Cilia as Wnt signaling organelles.

Cilia and Wnt signaling have a complex relationship, wherein Wnt regulates cilia and, conversely, cilia may affect Wnt signaling. Recently, it was shown that Wnt receptors are present in flagella, primary cilia, and multicilia, where they transmit an intraciliary signal that is independent of ß-catenin. Intraciliary Wnt signaling promotes ciliogenesis, affecting male fertility, adipogenesis, and mucociliary clearance. Wnt also stimulates the beating of motile cilia, highlighting that these nanomotors, too, are chemosensory. Intraciliary Wnt signaling employs a Wnt-protein phosphatase 1 (PP1) signaling axis, involving the canonical Wnt pathway's inhibition of glycogen synthase kinase 3 (GSK3) to repress PP1 activity. Collectively, these findings support that cilia are Wnt signaling organelles, with implications for ciliopathies and cancer.

Mucociliary Wnt signaling promotes cilia biogenesis and beating.

It is widely thought that Wnt/Lrp6 signaling proceeds through the cytoplasm and that motile cilia are signaling-inert nanomotors. Contrasting both views, we here show in the mucociliary epidermis of X. tropicalis embryos that motile cilia transduce a ciliary Wnt signal that is distinct from canonical ß-catenin signaling. Instead, it engages a Wnt-Gsk3-Ppp1r11-Pp1 signaling axis. Mucociliary Wnt signaling is essential for ciliogenesis and it engages Lrp6 co-receptors that localize to cilia via a VxP ciliary targeting sequence. Live-cell imaging using a ciliary Gsk3 biosensor reveals an immediate response of motile cilia to Wnt ligand. Wnt treatment stimulates ciliary beating in X. tropicalis embryos and primary human airway mucociliary epithelia. Moreover, Wnt treatment improves ciliary function in X. tropicalis ciliopathy models of male infertility and primary ciliary dyskinesia (ccdc108, gas2l2). We conclude that X. tropicalis motile cilia are Wnt signaling organelles that transduce a distinct Wnt-Pp1 response.

Primary cilia are WNT-transducing organelles whose biogenesis is controlled by a WNT-PP1 axis.

WNT signaling is important in development, stem cell maintenance, and disease. WNT ligands typically signal via receptor activation across the plasma membrane to induce ß-catenin-dependent gene activation. Here, we show that in mammalian primary cilia, WNT receptors relay a WNT/GSK3 signal that ß-catenin-independently promotes ciliogenesis. Characterization of a LRP6 ciliary targeting sequence and monitoring of acute WNT co-receptor activation (phospho-LRP6) support this conclusion. Ciliary WNT signaling inhibits protein phosphatase 1 (PP1) activity, a negative regulator of ciliogenesis, by preventing GSK3-mediated phosphorylation of the PP1 regulatory inhibitor subunit PPP1R2. Concordantly, deficiency of WNT/GSK3 signaling by depletion of cyclin Y and cyclin-Y-like protein 1 induces primary cilia defects in mouse embryonic neuronal precursors, kidney proximal tubules, and adult mice preadipocytes.

Combinatory Exposure to Urolithin A, Alternariol, and Deoxynivalenol Affects Colon Cancer Metabolism and Epithelial Barrier Integrity in vitro.

