Pangenomics analysis reveals diversification of enzyme families and niche specialization in globally abundant SAR202 bacteria

© The Author(s), 2019. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Saw, J. H. W., Nunoura, T., Hirai, M., Takaki, Y., Parsons, R., Michelsen, M., Longnecker, K., Kujawinski, E. B., Stepanauskas, R., Landry, Z., Carlson, C. A., & Giovannoni, S. J. Pangenomics analysis reveals diversification of enzyme families and niche specialization in globally abundant SAR202 bacteria. Mbio, 11(1), (2020): e02975-19, doi:10.1128/mBio.02975-19.It has been hypothesized that the abundant heterotrophic ocean bacterioplankton in the SAR202 clade of the phylum Chloroflexi evolved specialized metabolisms for the oxidation of organic compounds that are resistant to microbial degradation via common metabolic pathways. Expansions of paralogous enzymes were reported and implicated in hypothetical metabolism involving monooxygenase and dioxygenase enzymes. In the proposed metabolic schemes, the paralogs serve the purpose of diversifying the range of organic molecules that cells can utilize. To further explore SAR202 evolution and metabolism, we reconstructed single amplified genomes and metagenome-assembled genomes from locations around the world that included the deepest ocean trenches. In an analysis of 122 SAR202 genomes that included seven subclades spanning SAR202 diversity, we observed additional evidence of paralog expansions that correlated with evolutionary history, as well as further evidence of metabolic specialization. Consistent with previous reports, families of flavin-dependent monooxygenases were observed mainly in the group III SAR202 genomes, and expansions of dioxygenase enzymes were prevalent in those of group VII. We found that group I SAR202 genomes encode expansions of racemases in the enolase superfamily, which we propose evolved for the degradation of compounds that resist biological oxidation because of chiral complexity. Supporting the conclusion that the paralog expansions indicate metabolic specialization, fragment recruitment and fluorescent in situ hybridization (FISH) with phylogenetic probes showed that SAR202 subclades are indigenous to different ocean depths and geographical regions. Surprisingly, some of the subclades were abundant in surface waters and contained rhodopsin genes, altering our understanding of the ecological role of SAR202 species in stratified water columns. IMPORTANCE The oceans contain an estimated 662 Pg C in the form of dissolved organic matter (DOM). Information about microbial interactions with this vast resource is limited, despite broad recognition that DOM turnover has a major impact on the global carbon cycle. To explain patterns in the genomes of marine bacteria, we propose hypothetical metabolic pathways for the oxidation of organic molecules that are resistant to oxidation via common pathways. The hypothetical schemes we propose suggest new metabolic pathways and classes of compounds that could be important for understanding the distribution of organic carbon throughout the biosphere. These genome-based schemes will remain hypothetical until evidence from experimental cell biology can be gathered to test them. Our findings also fundamentally change our understanding of the ecology of SAR202 bacteria, showing that metabolically diverse variants of these cells occupy niches spanning all depths and are not relegated to the dark ocean.We thank the captain, crew, ROV and CTD operation teams, and science party of the JAMSTEC RV Kairei cruises KR11-11, KR12-19, and KR14-01. We thank the staff of the Bigelow Laboratory for Ocean Sciences’ Single Cell Genomics Center for the generation of single-cell genomic data. We thank Mark Dasenko from the Center for Genome Research and Biocomputing at Oregon State University for sequencing six of the Illumina SAG libraries. We thank the captain, crew and CTD operations team of the RV Atlantic Explorer (cruise AE1712). T.N. was supported in part by a Grant-in-Aid for Scientific Research (B) (30070015) from the Japan Society for the Promotion of Science (JSPS). The mass spectrometry samples were analyzed at the WHOI FT-MS Users’ Facility; funding for data collection and analysis came from the National Science Foundation (NSF Grant OCE-1154320 to E.B.K. and K.L.). This work was funded by Simons Foundation International as part of the BIOS-SCOPE initiative (S.J.G., C.A.C., and E.B.K.), and by NSF grants OCE-1335810 and DEB-1441717 to R.S. This work was funded by National Science Foundation grant OCE-1436865