Hansen, J. R. | bioRxiv Science

bioRxiv2018

Lactobacillus acidophilus disrupts collaborative multispecies bile acid metabolism

Bile acids are metabolic links between hosts and their gut microbiomes, yet little is known about the roles they play in microbe-to-microbe interactions. Here we present a study designed to investigate the effect that a common probiotic, Lactobacillus acidophilus, has on microbial interactions that lead to formation of secondary bile acids. A model microbial consortium was built from three human gut isolates, Clostridium scindens, Collinsella aerofaciens, and Blautia obeum, and cultured under different bile acid and probiotic treatments. A multi-omics platform that included mass spectrometry-based metabolomics and activity-based proteomic probes was used to produce two major results. The first, was that an uncommon secondary bile acid - ursocholate - was produced by a multi-species chemical synthesis pathway. This result highlights a new microbe-to-microbe interaction mediated by bile acids. The second finding was that the probiotic strain, L. acidophilus, quenched the observed interactions and effectively halted consortial synthesis of ursocholate. Little is known about the role that ursocholate plays in human health and development. However, we did discover that a decrease in ursocholate abundance corresponded with successful weight loss in patients after gastric bypass surgery versus those who did not lose weight after surgery. Hence, this study uncovered basic knowledge that may aid future designs of custom probiotic therapies to combat obesity.

microbiology↗

bioRxiv2018

Proximity-dependent proteomics of the Chlamydia trachomatis inclusion membrane reveals functional interactions with endoplasmic reticulum exit sites

Chlamydia trachomatis is the most common bacterial sexually transmitted infection, responsible for millions of infections each year. Despite this high prevalence, the elucidation of the molecular mechanisms of Chlamydia pathogenesis has been difficult due to limitations in genetic tools and its intracellular developmental cycle. Within a host epithelial cell, chlamydiae replicate within a vacuole called the inclusion. Many Chlamydia-host interactions are thought to be mediated by the Inc family of type III secreted proteins that are anchored in the inclusion membrane, but their array of host targets are largely unknown. To investigate how the inclusion membrane proteome changes over the course of an infected cell, we have adapted the APEX system of proximity-dependent biotinylation. APEX is capable of specifically labeling proteins within a 20 nm radius in living cells. We transformed C. trachomatis to express the enzyme APEX fused to known inclusion membrane proteins, allowing biotinylation and pull-down of inclusion-associated proteins. Using quantitative mass spectrometry against APEX labeled samples, we identified over 400 proteins associated with the inclusion membrane at early, middle, and late stages of epithelial cell infection. This system was sensitive enough to detect inclusion interacting proteins early in the developmental cycle, at 8 hours post infection, a previously intractable time point. Mass spectrometry analysis revealed a novel, early association between C. trachomatis inclusions and endoplasmic reticulum exit sites (ERES), functional regions of the ER where COPII-coated vesicles originate. Pharmacological and genetic disruption of ERES function severely restricted early chlamydial growth and the development of infectious progeny. APEX is therefore a powerful in situ approach for identifying critical protein interactions on the membranes of pathogen-containing vacuoles. Furthermore, the data derived from proteomic mapping of Chlamydia inclusions has illuminated an important functional role for ERES in promoting chlamydial developmental growth.

microbiology↗