Footprints of parasitism in the genome of the parasitic flowering plant Cuscuta campestris

Abstract

A parasitic lifestyle, where plants procure some or all of their nutrients from other living plants, has evolved independently in many dicotyledonous plant families and is a major threat for agriculture globally. Nevertheless, no genome sequence of a parasitic plant has been reported to date. Here we describe the genome sequence of the parasitic field dodder, Cuscuta campestris. The genome contains signatures of a fairly recent whole-genome duplication and lacks genes for pathways superfluous to a parasitic lifestyle. Specifically, genes needed for high photosynthetic activity are lost, explaining the low photosynthesis rates displayed by the parasite. Moreover, several genes involved in nutrient uptake processes from the soil are lost. On the other hand, evidence for horizontal gene transfer by way of genomic DNA integration from the parasite's hosts is found. We conclude that the parasitic lifestyle has left characteristic footprints in the C. campestris genome.

Fig. 1

Cuscuta campestris—host plant relationship. a Cuscuta campestris (yellow) infecting Pelargonium zonale. b Close-up of a parasitic vine entwining and infecting a host shoot. c Cross section through an infection site. The right half of the micrograph was overlayed with color to emphasize the border between C. campestris (yellow) and the P. zonale stem (green). d Simplified phylogenetic tree of reported host plant species of C. campestris, showing the variety of angiosperm orders (named on the right) that contain host species (according to refs. ^–). The gray background highlights the single described monocot host. The asterisk indicates the closest phylogenetic relative of C. campestris (Convolvulaceae) within the host groups

Fig. 2

Genome features. a Feulgen-stained metaphase chromosomes of C. campestris (2n = 56). The scale bar represents 10 µm. b Insertion age distribution of C. campestris full-length LTR retrotransposons in comparison to tomato (Solanum lycopersicum) as another Solanales representative. Insertion events of long terminal repeat (LTR) retrotransposons were dated by comparing the divergence between the 5′ and 3′ long terminal repeats. A random mutation rate of 7*10⁻⁹ was used to convert the sequence divergence into age (in million years)

Fig. 3

Duplication and loss of genes in the C. campestris genome. a Overview over 14,626 asterid orthogroups from C. campestris, Ipomoea nil, Solanum lycopersicum, Mimulus guttatus, and Daucus carota. The Venn diagram shows the number of overlapping and exclusive orthogroups found in the five species. The bar diagram below summarizes gene content in orthogroups in C. campestris and eight divergent dicot species. Indicated in black are orthogroups with exactly one member in each of these species (202 OGs). Orthogroups missing in the respective species only, but that are otherwise present in all species once (exactly 0:1, named:other species, striped) or more than once (ratio 0:1 or more, solid color) are shown in orange and are underlined in the Venn diagram. Duplicated orthogroups are defined as having exactly 2:1 genes per species (striped purple) or as having a ratio of at least 2n:n with at least one gene in every species (solid purple). All above ratios are (named species):(other species). b Synonymous substitution rate (dS) analysis. Density of dS rates for C. campestris paralogs are shown in purple, and for 1:1 orthologs with I. nil and S. lycopersicum in green and blue, respectively. c Functional enrichment analysis of lost and duplicated genes. MapMan categories for genes that are significantly enriched are depicted using the logarithmic values of their p values. The orange bars represent lost genes that are present in one or more copies in the eight other dicot species shown in a but are lost in C. campestris (ratio 0:1 or more). Similarly, purple bars represent categories significantly enriched for duplicated genes (at least 2n:n)

