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Structural insight into the mechanisms of Wnt signaling antagonism by Dkk.

ABSTRACT: Dickkopf (Dkk) proteins are antagonists of the canonical Wnt signaling pathway and are crucial for embryonic cell fate and bone formation. Wnt antagonism of Dkk requires the binding of the C-terminal cysteine-rich domain of Dkk to the Wnt coreceptor, LRP5/6. However, the structural basis of the interaction between Dkk and low density lipoprotein receptor-related protein (LRP) 5/6 is unknown. In this study, we examined the structure of the Dkk functional domain and elucidated its interactions with LRP5/6. Using NMR spectroscopy, we determined the solution structure of the C-terminal cysteine-rich domain of mouse Dkk2 (Dkk2C). Then, guided by mutagenesis studies, we docked Dkk2C to the YWTD beta-propeller domains of LRP5/6 and showed that the ligand binding site of the third LRP5/6 beta-propeller domain matches Dkk2C best, suggesting that this domain binds to Dkk2C with higher affinity. Such differential binding affinity is likely to play an essential role in Dkk function in the canonical Wnt pathway.

SUBMITTER: Chen L

PROVIDER: S-EPMC2517007 | biostudies-literature | 2008 Aug

REPOSITORIES: biostudies-literature

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Structural insight into the mechanisms of Wnt signaling antagonism by Dkk.

Chen Lijun L Wang Ke K Shao Youming Y Huang Jin J Li Xiaofeng X Shan Jufang J Wu Dianqing D Zheng Jie J JJ

The Journal of biological chemistry 20080603 34

Dickkopf (Dkk) proteins are antagonists of the canonical Wnt signaling pathway and are crucial for embryonic cell fate and bone formation. Wnt antagonism of Dkk requires the binding of the C-terminal cysteine-rich domain of Dkk to the Wnt coreceptor, LRP5/6. However, the structural basis of the interaction between Dkk and low density lipoprotein receptor-related protein (LRP) 5/6 is unknown. In this study, we examined the structure of the Dkk functional domain and elucidated its interactions wit ...[more]

PMID: 18524778

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Project description:Patients with Gaucher disease (GD) have progressive bone involvement that clinically presents with debilitating bone pain, structural bone changes, bone marrow infiltration (BMI), Erlenmeyer (EM) flask deformity, and osteoporosis. Pain is referred by the majority of GD patients and continues to persist despite the type of therapy. The pain in GD is described as chronic deep penetrating pain; however, sometimes, patients experience severe acute pain. The source of bone pain is mainly debated as nociceptive pain secondary to bone pathology or neuropathic or inflammatory origins. Osteocytes constitute a significant source of secreted molecules that coordinate bone remodeling. Osteocyte markers, sclerostin (SOST) and Dickkopf-1 (DKK-1), inactivate the canonical Wnt signaling pathway and lead to the inhibition of bone formation. Thus, circulated sclerostin and DKK-1 are potential biomarkers of skeletal abnormalities. This study aimed to assess the circulating levels of sclerostin and DKK-1 in patients with GD and their correlation with clinical bone pathology parameters: pain, bone mineral density (BMD), and EM deformity. Thirty-nine patients with GD were classified into cohorts based on the presence and severity of bone manifestations. The serum levels of sclerostin and DKK-1 were quantified by enzyme-linked immunosorbent assays. The highest level of sclerostin was measured in GD patients with pain, BMI, and EM deformity. The multiparameter analysis demonstrated that 95% of GD patients with pain, BMI, and EM deformity had increased levels of sclerostin. The majority of patients with elevated sclerostin also have osteopenia or osteoporosis. Moreover, circulating sclerostin level increase with age, and GD patients have elevated sclerostin levels when compared with healthy control from the same age group. Pearson's linear correlation analysis showed a positive correlation between serum DKK-1 and sclerostin in healthy controls and GD patients with normal bone mineral density. However, the balance between sclerostin and DKK-1 waned in GD patients with osteopenia or osteoporosis. In conclusion, the osteocyte marker, sclerostin, when elevated, is associated with bone pain, BMI, and EM flask deformity in GD patients. The altered sclerostin/DKK-1 ratio correlates with the reduction of bone mineral density. These data confirm that the Wnt signaling pathway plays a role in GD-associated bone disease. Sclerostin and bone pain could be used as biomarkers to assess patients with a high risk of BMI and EM flask deformities.

Project description:BackgroundLiver fibrosis is an outcome of restoring process in chronic liver injury. Human amniotic mesenchymal stem cells (hAMSCs) derived from amniotic membrane have multilineage differentiation, immunosuppressive, and anti-inflammatory potential which makes them suitable for treating liver fibrosis. This study aimed to explore the effect and mechanism of hAMSCs on liver fibrosis.MethodshAMSCs were transplanted into carbon tetrachloride (CCl4)-induced liver fibrosis mice via tail vein, and the effects of hAMSCs on hepatic fibrosis were assessed. The effects of hAMSCs and hAMSCs conditional medium (CM) on the activation of hepatic stellate cells (HSCs) were investigated in vivo and in vitro. Antibody array assay was used to identify the cytokines secreted by hAMSCs that may inhibit the activation of HSCs. Finally, the underlying mechanisms were explored by assessing IGF-1R/PI3K/AKT and GSK3β/β-catenin signaling pathways in the activated HSCs (LX-2) with hAMSCs and hAMSCs transfected with corresponding siRNAs.ResultsOur results showed that hAMSCs possessed the characterizations of mesenchymal stem cells. hAMSCs significantly reduced liver fibrosis and improved liver function in mice by inhibiting HSCs activation in vivo. Both hAMSCs and hAMSC-CM remarkably inhibited the collagen deposition and activation of LX-2 cells in vitro. Antibody array assay showed that insulin-like growth factor binding protein-3 (IGFBP-3), Dickkopf-3 (DKK-3), and Dickkopf-1 (DKK-1) were highly expressed in the co-culture group and hAMSC-CM group compared with LX-2 group. Western blot assay demonstrated that IGFBP-3, DKK-3, and DKK-1 derived from hAMSCs inhibit LX-2 cell activation through blocking canonical Wnt signaling pathway.ConclusionsOur results demonstrated that IGFBP-3, Dkk3, and DKK-1 secreted by hAMSCs attenuated liver fibrosis in mice through inhibiting HSCs activation via depression of Wnt/β-catenin signaling pathway, suggesting that hAMSCs or hAMSC-CM provides an alternative therapeutic approach for the treatment of liver fibrosis.

Dataset Information

Structural insight into the mechanisms of Wnt signaling antagonism by Dkk.

Publications

Structural insight into the mechanisms of Wnt signaling antagonism by Dkk.

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