Biology subjects

Sidebottom, A. M.

Publications and source records attributed to Sidebottom, A. M..

3 recordsLinked to original sources

bioRxiv2022

Exceptionally versatile respiratory metabolisms drive metabolite production by diverse gut bacteria

Respiratory reductases enable microbes to utilize molecules present in anaerobic ecosystems as energy-generating respiratory electron acceptors. Here we identify three taxonomically distinct families of human gut bacteria (Burkholderiaceae, Eggerthellaceae, Erysipelotrichaceae) that encode large arsenals of tens-to-hundreds of respiratory-like reductases per genome. Screening species from each family (Sutterella wadsworthensis, Eggerthella lenta, and Holdemania filiformis), we discover 22 metabolites used as respiratory electron acceptors in a species-specific manner. Identified reactions transform multiple classes of dietary- and host-derived metabolites, including bioactive molecules resveratrol and itaconate. Products of identified respiratory metabolisms highlight poorly characterized compounds, such as the itaconate-derived 2-methylsuccinate. Reductase substrate-profiling defines enzyme-substrate pairs and reveals a complex picture of reductase evolution, providing evidence that reductases with specificities for related cinnamate substrates independently emerged at least four times. These studies thus establish an exceptionally versatile form of anaerobic respiration that directly links microbial energy metabolism to the gut metabolome.

microbiology↗

bioRxiv2022

Distinct energy-coupling factor transporter subunits enable flavin acquisition and extracytosolic trafficking for extracellular electron transfer in Listeria monocytogenes

A variety of electron transfer mechanisms link bacterial cytosolic electron pools with functionally diverse redox activities in the cell envelope and extracellular space. In Listeria monocytogenes, the ApbE-like enzyme FmnB catalyzes extracytosolic protein flavinylation, covalently linking a flavin cofactor to proteins that transfer electrons to extracellular acceptors. L. monocytogenes uses an energy-coupling factor (ECF) transporter complex that contains distinct substrate-binding, transmembrane, ATPase A, and ATPase A subunits (RibU, EcfT, EcfA, and EcfA) to import environmental flavins, but the basis of extracytosolic flavin trafficking for FmnB flavinylation remains poorly defined. In this study, we show that the proteins EetB and FmnA are related to ECF transporter substrate-binding and transmembrane subunits, respectively, and are essential for exporting flavins from the cytosol for flavinylation. Comparisons of the flavin import versus export capabilities of L. monocytogenes strains lacking different ECF transporter subunits demonstrates a strict directionality of substrate-binding subunit transport but partial functional redundancy of transmembrane and ATPase subunits. Based on these results, we propose that ECF transporter complexes with different subunit compositions execute directional flavin import/export through a broadly conserved mechanism. Finally, we present genomic context analyses which show that related ECF exporter genes are distributed across the Firmicutes phylum and frequently co-localize with genes encoding flavinylated extracytosolic proteins. These findings clarify the basis of ECF transporter export and extracytosolic flavin cofactor trafficking in Firmicutes.

microbiology↗

bioRxiv2022

Dynamic genetic adaptation of Bacteroides thetaiotaomicron murine gut colonization

To understand how a bacterium ultimately succeeds or fails in adapting to a new environment, it is essential to assess the temporal dynamics of its fitness over the course of colonization. The mammalian gut, into which exogenous microorganisms are regularly introduced, represents a biologically and clinically relevant system to explore microbial adaptational processes. In this study, we introduce a human-derived commensal organism, Bacteroides thetaiotaomicron, into the guts of germ-free mice to 1) determine whether the genetic requirements for colonization shift over time and, if so, 2) characterize the biological functions required for microbial survival at different points of colonization. The results of a high-throughput functional genetics assay (BarSeq), transcriptomics, and metabolomics converge on several conclusions. First, adaptation to the host gut occurs in distinct stages. We observed drastic changes in gene usage during the first week, shifting from high expression of amino acid biosynthesis to polysaccharide utilization genes. These changes were sustained thereafter, except for the continued upregulation of a single polysaccharide utilization locus responsible for the degradation of raffinose-family oligosaccharides rich in the standard chow diet fed to our mice. Spontaneous mutations in wildtype Bt also evolve around this locus, highlighting the importance of efficient carbohydrate metabolism in long-term persistence within a monoassociated gut. To improve microbiome-based therapies, it will be important to appreciate and meet the distinct needs of the organism during each stage of colonization. ImportanceMicrobes regularly disperse across and adapt to new environments and ecological niches. A clinically significant microbial niche home to trillions of microbes is the mammalian gut. Temporal processes of microbial adaptation over the course of gut colonization are poorly understood on a genetic, transcriptional, and metabolite level. In this study, we leverage a three-pronged approach to characterize gut colonization as a dynamic process with shifting genetic determinants of microbial fitness. This study sheds light on host colonization by Bacteroides thetaiotaomicron, an organism that is prevalent and dominant across healthy human microbiomes, and not only identifies key pathways involved in colonization, but determines the timing of when these pathways are most vital to colonization success. By demonstrating that the key determinants of colonization success in the gut change over time, the results of this study highlight the importance of considering ecological dynamics in developing more effective microbiome-based therapies.

microbiology↗