Article Parallel Histories of Horizontal Gene Transfer Facilitated Extreme Reduction of Endosymbiont Genomes in Sap-Feeding Inse
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Parallel Histories of Horizontal Gene Transfer Facilitated Extreme Reduction of Endosymbiont Genomes in Sap-Feeding Insects Daniel B. Sloan,*,1 Atsushi Nakabachi,2 Stephen Richards,3 Jiaxin Qu,3 Shwetha Canchi Murali,3 Richard A. Gibbs,3 and Nancy A. Moran4 1Department of Biology, Colorado State University 2Electronics-Inspired Interdisciplinary Research Institute, Toyohashi University of Technology, Toyohashi, Japan 3Human Genome Sequencing Center, Baylor College of Medicine, Houston, Texas 4Section of Integrative Biology, University of Texas *Corresponding author: E-mail: [email protected]. Associate editor: Andrew Roger Downloaded from Abstract Bacteria confined to intracellular environments experience extensive genome reduction. In extreme cases, insect endo- symbionts have evolved genomes that are so gene-poor that they blur the distinction between bacteria and endosym- biotically derived organelles such as mitochondria and plastids. To understand the host’s role in this extreme gene loss, http://mbe.oxfordjournals.org/ we analyzed gene content and expression in the nuclear genome of the psyllid Pachypsylla venusta, a sap-feeding insect that harbors an ancient endosymbiont (Carsonella) with one of the most reduced bacterial genomes ever identified. Carsonella retains many genes required for synthesis of essential amino acids that are scarce in plant sap, but most of these biosynthetic pathways have been disrupted by gene loss. Host genes that are upregulated in psyllid cells housing Carsonella appear to compensate for endosymbiont gene losses, resulting in highly integrated metabolic pathways that mirror those observed in other sap-feeding insects. The host contribution to these pathwaysismediatedbyacombi- nation of native eukaryotic genes and bacterial genes that were horizontally transferred from multiple donor lineages early in the evolution of psyllids, including one gene that appears to have been directly acquired from Carsonella.By at University of San Diego - Copley Library on February 11, 2015 comparing the psyllid genome to a recent analysis of mealybugs, we found that a remarkably similar set of functional pathways have been shaped by independent transfers of bacterial genes to the two hosts. These results show that horizontal gene transfer is an important and recurring mechanism driving coevolution between insects and their bacterial endosymbionts and highlight interesting similarities and contrasts with the evolutionary history of mitochon- dria and plastids. Key words: amino acid biosynthesis, endosymbionts, lateral gene transfer, Pachypsylla venusta, psyllids. Article Introduction 2006; McCutcheon et al. 2009b; McCutcheon and Moran One of the most well-documented patterns in genome evo- 2010; McCutcheon and von Dohlen 2011; Bennett and lution is the reduction in genome size and gene content ex- Moran 2013). The first example of such extreme genome hibited by obligately intracellular bacteria (Andersson and reduction was found in Candidatus Carsonella ruddii (hereaf- Kurland 1998; Moran and Wernegreen 2000; McCutcheon ter referred to as Carsonella), a vertically transmitted gamma- and Moran 2012). This pattern can largely be explained by proteobacterial endosymbiont that is present in all psyllids a combination of relaxed functional constraint on redundant (Hemiptera: Sternorrhyncha) examined to date (Thao et al. or superfluous genes (Andersson et al. 1998), inefficient pu- 2000). Gene loss appears to be an ongoing process in this rifying selection in a host-restricted environment (Moran endosymbiont with observed Carsonella genome sizes rang- 1996), and a mutational bias favoring deletions over insertions ing in size from 158 to 174 kb across different host species (Mira et al. 2001; Kuo and Ochman 2009). However, extreme (Nakabachi et al. 2006, 2013; Sloan and Moran 2012). These cases of genome reduction involving the loss of seemingly tiny genomes lack many widely conserved genes involved in essential genes remain puzzling. In particular, many nutri- DNA replication, transcription, and translation, and most of tional endosymbionts in sap-feeding insects maintain ge- their essential amino acid biosynthesis pathways are incom- nomes that are less than 200 kb in size and that have lost plete (Tamames et al. 2007). Similar to other nutritional en- genes thought to be essential for basic cellular processes, in- dosymbionts in sap-feeding insects, Carsonella is sequestered cluding the primary symbiotic role of these bacteria—the within bacteriocytes, specialized host cells found in an ab- biosynthesis of amino acids that are lacking in their hosts’ dominal organ known as the bacteriome (Buchner 1965; diets and that cannot be made by animals (Nakabachi et al. Fukatsu and Nikoh 1998). ß The Author 2014. