Predicting and Manipulating Cardiac Drug Inactivation by the Human Gut Bacterium Eggerthella lenta

Despite numerous examples of the effects of the human gastrointestinal microbiome on drug efficacy and toxicity, there is often an incomplete understanding of the underlying mechanisms. Here, we dissect the inactivation of the cardiac drug digoxin by the gut Actinobacterium Eggerthella lenta. Transcriptional profiling, comparative genomics, and culture-based assays revealed a cytochrome-encoding operon up-regulated by digoxin, inhibited by arginine, absent in nonmetabolizing E. lenta strains, and predictive of digoxin inactivation by the human gut microbiome. Pharmacokinetic studies using gnotobiotic mice revealed that dietary protein reduces the in vivo microbial metabolism of digoxin, with significant changes to drug concentration in the serum and urine. These results emphasize the importance of viewing pharmacology from the perspective of both our human and microbial genomes.Chemistry and Chemical Biolog

Small non-coding RNAs in Streptomyces coelicolor

In bacteria, small RNAs (sRNAs) make important regulatory contributions to an ever increasing number of cellular processes. To expand the repertoire of known sRNAs, we sought to identify novel sRNAs in the differentiating, multicellular bacterium Streptomyces coelicolor. We describe a combined bioinformatic and experimental approach that enabled the identification and characterization of nine novel sRNAs in S. coelicolor, including a cis-encoded antisense sRNA. We examined sRNA expression throughout the S. coelicolor developmental cycle, which progresses from vegetative mycelium formation, to aerial mycelium formation and finally sporulation. We further determined the effects of growth medium composition (rich versus minimal medium) on sRNA gene expression, and compared wild-type sRNA expression profiles with those of four developmental mutants. All but two of the sRNAs exhibited some degree of medium dependence, with three sRNAs being expressed exclusively during growth on one medium type. Unlike most sRNAs characterized thus far, several sRNA genes in S. coelicolor were expressed constitutively (apart from during late sporulation), suggesting a possible housekeeping role for these transcripts. Others were expressed at specific developmental stages, and their expression profiles were altered in response to developmental mutations. Expression of one sRNA in particular was dependent upon the sporulation-specific sigma factor σWhiG

Developmentally regulated cleavage of tRNAs in the bacterium Streptomyces coelicolor

The ability to sense and respond to environmental and physiological signals is critical for the survival of the soil-dwelling Gram-positive bacterium Streptomyces coelicolor. Nutrient deprivation triggers the onset of a complex morphological differentiation process that involves the raising of aerial hyphae and formation of spore chains, and coincides with the production of a diverse array of clinically relevant antibiotics and other secondary metabolites. These processes are tightly regulated; however, the genes and signals involved have not been fully elucidated. Here, we report a novel tRNA cleavage event that follows the same temporal regulation as morphological and physiological differentiation, and is growth medium dependent. All tRNAs appear to be susceptible to cleavage; however, there appears to be a bias towards increased cleavage of those tRNAs that specify highly utilized codons. In contrast to what has been observed in eukaryotes, accumulation of tRNA halves in S. coelicolor is not significantly affected by amino acid starvation, and is also not affected by induction of the stringent response or inhibition of ribosome function. Mutants defective in aerial development and antibiotic production exhibit altered tRNA cleavage profiles relative to wild-type strains

Genomic variation and strain-specific functional adaptation in the human gut microbiome during early life

The human gut microbiome matures towards the adult composition during the first years of life and is implicated in early immune development. Here, we investigate the effects of microbial genomic diversity on gut microbiome development using integrated early childhood data sets collected in the DIABIMMUNE study in Finland, Estonia and Russian Karelia. We show that gut microbial diversity is associated with household location and linear growth of children. Single nucleotide polymorphism- and metagenomic assembly-based strain tracking revealed large and highly dynamic microbial pangenomes, especially in the genus Bacteroides, in which we identified evidence of variability deriving from Bacteroides-targeting bacteriophages. Our analyses revealed functional consequences of strain diversity; only 10% of Finnish infants harboured Bifidobacterium longum subsp. infantis, a subspecies specialized in human milk metabolism, whereas Russian infants commonly maintained a probiotic Bifidobacterium bifidum strain in infancy. Groups of bacteria contributing to diverse, characterized metabolic pathways converged to highly subject-specific configurations over the first two years of life. This longitudinal study extends the current view of early gut microbial community assembly based on strain-level genomic variation.Peer reviewe

Developing a metagenomic view of xenobiotic metabolism

The microbes residing in and on the human body influence human physiology in many ways, particularly through their impact on the metabolism of xenobiotic compounds, including therapeutic drugs, antibiotics, and diet-derived bioactive compounds. Despite the importance of these interactions and the many possibilities for intervention, microbial xenobiotic metabolism remains a largely underexplored component of pharmacology. Here, we discuss the emerging evidence for both direct and indirect effects of the human gut microbiota on xenobiotic metabolism, and the initial links that have been made between specific compounds, diverse members of this complex community, and the microbial genes responsible. Furthermore, we highlight the many parallels to the now well-established field of environmental bioremediation, and the vast potential to leverage emerging metagenomic tools to shed new light on these important microbial biotransformations

Cell Wall Hydrolases Affect Germination, Vegetative Growth, and Sporulation in Streptomyces coelicolor▿ †

Peptidoglycan is a major cell wall constituent of gram-positive bacteria. It is a dynamic macromolecule that is actively remodeled to enable cell growth and differentiation through a tightly choreographed interplay of hydrolytic and biosynthetic enzyme activities. The filamentous bacterium Streptomyces coelicolor has a complex life cycle that likely requires considerable cell wall remodeling to enable both extension of vegetative hyphae and formation of differentiated cell types. In silico analysis of the S. coelicolor genome enabled identification of 56 candidate cell wall hydrolase genes. We found that seven of these genes shared a highly conserved 5′ untranslated region and were expressed during both vegetative growth and sporulation; four of these genes were selected for more extensive biochemical and biological characterization. The proteins encoded by these genes, termed RpfA, SwlA, SwlB, and SwlC, were confirmed to be hydrolytic enzymes, as they could efficiently cleave S. coelicolor cell walls. Phenotypic analyses revealed that these enzymes are important throughout development; deletion of each hydrolase gene resulted in a mutant strain that was heat sensitive, defective in spore formation, and either altered in vegetative growth or delayed in spore germination. Our results indicate that these enzymes play key roles at multiple stages in the growth and development of S. coelicolor, highlighting both the lack of redundancy in hydrolase activity and the importance of cell wall remodeling in the S. coelicolor life cycle

Mechanistic insight into digoxin inactivation by Eggerthella lenta augments our understanding of its pharmacokinetics

The human gut microbiota plays a key role in pharmacology, yet the mechanisms responsible remain unclear, impeding efforts toward personalized medicine. We recently identified a cytochrome-encoding operon in the common gut Actinobacterium Eggerthella lenta that is transcriptionally activated by the cardiac drug digoxin. These genes represent a predictive microbial biomarker for the inactivation of digoxin. Gnotobiotic mouse experiments revealed that increased protein intake can limit microbial drug inactivation. Here, we present a biochemical rationale for how the proteins encoded by this operon might inactivate digoxin through substrate promiscuity. We discuss digoxin signaling in eukaryotic systems, and consider the possibility that endogenous digoxin-like molecules may have selected for microbial digoxin inactivation. Finally, we highlight the diverse contributions of gut microbes to drug metabolism, present a generalized approach to studying microbe-drug interactions, and argue that mechanistic studies will pave the way for the clinical application of this work