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Summary The balance and distribution of epithelial cell types is required to maintain tissue homeostasis. A hallmark of airway diseases is epithelial remodeling, leading to increased goblet cell numbers and an overproduction of mucus. In the conducting airway, basal cells act as progenitors for both secretory and ciliated cells. To identify mechanisms regulating basal cell fate, we developed a screenable 3D culture system of airway epithelial morphogenesis. We performed a high-throughput screen using a collection of secreted proteins and identified inflammatory cytokines that specifically biased basal cell differentiation toward a goblet cell fate, culminating in enhanced mucus production. We also demonstrate a specific requirement for Notch2 in cytokine-induced goblet cell metaplasia in vitro and in vivo. We conclude that inhibition of Notch2 prevents goblet cell metaplasia induced by a broad range of stimuli and propose Notch2 neutralization as a therapeutic strategy for preventing goblet cell metaplasia in airway diseases. [ABSTRACT FROM AUTHOR]

R-spondin proteins sensitize cells to Wnt signalling and act as potent stem cell growth factors. Various membrane proteins have been proposed as potential receptors of R-spondin, including LGR4/5, membrane E3 ubiquitin ligases ZNRF3/RNF43 and several others proteins. Here, we show that R-spondin interacts with ZNRF3/RNF43 and LGR4 through distinct motifs. Both LGR4 and ZNRF3 binding motifs are required for R-spondin-induced LGR4/ZNRF3 interaction, membrane clearance of ZNRF3 and activation of Wnt signalling. Importantly, Wnt-inhibitory activity of ZNRF3, but not of a ZNRF3 mutant with reduced affinity to R-spondin, can be strongly suppressed by R-spondin, suggesting that R-spondin primarily functions by binding and inhibiting ZNRF3. Together, our results support a dual receptor model of R-spondin action, where LGR4/5 serve as the engagement receptor whereas ZNRF3/RNF43 function as the effector receptor. [ABSTRACT FROM AUTHOR]

The phosphodiesterases (PDEs) are metal ion-dependent enzymes that regulate cellular signaling by metabolic inactivation of the ubiquitous second messengers cAMP and cGMP. In this role, the PDEs are involved in many biological and metabolic processes and are proven targets of successful drugs for the treatments of a wide range of diseases. However, because of the rapidity of the hydrolysis reaction, an experimental knowledge of the enzymatic mechanisms of the PDEs at the atomic level is still lacking. Here, we report the structures of reaction intermediates accumulated at the reaction steady state in PDE9/crystal and preserved by freeze-trapping. These structures reveal the catalytic process of a PDE and explain the substrate specificity of PDE9 in an actual reaction and the cation requirements of POEs in general. [ABSTRACT FROM AUTHOR]

Lasofoxifene is a new and potent selective estrogen receptor modulator (SERM). The structural basis of its interaction with the estrogen receptor has been investigated by crystallographic analysis of its complex with the ligand-binding domain of estrogen receptor α at a resolution of 2.0 Å. As with other SERMs, lasofoxifene diverts the receptor from its agonist-bound conformation by displacing the C-terminal AF-2 helix into the site at which the LXXLL motif of coactivator proteins would otherwise be able to bind. Lasofoxifene achieves this effect by occupying the space normally filled by residue Leu 540, as well as by modulating the conformation of residues of helix 11 (His 524, Leu 525). A well-defined salt bridge between lasofoxifene and Asp 351 suggests that charge neutralization in this region of the receptor may explain the some of the antiestrogenic effects of lasofoxifene. The results suggest general features of ERα/SERM recognition, and add a new dimension to efforts to rationalize differences between the biological activity profiles exhibited by these important pharmacological agents. [ABSTRACT FROM AUTHOR]

Analyzes the kinetics of the interaction between estrogen receptors and ligands. Use of a surface plasmon resonance biosensor technology; Details of the association and disassociation phase of binding responses; Types of ligands.

In order to confirm and further explore the significance of the overexpression of the CRABP II (cellular retinoic acid binding protein type II) and psoriasin genes in psoriatic versus normal skin, we examined the mRNA expression levels of these two genes by in situ hybridization in skin samples from psoriatic plaques and in one case from the border between a psoriatic plaque and uninvolved skin. Both genes were markedly upregulated in lesional skin, with a shift from low to high expression in the transitional zone of the plaque. Expression of the cytokeratin 1 (K1) gene was, in contrast, high in normal skin and decreased in the transition from uninvolved skin to psoriatic plaque, Examination of mRNA levels of CRABP II and psoriasin in other hyperproliferative and inflammatory skin diseases showed high expression of psoriasin, and in some cases also of CRABP II, in atopic dermatitis, mycosis fungoides, Darier's disease and inflammatory lichen sclerosus et atrophicus. In atrophic lesions of lichen sclerosus et atrophicus that lacked an inflammatory infiltrate, these changes were only weakly expressed. These findings demonstrate that altered epidermal gene expression of K1, psoriasin and CRABP II is not disease-specific and may reflect instead an altered state of epidermal differentiation and/or may be linked to the inflammation and cellular infiltration common to all the conditions studied. [ABSTRACT FROM AUTHOR]

Metabolic defects in phytanic acid catabolism have been shown to be connected with a number of human diseases which can lead to lethal defects of the nervous system and other organs. These effects are probably a result of the very high accumulation of phytanic acid in tissues throughout the body, due to defects in phytanic acid oxidation, the peroxisome being a major site for this process. The nuclear hormone receptors peroxisome proliferator-activated receptor and retinoid X receptor (RXR) have been shown to function as transcription factors in the control of the peroxisomal enzyme expression. Known activators of peroxisome proliferator-activated receptor include polyunsaturated fatty acids and, for RXR, the 9-cis isomer of retinoic acid. Here we report that phytanic acid is also a natural ligahd for RXRa, being able to activate a RXR-responsive promoter. We present evidence that phytanic acid binds to RXRa, promotes formation of an RXRa/RXR response element complex (as detected by gel retardation), and induces a RXRa conformational change similar to that induced by 9-cis-retinoic acid (as detected by protease sensitivity). These results suggest an involvment of RXRa in the control of fatty acid metabolism and could simply that RXRs have a role in the disease effects resulting from defective phytanic acid catabolism. [ABSTRACT FROM AUTHOR]