Dynamic genome evolution and complex virocell metabolism of globally-distributed giant viruses

Abstract

The discovery of eukaryotic giant viruses has transformed our understanding of the limits of viral complexity, but the extent of their encoded metabolic diversity remains unclear. Here we generate 501 metagenome-assembled genomes of Nucleo-Cytoplasmic Large DNA Viruses (NCLDV) from environments around the globe, and analyze their encoded functional capacity. We report a remarkable diversity of metabolic genes in widespread giant viruses, including many involved in nutrient uptake, light harvesting, and nitrogen metabolism. Surprisingly, numerous NCLDV encode the components of glycolysis and the TCA cycle, suggesting that they can re-program fundamental aspects of their host's central carbon metabolism. Our phylogenetic analysis of NCLDV metabolic genes and their cellular homologs reveals distinct clustering of viral sequences into divergent clades, indicating that these genes are virus-specific and were acquired in the distant past. Overall our findings reveal that giant viruses encode complex metabolic capabilities with evolutionary histories largely independent of cellular life, strongly implicating them as important drivers of global biogeochemical cycles.

Fig. 1. Evolutionary relationships of NCLDV MAGs and reference genomes.

a Phylogeny of the 501 NCLDV MAGs presented in this study together with 121 reference genomes. The phylogeny was constructed from a concatenated alignment of 5 highly conserved marker genes that are present throughout the NCLDV families using the VT + F + I + G4 model in IQ-TREE. The tree is rooted at Poxviridae/Asfarviridae branch, consistent with previous studies. The inner strip is colored according to the phylogeny of the MAGs, while the outer strip is colored according to the habitat in which they were found. The bar chart represents genome size, which ranges from 100–2474 Kbp, and the dotted line denotes the 500 Kbp mark. Clades with >5 genomes are indicated with two letter abbreviations and clade numbers. MM: Mimiviridae, EP: Early Phycodnaviridae, LP: Late Phycodnaviridae, IR: Iridoviridae, MR: Marseilleviridae, PT: Pithoviridae. For the list of all the clades, see Dataset 1. b Average amino acid identity (AAI) heatmap of the MAGs and reference genomes, with rows and columns clustered according to the phylogeny.

Fig. 2. Gene-sharing patterns of the NCLDV MAGs and reference genomes.

a The distribution of the orthologous groups (OGs) in the NCLDV MAGs and reference genomes. The barplot on the left shows the proportion of OGs in each frequency category that could be assigned an annotation, while the barplot on the right shows the total number of OGs in each frequency category (log scale). b The degree distribution of the OG occurrence in the genomes analyzed. The best fit to a power law distribution is also shown. c A bipartite network of the OGs, with large nodes corresponding to genomes and small nodes corresponding to OGs. The size of the genome nodes is proportional to their genome size, and they are colored according to their family-level classification.

Fig. 3. Distribution of orthologous groups and metabolic enzymes in NCLDV clades.

a The barplot shows the number of enriched OGs in each of the major NCLDV clades analyzed in this study. Only a subset of total functional categories are shown here; a full table can be found in Supplementary Dataset 2. The heatmap shows the occurrence of OGs with >5 total members across the major NCLDV clades, with shading corresponding to the percent of MAGs in that clade that encode a given OG. b A bubble plot of select metabolic genes detected in the NCLDV clades, with bubble size proportional to the percent of genomes in a clade that encode that protein. The numbers in brackets next to each enzyme name denote the number of these proteins observed in the MAGs we present here and the number observed in reference NCLDV genomes, respectively. G3P: glycerol-3-phosphate; LCM: Large conductance mechanosensitive; SCM; small conductance mechanosensitive.

Fig. 4. Presence of central carbon metabolic enzymes in the NCLDVs.

a Schematic of Glycolysis, Gluconeogenesis, and the TCA cycle, with the number of NCLDV MAGs harboring a particular enzyme provided beside abbreviated enzyme names. Enzymes that were not detected in any of the studied NCLDVs are in gray. b Representative CDS from genome ERX552243.92 illustrating the domain organization (PFAM and Interpro) of the fused-domain gene (G3P + PGK) involved in glycolysis, that was detected in 16 of the NCLDV MAGs. c Example of co-localization of genes involved in TCA cycle on genomic contigs from five representative NCLDV MAGs. Location of a number of other genes commonly present in NCLDVs are also shown. d Presence/absence of genes involved in central-carbon metabolism in NCLDV genomes assembled in this study. Only the genomes harboring 3 or more enzymes are shown. G3P + PGK indicates the fused-domain gene illustrated in panel B. Blue arrow indicates the genome that harbors 7 out of 10 enzymes involved in glycolysis. HK: hexokinase, PGI: Phosphoglucoisomerase, PFK: Phosphofructokinase, ALD: aldolase, TPI: Triose-phosphate isomerase, G3P: Glyceraldehyde 3-phosphate dehydrogenase, PGK: Phosphoglycerate kinase, PGM: Phosphoglycerate mutase, ENO: Enolase, PYK: Pyruvate kinase, PEPCK: PEP carboxykinase, FBP: Fructose 1,6-bisphosphatase, G6P: Glucose 6-phosphatase, PDH: Pyruvate dehydrogenase, PC: Pyruvate carboxylase, CS: Citrate synthase, ACON: Aconitase, ICL: Isocitrate lyase, ICD: Isocitrate dehydrogenase, αKDH: α-ketoglutarate dehydrogenase, SCS: Succinyl-CoA synthetase, SD: Succinate dehydrogenase (subunits A, B and C), FH: Fumarate hydratase, MS: Malate synthase, MDH: Malate dehydrogenase.

Fig. 5. Phylogenetic reconstruction of a number of representative NCLDV genes likely involved in carbon and nutrient metabolism and light harvesting.

NCLDV-specific clusters are encircled with dashed ovals in each of the trees, while number of genes from different NCLDV-clades contributing to these monophyletic groups are also provided (MM: Mimiviridae, EP: Early Phycodnaviridae, LP: Late Phycodnaviridae) Colors of the clade names correspond to those in Fig. 1. Although node support values are not provided for better visual clarity, all the NCLDV-specific nodes are supported by >90% ultrafast bootstrap values (see Methods and Data availability statement for details). One asterisk denotes bacteriophage sequences, which are only present in the PhoH tree. Double asterisks denote unclassified sequences (environmental), which are only present in the rhodopsin tree.