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The bioactive lysophospholipid sphingosine-1-phosphate (S1P) acts via five different subtypes of S1P receptors (S1PRs) - S1P1-5. S1P5 is predominantly expressed in nervous and immune systems, regulating the egress of natural killer cells from lymph nodes and playing a role in immune and neurodegenerative disorders, as well as carcinogenesis. Several S1PR therapeutic drugs have been developed to treat these diseases; however, they lack receptor subtype selectivity, which leads to side effects. In this article, we describe a 2.2 Å resolution room temperature crystal structure of the human S1P5 receptor in complex with a selective inverse agonist determined by serial femtosecond crystallography (SFX) at the Pohang Accelerator Laboratory X-Ray Free Electron Laser (PAL-XFEL) and analyze its structure-activity relationship data. The structure demonstrates a unique ligand-binding mode, involving an allosteric sub-pocket, which clarifies the receptor subtype selectivity and provides a template for structure-based drug design. Together with previously published S1PR structures in complex with antagonists and agonists, our structure with S1P5-inverse agonist sheds light on the activation mechanism and reveals structural determinants of the inverse agonism in the S1PR family. S1P5 is a sphingosine-1-phosphate (S1P) receptor implicated in immune and neurodegenerative disorders. Here, authors report a crystal structure of the S1P5 receptor in complex with a selective inverse agonist, revealing an allosteric subpocket and shedding light on inverse agonism in S1P receptors. [ABSTRACT FROM AUTHOR] more...

Venoms are a rich source of highly specific toxins, which allow the identification of novel therapeutic targets. We have now applied high content screening (HCS) microscopy to identify toxins that modulate pain sensitization signaling in primary sensory neurons of rat and elucidated the underlying mechanism. A set of venoms and fractions thereof were analyzed for their ability to activate type II protein kinase A (PKA‐II) and extracellular signal‐regulated kinases (ERK1/2). We identified MeuNaTxα‐1, a sodium channel‐selective scorpion α‐toxin from Mesobuthus eupeus, which affected both PKA‐II and ERK1/2. Recombinant MeuNaTxα‐1 showed identical activity to the native toxin on mammalian voltage‐gated sodium channels expressed in Xenopus laevis oocytes and induced thermal hyperalgesia in adult mice. The effect of MeuNaTxα‐1 on sensory neurons was dose‐dependent and tetrodotoxin‐sensitive. Application of inhibitors and toxin mutants with altered sodium channel selectivity demonstrated that signaling activation in sensory neurons depends on NaV1.2 isoform. Accordingly, the toxin was more potent in neurons from newborn rats, where NaV1.2 is expressed at a higher level. Our results demonstrate that HCS microscopy‐based monitoring of intracellular signaling is a novel and powerful tool to identify and characterize venoms and their toxins affecting sensory neurons. [ABSTRACT FROM AUTHOR] more...