Fig. 4

Comparison of gene losses between C. campestris and autotrophic plants. a Presence (gray) and absence (red) of 42 lost genes connected to photosynthesis, metabolism, transport processes, and symbiotic interactions in 19 sequenced genomes (from Phytozome v.12.0) of the angiosperms, lower plants, and green algae. Species names are shown on the right in the order of increasing evolutionary distance from C. campestris (from top to bottom). Gene abbreviations and MapMan bincodes are given on the top. Full gene names and descriptions of the bincodes are given in Supplementary Table 5. Asterisks behind species names indicate that the sequences from these species were used as query for manual homology searches against the C. campestris genome. b Schematic view of primary carbon metabolism, i.e., Calvin cycle, photorespiration, plastid and cytosolic glycolysis and gluconeogenesis, and synthesis of the compatible solutes mannitol, raffinose and stachyose. Enzymatic reactions for which genes are missing are indicated by circled red crosses. E4P erythrose-4-phosphate, F6P fructose-6-phosphate, FBP fructose-1,6-bisphosphate, G1P glucose-1-phosphate, G6P glucose-6-phosphate, Glyc glycerate, Gly glycine, Gl glycolate, Gl2P glycolate-2-phosphate, Glyox glyoxylate, HyPyr hydroxypyruvate, Ino inositol, M6P mannose-6-phosphate, PEP phosphoenolpyruvate, PGA 3-phosphoglycerate, BPGA 1,3-bisphosphoglycerate, Pyr pyruvate, Raf raffinose, R5P ribose-5-phosphate, Ru5P ribulose-5-phosphate, RuBP ribulose-1,5-bisphosphate, S7P sedulose-7-phosphate, SBP sedulose-1,7-bisphosphate, Ser serine, Sta stachyose, Suc sucrose, TrioseP triose phosphates, UDP-Gal UDP-galactose, Xu5P xylulose-5-phosphate. c Schematic view of photosynthetic electron transport, chlorophyll regeneration, and degradation pathways and phylloquinone synthesis. Enzymatic reactions for which genes are missing are indicated by circled red crosses. Chl a chlorophyll a, Chlide a Chlorophyllide a, Cytb₆f cytochrome b₆f complex, D1 D1 protein of photosystem II, D1* damaged D1 protein, DePhyQ demethylphylloquinone, e⁻ electron, Fd ferredoxin, Glu glutamate, NDH NADH-dehydrogenase complex, PC plastocyanin, PGR5/PGRL1 proton gradient regulation 5/PGR5-like photosynthetic phenotype 1-complex, Pheide a pheophorbide a, Pheo a pheophytin a, PhyQ phylloquinone, PpIX Protoporphyrin IX, PQ plastoquinone, PS I photosystem I, PS II photosystem II, PTOX plastid terminal oxidase

Fig. 5

Horizontal gene transfer (HGT) in C. campestris. a Depiction of the phylogenetic origin of 36 HGT events and their functional category assignment. Phylogenetic orders from which HGT originated are color-coded according to their evolutionary distance to Cuscuta using the median values in million years (MYA) for the split as predicted by Timetree (http://timetree.org) and supported by a species tree calculated using Astral-II (Supplementary Fig. 5). White numbers refer to the number of genes per donor order, black numbers to the right indicate the number of genes in the recipient, C. campestris. Cake diagrams represent functional assignment of donor genes in MapMan bins. b Phylogenetic placement of HGT candidate protein Cc027215, a G-type lectin S-receptor-like serine/threonine-protein kinase, Callus Expression of RBCS 101 (CES101). The amino acid sequence of Cc027215 (highlighted in yellow) is nested deeply within a cluster of orthologous proteins from the hypothetical donor order Fabales (in red), while all orthologs from Solanales species (in gray), that are much more closely related to C. campestris, are located on the second branch. The schematic sequence comparison shows identities relative to Cc027215 in black boxes, discrepancies in gray boxes and gaps in the alignment as black lines. Bootstrap values after 1000 replicates were 86% or higher for all nodes. c Nucleotide sequence comparison of a 36 kb region of chromosome 3 from Daucus carota (top) and a region of C. campestris scaffold123. Coding sequences are shown in dark green and introns as well as up- and down-stream untranslated flanking regions are shown in light green. Areas with very high similarity, indicating conserved remnants of a previous horizontal DNA transfer between both species are indicated by gray fields. Within the depicted area, a gene duplication (Cc021601 and Cc021599 both mapping with all three exons and two introns to DCAR_012700), a larger sequence insertion between Cc021599 and Cc021598, as well as sequence inversions can be seen