Ancient Gene Capture and Recent Gene Loss Shape the Evolution of Orthopoxvirus-Host Interaction Genes

Orthopoxviruses (ORPV) include smallpox (variola) virus, one of the most devastating human pathogens, and vaccinia virus, comprising the vaccine used for smallpox eradication. Among roughly 200 ORPV genes, about half are essential for genome replication and expression as well as virion morphogenesis, whereas the remaining half consists of accessory genes counteracting the host immune response.

Cascade evolution in poxviruses: retrotransposon-mediated host gene capture, complementation and recombination

There is ample phylogenetic evidence that many critical virus functions, like immune evasion, evolved by the acquisition of genes from their hosts by horizontal gene transfer (HGT). However, the lack of an experimental system has prevented a mechanistic understanding of this process. We developed a model to elucidate the mechanisms of HGT into poxviruses. All identified gene capture events showed signatures of LINE-1-mediated retrotransposition. Integrations occurred across the genome, in some cases knocking out essential viral genes. These essential gene knockouts were rescued through a process of complementation by the parent virus followed by non-homologous recombination to generate a single competent virus. This work links multiple evolutionary mechanisms into one adaptive cascade and identifies host retrotransposons as major drivers for virus evolution.

Differentially Marked IncP-1β R751 Plasmids for Cloning via Recombineering and Conjugation

We demonstrate here for the first time the use of an IncP-1β plasmid, R751, as a gene capture vehicle for recombineering/conjugation strategies to clone large segments of bacterial genomes (20 – 100 + Kb). We designed R751 derivatives containing alternative markers for greater flexibility when using the R751 vehicle across different bacteria. These markers are removable if desired as part of the cloning procedure (with no extra steps needed). We demonstrated utility via cloning of 38 and 22 kb genomic segments from Salmonella enterica serovar Typhimurium and Escherichia coli, respectively. The plasmids expand the options available for use in recombineering/conjugation-based cloning applications.

Gene capture by transposable elements leads to epigenetic conflict in maize

Plant transposable elements (TEs) regularly capture fragments of genes. When the host silences these TEs, siRNAs homologous to the captured regions may also target the genes. This epigenetic cross-talk establishes an intragenomic conflict: silencing the TEs has the cost of silencing the genes. If genes are important, however, natural selection may maintain function by moderating the silencing response, which may also advantage the TEs. Here, we examined this model by focusing on three TE families in maize: Helitrons, Pack-MULEs and Sirevirus LTR retrotransposons. We documented 1,263 TEs containing exon fragments from 1,629 donor genes. Consistent with epigenetic conflict, donor genes mapped more siRNAs and were more methylated than genes with no evidence of capture. However, these patterns differed between syntelog vs. translocated donor genes. Syntelogs appeared to maintain function, as measured by gene expression, consistent with moderation of silencing for functionally important genes. Epigenetic marks did not spread beyond their captured regions and 24nt cross-talk siRNAs were linked with CHH methylation. Translocated genes, in contrast, bore the signature of silencing by being highly methylated and less expressed. They were also overrepresented among donor genes, suggesting a link between capture and gene movement. The evidence for an advantage to TEs was less obvious. TEs with captured fragments were older, mapped fewer siRNAs and were slightly less methylated than TEs without captured fragments but showed no evidence of increased copy numbers. Altogether, our results demonstrate that TE capture triggers an epigenetic conflict for important genes, but it may lead to pseudogenization for less constrained genes.

orthoCapture: Facilitating Gene Capture Probe Design for Non-Model Species

In non-model species, targeted gene capture (selective enrichment of specific genomic regions of interest) applications in molecular ecology have been limited by the practicalities of capture design. Currently, the minimal requirement for designing capture probes is a transcriptome, or established reference genome for the species of interest. When an established, annotated reference genome is unavailable, one common approach is to design probes from annotated reference genomes (or transcriptomes) of related species. Unfortunately, as divergence between probes and the genome of interest increases, such as occurs during directional selection, capture performance decreases. Here I introduce orthoCapture, a tool to overcome such limitations by mining unannotated whole-genome sequence (WGS) data from non-model species and/or their close relatives to allow probe design using multiple genomic sources. orthoCapture finds orthologs in WGS data from multiple related species to create a set of exon sequences that encompasses the diversity of the exons of interest. These “design sequences” can then be used to design capture probes for the species of interest. orthoCapture thus eliminates the need for transcriptome or whole-genome sequencing for bait capture experiments, making this technique accessible for molecular ecology and conservation studies. Use of orthoCapture is via command-line interface on Unix systems, and requires the input of a gene sequence from an unrelated annotated genome and a fasta database from a target, unannotated genome (e.g., whole-genome shotgun contigs). The output, sequence templates from the nonannotated genomic data, allows probe creation by any commercial company providing gene capture services.