The human gastrointestinal tract is an important site of nutrient absorption and a crucial barrier against xenobiotics. It regularly faces "chemical cocktails" composed of food constituents, their human and microbial metabolites, and foodborne contaminants, such as mycotoxins. Hence, the colonic epithelium adapts to dietary molecules tuning its immune response, structural integrity, and metabolism to maintain intestinal homeostasis. While gut microbiota metabolites of berry ellagitannins, such as urolithin A (Uro A) might contribute to physiological epithelial barrier integrity, foodborne co-contaminating mycotoxins like alternariol (AOH) and deoxynivalenol (DON) could hamper epithelial function. Hence, we investigated the response of differentiated Caco-2 cells (clone C2BBe1) in vitro to the three compounds alone or in binary mixtures. In virtue of the possible interactions of Uro A, AOH, and DON with the aryl hydrocarbon receptor (AhR) pathway, potential effects on phase-I-metabolism enzymes and epithelial structural integrity were taken as endpoints for the evaluation. Finally, Liquid chromatography tandem mass spectrometry measurements elucidated the absorption, secretion, and metabolic capacity of the cells under single and combinatory exposure scenarios. Uro A and AOH as single compounds, and as a binary mixture, were capable to induce CYP1A1/1A2/1B1 enzymes triggered by the AhR pathway. In light of its ribosome inhibiting capacity, the trichothecene suppressed the effects of both dibenzo-α-pyrones. In turn, cellular responsiveness to Uro A and AOH could be sustained when co-exposed to DON-3-sulfate, instead of DON. Colonic epithelial structural integrity was rather maintained after incubation with Uro A and AOH: this was reinforced in the combinatory exposure scenario and disrupted by DON, an effect, opposed in combination. Passage through the cells as well as the metabolism of Uro A and AOH were rather influenced by co-exposure to DON, than by interaction with each other. Therefore, we conclude that although single foodborne bioactive substances individually could either support or disrupt the epithelial structure and metabolic capacity of colon cancer, exposure to chemical mixtures changes the experimental outcome and calls for the need of combinatory investigations for proper risk assessment.

R-spondins are BMP receptor antagonists in Xenopus early embryonic development.

BMP signaling plays key roles in development, stem cells, adult tissue homeostasis, and disease. How BMP receptors are extracellularly modulated and in which physiological context, is therefore of prime importance. R-spondins (RSPOs) are a small family of secreted proteins that co-activate WNT signaling and function as potent stem cell effectors and oncogenes. Evidence is mounting that RSPOs act WNT-independently but how and in which physiological processes remains enigmatic. Here we show that RSPO2 and RSPO3 also act as BMP antagonists. RSPO2 is a high affinity ligand for the type I BMP receptor BMPR1A/ALK3, and it engages ZNRF3 to trigger internalization and degradation of BMPR1A. In early Xenopus embryos, Rspo2 is a negative feedback inhibitor in the BMP4 synexpression group and regulates dorsoventral axis formation. We conclude that R-spondins are bifunctional ligands, which activate WNT- and inhibit BMP signaling via ZNRF3, with implications for development and cancer.

MicroRNA-182-5p regulates hedgehog signaling pathway and chemosensitivity of cisplatin-resistant lung adenocarcinoma cells via targeting GLI2.

Lung cancer is one of the deadliest cancers worldwide. Late diagnosis at an advanced, inoperable stage makes chemotherapy a treatment of choice, yet, with low response rates. The hedgehog signaling pathway (HHSP) is often reactivated in cancer. We identified miR-182-5p as a regulator of GLI2, a transcriptional regulator of the HHSP, and explored the role of the miR-182-5p/GLI2 axis in carcinogenesis and cisplatin resistance of lung adenocarcinoma (LADC). Expression of miRNAs and target genes was analyzed by RT-qPCR, expression of the GLI-protein family in LADC and adjacent lung tissue (n = 27 pairs) by immunohistochemistry. MiR-182-5p was manipulated, and data were generated by immunoblotting, immunofluorescence, apoptosis, proliferation/viability, dual-luciferase-, and colony forming assays. MiR-182-5p was down-regulated in cisplatin-resistant LADC cells and directly targeted GLI2. Interference with miR-182-5p or GLI2 silencing resulted in modulation of cell proliferation, clonogenic potential, and cisplatin-sensitivity. HHSP was markedly reactivated in LADC tissue compared to adjacent non-malignant lung tissue. Our results indicate that the miR-182-5p/GLI2 axis modulates tumorigenesis and cisplatin-resistance in LADC cells, by influencing the HHSP. Therefore, this axis might be considered as a potential biomarker and future therapeutic target in LADC patients.