Coxsackievirus A10 impairs nail regeneration and induces onychomadesis by mimicking DKK1 to attenuate Wnt signaling.

Coxsackievirus A10 (CV-A10) infection, a prominent cause of childhood hand-foot-and-mouth disease (HFMD), frequently manifests with the intriguing phenomenon of onychomadesis, characterized by nail shedding. However, the underlying mechanism is elusive. Here, we found that CV-A10 infection in mice could suppress Wnt/ß-catenin signaling by restraining LDL receptor-related protein 6 (LRP6) phosphorylation and ß-catenin accumulation and lead to onychomadesis. Mechanistically, CV-A10 mimics Dickkopf-related protein 1 (DKK1) to interact with Kringle-containing transmembrane protein 1 (KRM1), the CV-A10 cellular receptor. We further found that Wnt agonist (GSK3ß inhibitor) CHIR99021 can restore nail stem cell differentiation and protect against nail shedding. These findings provide novel insights into the pathogenesis of CV-A10 and related viruses in onychomadesis and guide prognosis assessment and clinical treatment of the disease.

Multifaceted effects of LRP6 in cancer: exploring tumor development, immune modulation and targeted therapies.

Low-density lipoprotein receptor (LDLR)-related protein 6 (LRP6), a member of the LDLR superfamily of cell surface receptors, is most widely known as a crucial co-receptor in the activation of canonical Wnt/ß-catenin signaling. This signaling pathway is implicated in multiple biological processes, such as lipoprotein metabolism, protease regulation, cell differentiation, and migration. LRP6 is frequently overexpressed in a variety of tumors, including liver cancer, colorectal cancer, and prostate cancer, and is generally considered an oncogene that promotes tumor proliferation, migration, and invasion. However, there are exceptions; some studies have reported that LRP6 inhibits lung metastasis of breast cancer through its ectodomain (LRP6N), and patients with low LRP6 expression tend to have a poor prognosis. Thus, the role of LRP6 in tumors remains controversial. Although limited studies have shown that LRP6 is associated with the expression and roles of a variety of immune cells in tumors, the interaction of LRP6 with the tumor microenvironment (TME) is not fully understood. Furthermore, it is crucial to acknowledge that LRP6 can engage with alternative pathways, including the mTORC1, CXCL12/CXCR4, and KRAS signaling pathways mentioned earlier, resulting in the regulation of biological functions independent of canonical Wnt/ß-catenin signaling. Due to the potential of LRP6 as a molecular target for cancer therapy, various treatment modalities have been developed to directly or indirectly inhibit LRP6 function, demonstrating promising anti-cancer effects across multiple cancer types. This review will concentrate on exploring the expression, function, and potential therapeutic applications of LRP6 in different cancer types, along with its influence on the TME.

WNT Signaling Cascade Proteins and LRP6 in the Formation of Various Types of Coronary Lesions in Patients With Coronary Artery Disease.

AIM: Assessment of WNT1, WNT3a, and LRP6 concentrations in patients with ischemic heart disease (IHD) and obstructive and non-obstructive coronary artery (CA) disease. MATERIAL AND METHODS: This cross-sectional observational study included 50 IHD patients (verified by coronary angiography, CAG), of which 25 (50%) were men, mean age 64.9±8.1 years; 20 patients had non-obstructive CA disease (stenosis <50%), and 30 patients had hemodynamically significant stenosis. Concentrations of WNT1, WNT3a and LRP6 were measured in all patients. RESULTS: The concentrations of WNT1 and WNT3a proteins were significantly higher in patients with IHD and obstructive CA disease (p < 0.001), while the concentration of LRP6 was higher in the group with non-obstructive CA disease (p = 0.016). Data analysis of the group with obstructive CA disease showed a moderate correlation between WNT1 and LRP6 (ρ=0.374; p=0.042). Correlation analysis of all groups of patients with CA disease revealed a moderate association between the concentrations of WNT1 and uric acid (ρ=0.416; p=0.007). Regression analysis showed that risk factors for the development of IHD, such as increased body mass index, age, smoking, dyslipidemia, and hypertension, did not significantly influence the type of CA disease in IHD patients. According to ROC analysis, the obstructive form of IHD was predicted by a WNT3a concentration higher than 0.155 ng/ml and a LRP6 concentration lower than 12.94 ng/ml. CONCLUSION: IHD patients with non-obstructive CA disease had the greatest increase in LRP6, while patients with obstructive CA disease had significantly higher concentrations of the canonical WNT cascade proteins, WNT1 and WNT3a. According to the ROC analysis, a WNT3a concentration >0.155 ng/ml can serve as a predictor for the presence of hemodynamically significant CA stenosis in IHD patients (sensitivity 96.7%; specificity 70%), whereas a LRP6 concentration >12.94 ng/ml can predict the development of non-obstructive CA disease (sensitivity 76.7%; specificity 65%).