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: [email protected] Mol. Biol. Evol. 31(4):857–871 doi:10.1093/molbev/msu004 Advance Access publication January 6, 2014 857 Sloan et al. doi:10.1093/molbev/msu004 MBE The exceptional gene loss observed in Carsonella suggests many of which appear to directly complement Tremblaya three possible mechanisms (none of which are mutually gene losses (Husnik et al. 2013). Interestingly, there is no ev- exclusive). First, modification of highly conserved cellular pro- idence that any of the HGT events in aphids or mealybugs cesses or selection for multifunctional proteins could have involve the direct transfer of functional genes from their re- allowed the endosymbiont to dispense with otherwise spective obligate endosymbionts, contrasting with the mas- “essential” genes (McCutcheon 2010; Kelkar and Ochman sive gene movement from mitochondria and plastids to the 2013). Second, as commonly observed in sap-feeding insects, nucleus during the history of eukaryotic genome evolution the presence of additional endosymbionts may compensate (Timmis et al. 2004). Instead, genes appear to have been ac- for gene losses (Wu et al. 2006; McCutcheon et al. 2009a; quired from other insect-associated bacteria, such as McCutcheon and Moran 2010; Lamelas et al. 2011; Arsenophonus, Cardinium, Rickettsia, Sodalis, Serratia,and McCutcheon and von Dohlen 2011; Nakabachi et al. 2013). Wolbachia (Nikoh and Nakabachi 2009; Nikoh et al. 2010; There is evidence supporting a role for other endosymbionts Husnik et al. 2013). However, the pea aphid genome does in some psyllids. For example, the loss of Carsonella genes contain two small, nonfunctional fragments of Buchnera Downloaded from necessary for arginine and tryptophan biosynthesis in the DNA, indicating that genetic transfer to insects from their Eucalyptus-feeding psyllid Ctenarytaina eucalypti appears to obligate endosymbionts is possible (Nikoh et al. 2010). be offset by a second bacterial endosymbiont that retains By analyzing genome-wide patterns of host gene expres- these genes (Sloan and Moran 2012). However, some psyllids, sion in the hackberry petiole gall psyllid Pachypsylla venusta, including hackberry-feeding species in the genus Pachypsylla, we show that extreme genome reduction in Carsonella and http://mbe.oxfordjournals.org/ lack additional symbiotic partners (Thao et al. 2000) but still Tremblaya has involved a number of striking evolutionary maintain Carsonella strains with highly reduced genomes that parallels, including independent HGT events affecting the have lost genes required for synthesis of some essential amino same functional pathways. Although multiple bacterial line- acids (Nakabachi et al. 2006). Finally, missing Carsonella genes ages have acted as HGT donors, we find that psyllids have may have been functionally replaced by host-encoded pro- acquired at least one gene directly from Carsonella. teins, including pre-existing eukaryotic homologs (Wilson et al. 2010; Price et al. 2011) and/or genes of bacterial origin Results acquired by horizontal gene transfer (HGT) (e.g., Nowack and Grossman 2012). Identification of Preferentially Expressed Genes in the at University of San Diego - Copley Library on February 11, 2015 Here, we focus on the last of these three potential mech- Psyllid Bacteriome by mRNA-Seq anisms to address the question of whether Carsonella has Deep strand-specific sequencing of polyadenylated tran- followed a similar trajectory as the evolution of mitochondria scripts (mRNA-seq) in P. venusta nymphs isolated from hack- and plastids from bacterial progenitors, in which genetic con- berry galls was used to broadly identify and characterize trol was largely shifted from the endosymbiont (organelle) to psyllid genes. Furthermore, by comparing expression levels the host (nuclear) genome through a combination of direct between the bacteriome and the rest of the insect body, we gene transfer and functional gene replacement. Recent func- identified dramatic differences in host gene expression (fig. 1). tional genomic analyses in two sap-feeding insects, aphids and After quality filtering, we obtained between 14.3 and 20.9 mealybugs, have supported the prediction that host genomes million Illumina read pairs for each of three replicate play a central role in endosymbiont metabolism and provided mRNA-seq libraries derived from isolated P. venusta bacter- evidence that both insects have acquired multiple genes of iomes and three corresponding libraries derived from the re- bacterial origin, many of which are now preferentially maining body tissues (table 1). Despite the polyA selection expressed in