Canonical and noncanonical Wnts use a common mechanism to activate completely unrelated coreceptors.

Wnt ligands signal through ß-catenin and are critically involved in cell fate determination and stem/progenitor self-renewal. Wnts also signal through ß-catenin-independent or noncanonical pathways that regulate crucial events during embryonic development. The mechanism of noncanonical receptor activation and how Wnts trigger canonical as opposed to noncanonical signaling have yet to be elucidated. We demonstrate here that prototype canonical Wnt3a and noncanonical Wnt5a ligands specifically trigger completely unrelated endogenous coreceptors-LRP5/6 and Ror1/2, respectively-through a common mechanism that involves their Wnt-dependent coupling to the Frizzled (Fzd) coreceptor and recruitment of shared components, including dishevelled (Dvl), axin, and glycogen synthase kinase 3 (GSK3). We identify Ror2 Ser 864 as a critical residue phosphorylated by GSK3 and required for noncanonical receptor activation by Wnt5a, analogous to the priming phosphorylation of low-density receptor-related protein 6 (LRP6) in response to Wnt3a. Furthermore, this mechanism is independent of Ror2 receptor Tyr kinase functions. Consistent with this model of Wnt receptor activation, we provide evidence that canonical and noncanonical Wnts exert reciprocal pathway inhibition at the cell surface by competition for Fzd binding. Thus, different Wnts, through their specific coupling and phosphorylation of unrelated coreceptors, activate completely distinct signaling pathways.

Allosteric modulation of a human odorant receptor.

Odor perception is first determined by how the myriad of environmental volatiles are detected at the periphery of the olfactory system. The combinatorial activation of dedicated odorant receptors generates enough encoding power for the discrimination of tens of thousands of odorants. Recent studies have revealed that odorant receptors undergo widespread inhibitory modulation of their activity when presented with mixtures of odorants, a property likely required to maintain discrimination and ensure sparsity of the code for complex mixtures. Here, we establish the role of human OR5AN1 in the detection of musks and identify distinct odorants capable of enhancing its activity in binary mixtures. Chemical and pharmacological characterization indicate that specific α-ß unsaturated aliphatic aldehydes act as positive allosteric modulators. Sensory experiments show decreased odor detection threshold in humans, suggesting that allosteric modulation of odorant receptors is perceptually relevant and likely adds another layer of complexity to how odors are encoded in the peripheral olfactory system.

Odorant Receptor Inhibition Is Fundamental to Odor Encoding.

Most natural odors are complex mixtures of volatile components, competing to bind odorant receptors (ORs) expressed in olfactory sensory neurons (OSNs) of the nose. To date, surprisingly little is known about how OR antagonism shapes neuronal representations in the detection layer of the olfactory system. Here, we investigated its prevalence, the degree to which it disrupts OR ensemble activity, and its conservation across phylogenetically related ORs. Calcium imaging microscopy of dissociated OSNs revealed significant inhibition, often complete attenuation, of responses to indole-a commonly occurring volatile associated with both floral and fecal odors-by a set of 36 tested odorants. To confirm an OR mechanism for the observed inhibition, we performed single-cell transcriptomics on OSNs exhibiting specific response profiles to a diagnostic panel of odorants and identified three paralogous receptors-Olfr740, Olfr741, and Olfr743-which, when tested in vitro, recapitulated OSN responses. We screened ten ORs from the Olfr740 gene family with ∼800 perfumery-related odorants spanning a range of chemical scaffolds and functional groups. Over half of these compounds (430) antagonized at least one of the ten ORs. OR activity fitted a mathematical model of competitive receptor binding and suggests normalization of OSN ensemble responses to odorant mixtures is the rule rather than the exception. In summary, we observed OR antagonism occurred frequently and in a combinatorial manner. Thus, extensive receptor-mediated computation of mixture information appears to occur in the olfactory epithelium prior to transmission of odor information to the olfactory bulb.

Canonical Wnts function as potent regulators of osteogenesis by human mesenchymal stem cells.

Genetic evidence indicates that Wnt signaling is critically involved in bone homeostasis. In this study, we investigated the functions of canonical Wnts on differentiation of adult multipotent human mesenchymal stem cells (hMSCs) in vitro and in vivo. We observe differential sensitivities of hMSCs to Wnt inhibition of osteogenesis versus adipogenesis, which favors osteoblastic commitment under binary in vitro differentiation conditions. Wnt inhibition of osteogenesis is associated with decreased expression of osteoblastic transcription factors and inhibition of c-Jun N-terminal kinase and p38 mitogen-activated protein kinase activation, which are involved in osteogenic differentiation. An hMSC subpopulation exhibits high endogenous Wnt signaling, the inhibition of which enhances osteogenic and adipogenic differentiation in vitro. In an in vivo bone formation model, high levels of Wnt signaling inhibit de novo bone formation by hMSCs. However, hMSCs with exogenous expression of Wnt1 but not stabilized beta-catenin markedly stimulate bone formation by naive hMSCs, arguing for an important role of a canonical Wnt gradient in hMSC osteogenesis in vivo.