Cx. quinquefasciatus, but is absent from the equivalent genomic region of a susceptible strain. Via expression of CYP9M10 in E.coli we have now demonstrated time- and NADPH-dependant permethrin metabolism, prerequisites for confirmation of a role in metabolic resistance, and through qPCR shown that CYP9M10 is >20-fold over-expressed in ISOP450 compared to a susceptible strain. In a fluorescent reporter assay the region upstream of CYP9M10 from ISOP450 drove 10x expression compared to the equivalent region (lacking CuRE1) from the susceptible strain. Close correspondence with the gene expression fold-change implicates the upstream region including CuRE1 as a cis-regulatory element involved in resistance. Only a single CuRE1 bearing allele, identical to the CuRE1 bearing allele in the resistant strain, is found throughout Sub-Saharan Africa, in contrast to the diversity encountered in non-CuRE1 alleles. This suggests a single origin and subsequent spread due to selective advantage. CuRE1 is detectable using a simple diagnostic. When applied to Cx. quinquefasciatus larvae from Ghana we have demonstrated a significant association with permethrin resistance in multiple field sites (mean Odds Ratio = 3.86) suggesting this marker has relevance to natural populations of vector mosquitoes. However, when CuRE1 was excised from the allele used in the reporter assay through fusion PCR, expression was unaffected, indicating that the TE has no direct role in resistance and hence that CuRE1 is acting only as a marker of an as yet unidentified regulatory motif in the association analysis. This suggests that a re-evaluation of the assumption that TEs contribute regulatory motifs involved in gene expression may be necessary

Quantifying Adaptive Evolution in the Drosophila Immune System

It is estimated that a large proportion of amino acid substitutions in Drosophila have been fixed by natural selection, and as organisms are faced with an ever-changing array of pathogens and parasites to which they must adapt, we have investigated the role of parasite-mediated selection as a likely cause. To quantify the effect, and to identify which genes and pathways are most likely to be involved in the host–parasite arms race, we have re-sequenced population samples of 136 immunity and 287 position-matched non-immunity genes in two species of Drosophila. Using these data, and a new extension of the McDonald-Kreitman approach, we estimate that natural selection fixes advantageous amino acid changes in immunity genes at nearly double the rate of other genes. We find the rate of adaptive evolution in immunity genes is also more variable than other genes, with a small subset of immune genes evolving under intense selection. These genes, which are likely to represent hotspots of host–parasite coevolution, tend to share similar functions or belong to the same pathways, such as the antiviral RNAi pathway and the IMD signalling pathway. These patterns appear to be general features of immune system evolution in both species, as rates of adaptive evolution are correlated between the D. melanogaster and D. simulans lineages. In summary, our data provide quantitative estimates of the elevated rate of adaptive evolution in immune system genes relative to the rest of the genome, and they suggest that adaptation to parasites is an important force driving molecular evolution