UC Berkeley

Although most adaptation has long been thought to occur gradually over evolutionary time, a growing body of research shows that adaptation can also happen by “leaps and bounds” - specifically, by horizontal gene transfer (HGT) between microbes to eukaryotic genomes. Even more recently, such HGT has also been shown to have played an important role in the evolution of eukaryotes. This dissertation research characterizes the evolution and function of cytolethal distending toxin B (cdtB), apoptosis inducing protein of 56 kDa (aip56), and other microbially-derived, toxin-encoding genes, which were transferred from phages to insects.

The first chapter describes and characterizes the HGT of cdtB, which encodes an apoptotic DNase toxin, into the genomes of vinegar flies (Diptera: Drosophilidae) and aphids (Hemiptera: Aphididae) (Verster et al 2019). Phylogenetic analysis of CdtB shows that the orthologs most closely related to insect CdtB are from bacteriophage that infect Hamiltonella defensa, a bacterial mutualistic symbiont of aphids that confers resistance to parasitoid wasps. In drosophilids, cdtB orthologs are highly expressed during the parasitoid-prone larval stage and encode a protein with ancestral DNase activity.

The second chapter broadens the focus of HGT in insects, reporting HGT of five microbially-derived toxin genes (aip56, cdtB, lysozyme, rhs, sltxB) in the galling midge fly family Cecidomyiidae (Verster et al 2021). These genes were transferred to the genomes of midge species from a variety of donors including fungi, bacteria, and viruses – highlighting the remarkable evolutionary convergence and repeatability of toxin gain in insects. The question of how HGT occurs is also addressed. While the actual mechanisms leading to HGT between bacteria and eukaryotes are relatively unknown, a novel phylogenetic analysis is utilized to determine that species living in similar environments, such as endosymbionts and their hosts, are more likely to exchange genes than species living in dissimilar environments. This research highlights that HGT between insects and phage is common, and likely facilitated by their intimate mutualisms (Verster et al 2021).

The third chapter shows how horizontally transferred cdtB and aip56 have led to the evolution of a novel arm of the insect immune system in the fly Drosophila ananassae. When encoded by a prophage that infects aphid bacterial endosymbionts, CdtB and AIP56 are associated with increased aphid survival against parasitic wasps. However, an HGT event brought cdtB and aip56 directly into the genome of the insect, side-stepping the symbiont. The function of these genes in insects was unknown, though an immunity role (defense against parasitoid wasps) is clearly possible. Phylogenetic analysis, gene expression, biochemistry, immunofluorescence, and CRISPR/Cas9-mediated knockout studies show that these two horizontally transferred, phage-derived toxin genes are now essential for resistance to parasitoid wasp infection in D. ananassae.