Function and Importance of Marine Bacterial Transporters of Plankton Exometabolites

Abstract

Metabolite exchange within marine microbial communities transfers carbon and other major elements through global cycles and forms the basis of microbial interactions. Yet lack of gene annotations and concern about the quality of existing ones remain major impediments to revealing the metabolite-microbial network. We employed an arrayed mutant library of the marine bacterium Ruegeria pomeroyi DSS-3 to experimentally annotate substrates of organic compound transporter systems, using mutant growth and compound drawdown analyses to link transporters to their substrates. Mutant experiments verified substrates for thirteen R. pomeroyi transporters. Four were previously hypothesized based on gene expression data (taurine, glucose/xylose, isethionate, and cadaverine/putrescine/spermidine); five were previously hypothesized based on homology to experimentally annotated transporters in other bacteria (citrate, glycerol, N-acetylglucosamine, fumarate/malate/succinate, and dimethylsulfoniopropionate); and four had no previous annotations (thymidine, carnitine, cysteate, and 3-hydroxybutyrate transporter). These bring the total number of experimentally-verified organic carbon influx transporters to 17 of 126 in the R. pomeroyi genome. In a longitudinal study of a coastal phytoplankton bloom, expression patterns of the experimentally annotated transporters linked them to different stages of the bloom, and also led to the hypothesis that citrate and 3-hydroxybutyrate were among the most highly available bacterial substrates. Improved functional knowledge of these gatekeepers of organic carbon uptake is facilitating better characterization of the surface ocean metabolite network.

Digital Microbe: A Genome-Informed Data Integration Framework for Collaborative Research on Emerging Model Organisms

“…This figure was generated from version 5.0 of the R. pomeroyi Digital Microbe databases on Zenodo. (Schroer et al 2023) which were added to the Digital Microbe (required feature 2). Previous homology-based annotations of most of these transporter systems were either incorrect or vague, and therefore of minimal ecological value.…”

“…Introduced into a natural dinoflagellate bloom (Nowinski and Moran 2021), the Arrows point to transcriptomes collected when substrates were derived from dinoflagellates (red) or diatoms (brown); significant differences are indicated with asterisks colored red (enriched with citrate transporter had the highest relative expression; in a diatom co-culture, the acetate transporter was the most highly expressed; co-cultured with a green alga, transporter genes indicated that taurine, glycerol, carnitine, and dimethylsulfoniopropionate (DMSP) were on the menu. The organic acid transporter that enables R. pomeroyi uptake of 3-hydroxybutyrate (Schroer et al 2023) was expressed across most growth conditions, yet this metabolite has not previously been identified as a relevant currency in bacterially-driven carbon flux. The metaanalysis also showed a pattern in expression for transporters that contain a substrate binding protein gene (i.e., the ABC and TRAP transporter classes): the gene is expressed at consistently higher levels than other genes in the same transporter (i.e., higher than permeases and ATP-binding proteins) despite all having membership in the same operon.…”

“…Three additional genome-linked data types recently added to the R. pomeroyi Digital Microbe include the locations of insertion sites of knockout mutants (covering 3,570 genes of 4,288 genes; (Schroer et al 2023;Mejia et al 2022), proteomic data collected concurrently with one of the transcriptomic studies (Cooper et al, unpublished), and TnSeq mutant fitness measurements in synthetic microbial communities (Schreier et al 2023) (required feature 2), and these are enhancing collaborations among team members.…”

Substrate uptake patterns shape niche separation in marine prokaryotic microbiome

“…Therefore, POM and high-molecular weight DOM need to be hydrolyzed before assimilation (5). In addition, the substrate specificity of membrane-bound transporter protein systems determines substrate uptake even if the size of the molecule is theoretically small enough to be transferred into the cell (6)(7)(8)(9)(10)(11)(12)(13).…”

Ultrahigh-affinity transport proteins from ubiquitous marine bacteria reveal mechanisms and global patterns of nutrient uptake

“…However, the properties of these transporters and their specific functions (i.e., transported metabolites) are mostly unknown, limiting our knowledge of the full range of DOM that can be assimilated by SAR11 bacteria, nutrient exchange within the marine microbial community, and the molecular mechanisms for high-affinity nutrient uptake. Although homology-based predictions are available, homology-based predictions of protein function have limited accuracy, especially for functionally diverse protein superfamilies like ABC transporters 18,19 . Transport proteins can also be characterized experimentally through radioassays of nutrient uptake in cultured cells, and this approach has been used to characterize the broad-specificity osmolyte transporter of the SAR11 bacterium ‘ Candidatus Pelagibacter ubique’ 20 .…”