The organic anion transporters OAT1 (SLC22A6) and OAT3 (SLC22A8) are drug transporters that are expressed in the kidney, with well-established roles in the in vivo transport of drugs and endogenous metabolites. A comparatively unexplored potential function of these drug transporters is their contribution to the in vivo regulation of natural products (NPs) and their effects on endogenous metabolism. This is important for the evaluation of potential NP interactions with other compounds at the transporter site. Here, we have analyzed the NPs present in several well-established databases from Asian (Chinese, Indian Ayurvedic) and other traditions. Loss of OAT1 and OAT3 in murine knockouts caused serum alterations of many NPs, including flavonoids, vitamins, and indoles. OAT1- and OAT3-dependent NPs were largely separable based on a multivariate analysis of chemical properties. Direct binding to the transporter was confirmed using in vitro transport assays and protein binding assays. Our in vivo and in vitro results, considered in the context of previous data, demonstrate that OAT1 and OAT3 play a pivotal role in the handling of non-synthetic small molecule natural products, NP-derived antioxidants, phytochemicals, and nutrients (e.g., pantothenic acid, thiamine). As described by remote sensing and signaling theory, drug transporters help regulate redox states by meditating the movement of endogenous antioxidants and nutrients between organs and organisms. Our results demonstrate how dietary antioxidants and other NPs might feed into these inter-organ and inter-organismal pathways.

The threat to public health posed by drug-resistant bacteria is rapidly increasing, as some of healthcare’s most potent antibiotics are becoming obsolete. Approximately two-thirds of the world’s antibiotics are derived from natural products produced by Streptomyces encoded biosynthetic gene clusters. Thus, in order to identify novel gene clusters, we sequenced the genomes of four bioactive Streptomyces strains isolated from the soil in San Diego County and used Bacterial Cytological Profiling adapted for agar plate culturing in order to examine the mechanisms of bacterial inhibition exhibited by these strains. In the four strains, we identified 101 biosynthetic gene clusters. Some of these clusters were predicted to produce previously studied antibiotics; however, the known mechanisms of these molecules could not fully account for the antibacterial activity exhibited by the strains, suggesting that novel clusters might encode antibiotics. When assessed for their ability to inhibit the growth of clinically isolated pathogens, three Streptomyces strains demonstrated activity against methicillin-resistant Staphylococcus aureus . Additionally, due to the utility of bacteriophages for genetically manipulating bacterial strains via transduction, we also isolated four new phages (BartholomewSD, IceWarrior, Shawty, and TrvxScott) against S. platensis . A genomic analysis of our phages revealed nearly 200 uncharacterized proteins, including a new site-specific serine integrase that could prove to be a useful genetic tool. Sequence analysis of the Streptomyces strains identified CRISPR-Cas systems and specific spacer sequences that allowed us to predict phage host ranges. Ultimately, this study identified Streptomyces strains with the potential to produce novel chemical matter as well as integrase-encoding phages that could potentially be used to manipulate these strains.

Short-chain fatty acids (SCFAs) and medium-chain fatty acids (MCFAs) include small organic anions derived from the gut microbiome that interact with organic anion transporters of the SLC22 family, many of which are expressed in the kidney proximal tubule. According to the Remote Sensing and Signaling Theory (RSST), crosstalk between organs (e.g., gut–liver–kidney axis, gut–brain axis) and the gut microbiome is mediated by metabolites and signaling molecules transported by multi-specific “drug” transporters. The renal drug transporter OAT1 (SLC22A6) is also a major transporter of gut-microbiome products and uremic toxins (e.g., indoxyl sulfate); it has been shown to act as part of a regulatory feedback loop involving the gut microbiome. SCFAs, especially propionate and butyrate, have been shown to play a central role in the transcriptional regulation of OAT1 through HDAC inhibition. By fecal metagenomics analyses of Oat1 knockout mice, we now find that propionate synthesis is among the most altered pathways in the gut microbiome. In contrast, these pathways were only minimally altered in the Oat3 (Slc22a8) knockout. Metabolomics analyses indicate that serum propionate derivatives (e.g., propionyl glycine) and 3-hydroxybutyrate are dependent on OAT1 in the knockout mice and in humans treated with probenecid, an OAT1 inhibitor. The gut microbiome of the Oat1 knockout mice also exhibited greater fatty acid synthesis, which generates odd-chain-length fatty acids (e.g. heptanoate) when propionate is available. Overall, the data, especially when considered in light of in vitro experiments of others, indicates the in vivo existence of a feedback loop connecting gut-microbiome-derived SCFAs and MCFAs to kidney proximal tubule uptake via OAT1. This bidirectional feedback loop in turn regulates OAT1 expression through HDAC inhibition. The feedback loop is clearly consistent with the Remote Sensing and Signaling Theory—in particular, the centrality of multi-specific “drug” transporters in organ crosstalk and host–microbiome interactions via small molecules with “high information content.” The key role of OAT1 function in maintaining tubular secretion in CKD supports the importance of this RSST loop in renal pathophysiology. Modulating this RSST loop could have therapeutic value in chronic kidney disease and other contexts.

Uridine 5′-diphospho-glucuronosyltransferase (UGT) enzymes play a critical role in hepatic and other metabolism of endogenous and exogenous small molecules. The addition of glucuronic acid via UGT increases the polarity and subsequent solubility of small molecules, helping direct them for biliary excretion and renal excretion by organic anion transporter OAT3 (SLC22A8). Mutations in UGTs are associated with disorders in biliary metabolism (e.g., Gilbert’s syndrome, Crigler-Najjar syndrome). Here, we used comprehensive serum metabolomic profiling of pan Ugt1-knockout (Ugt1–/–) mice to define the metabolic consequences of UGT deficiency. All 7 knockout mice exhibited markedly elevated bilirubin (mean = 5.96 mg/dL). Partial Least Squares Discriminant Analysis (PLS-DA) showed clear separation between Ugt1–/– and wild type (WT), and volcano plots revealed significant endogenous metabolite alterations. These included upregulation of numerous bilirubin degradation products as well as major disturbances in lipid and amino acid metabolism. In particular, pathway enrichment analyses identified disruptions in fatty acid biosynthesis, glycine-serine metabolism, heme breakdown, porphyrin metabolism, and β-oxidation of fatty acids. Thus, the UGT1 proteins regulate different aspects of endogenous metabolism and signaling. Consistent with this notion, there were increases in liver expression of several nuclear receptors (e.g., HNF4a, PPARa, PXR) involved in metabolic signaling. These results support the remote sensing and signaling theory (RSST), which proposes that UGT1 proteins play a key role in organ crosstalk via routing small molecules between the liver and kidney, with glucuronidated molecules being mainly directed to the multispecific SLC kidney transporter OAT3. We propose that this glucuronidation-regulated RSST loop is essential to signaling along the gut-liver-kidney axis.

Organic anion transporters (OATs, SLC22) in the kidney and organic anion-transporting polypeptides (OATPs, SLCO) in the liver play crucial roles in the disposition of small molecule drugs that are organic anions. According to the Remote Sensing and Signaling Theory, these multispecific “drug” transporters are also central to crosstalk between the liver, kidney, and other organs via endogenous small molecules (e.g., metabolites, signaling molecules, gut microbiome products). These multispecific drug transporters govern access of small molecules with high informational content across multiple scales (organism to organelle). Previous chemoinformatic and machine learning methods have proven useful for identifying molecular properties of organic anion drugs that predispose them to handling by the OAT (renal) and the OATP (hepatic) transporters. This is important for understanding pharmacokinetics (ADME) in the context of chronic kidney disease (CKD) and liver disease. Given that OATs and OATPs are involved in many metabolic diseases, we sought to determine whether molecular properties could be identified for distinguishing OAT- versus OATP-interacting endogenous metabolites in vivo. This is essential for understanding endogenous small molecule communication between the kidney proximal tubule and hepatocytes in a larger Remote Sensing and Signaling System. We analyzed in vivo metabolomics data from OAT and OATP knockout mice, focusing on endogenous metabolites selective for OATs (e.g., OAT1 or SLC22A6; OAT3 or SLC22A8) vs OATPs (including the locus containing Oatp1b2, the closest homologue of human OATP1B1 or SLCO1B1 and OATP1B3 or SLCO1B3). Applying chemoinformatic methods to a data set of 210 metabolites based on knockout mouse metabolomics (92 OAT-selective, 118 OATP-selective), we identified a set of distinguishing molecular properties (e.g., MolLogP, RingCount, NumRotatableBonds). We then used machine learning approaches (e.g., Random Forest, Naive Bayes, Logistic Regression) to classify OAT vs OATP metabolites, achieving over 75% accuracy. These results support the view that transporter knockout mouse metabolomics can help define selectivity of SLC drug transporters for endogenous metabolites, signaling molecules, antioxidants, nutrients, and gut microbiome products. In the context of the Remote Sensing and Signaling Theory, we discuss the implications for understanding organ crosstalk and interorganismal communication as well as drug disposition, drug-metabolite interactions, and metabolite-based drug design.