Cachd1 interacts with Wnt receptors and regulates neuronal asymmetry in the zebrafish brain.

Neurons on the left and right sides of the nervous system often show asymmetric properties, but how such differences arise is poorly understood. Genetic screening in zebrafish revealed that loss of function of the transmembrane protein Cachd1 resulted in right-sided habenula neurons adopting left-sided identity. Cachd1 is expressed in neuronal progenitors, functions downstream of asymmetric environmental signals, and influences timing of the normally asymmetric patterns of neurogenesis. Biochemical and structural analyses demonstrated that Cachd1 can bind simultaneously to Lrp6 and Frizzled family Wnt co-receptors. Consistent with this, lrp6 mutant zebrafish lose asymmetry in the habenulae, and epistasis experiments support a role for Cachd1 in modulating Wnt pathway activity in the brain. These studies identify Cachd1 as a conserved Wnt receptor-interacting protein that regulates lateralized neuronal identity in the zebrafish brain.

PI15, a novel secreted WNT-signaling antagonist, regulates chondrocyte differentiation.

PURPOSE/AIM OF STUDY: During the development of the vertebrate skeleton, the progressive differentiation and maturation of chondrocytes from mesenchymal progenitors is precisely coordinated by multiple secreted factors and signaling pathways. The WNT signaling pathway has been demonstrated to play a major role in chondrogenesis. However, the identification of secreted factors that fine-tune WNT activity has remained elusive. Here, in this study, we have identified PI15 (peptidase inhibitor 15, protease Inhibitor 15, SugarCrisp), a member of the CAP (cysteine rich secretory proteins, antigen 5, and pathogenesis related 1 proteins) protein superfamily, as a novel secreted WNT antagonist dynamically upregulated during chondrocyte differentiation. MATERIALS AND METHODS: ATDC5 cells, C3H10T1/2 micromass cultures and primary chondrocyte cells were used as in vitro models of chondrogenesis. PI15 levels were stably depleted or overexpressed by viral shRNA or expression vectors. Chondrogenesis was evaluated by qPCR gene expression analysis and Alcian blue staining. Protein interactions were determined by coimmunoprecipitation assays. RESULTS AND CONCLUSIONS: shRNA-mediated knockdown of PI15 in ATDC5 cells, C3H10T1/2 cells or primary chondrocytes inhibits chondrogenesis, whereas the overexpression of PI15 strongly enhances chondrogenic potential. Mechanistically, PI15 binds to the LRP6 WNT co-receptor and blocks WNT-induced LRP6 phosphorylation, thus repressing WNT-induced transcriptional activity and alleviating the inhibitory effect of WNT signaling on chondrogenesis. Altogether, our findings suggest that PI15 acts as a key regulator of chondrogenesis and unveils a mechanism through which chondrocyte-derived molecules can modulate WNT activity as differentiation proceeds, thereby creating a positive feedback loop that further drives differentiation.

Targeting LRP6: A new strategy for cancer therapy.

Targeting specific molecular drivers of tumor growth is a key approach in cancer therapy. Among these targets, the low-density lipoprotein receptor-related protein 6 (LRP6), a vital component of the Wnt signaling pathway, has emerged as an intriguing candidate. As a cell-surface receptor and vital co-receptor, LRP6 is frequently overexpressed in various cancer types, implicating its pivotal role in driving tumor progression. The pursuit of LRP6 as a target for cancer treatment has gained substantial traction, offering a promising avenue for therapeutic intervention. Here, this comprehensive review explores recent breakthroughs in our understanding of LRP6's functions and underlying molecular mechanisms, providing a profound discussion of its involvement in cancer pathogenesis and drug resistance. Importantly, we go beyond discussing LRP6's role in cancer by discussing diverse potential therapeutic approaches targeting this enigmatic protein. These approaches encompass a wide spectrum, including pharmacological agents, natural compounds, non-coding RNAs, epigenetic factors, proteins, and peptides that modulate LRP6 expression or disrupt its interactions. In addition, also discussed the challenges associated with developing LRP6 inhibitors and their advantages over Wnt inhibitors, as well as the drugs that have entered phase II clinical trials. By shedding light on these innovative strategies, we aim to underscore LRP6's significance as a valuable and multifaceted target for cancer treatment, igniting enthusiasm for further research and facilitating translation into clinical applications.

ATP6AP2, a regulator of LRP6/ß-catenin protein trafficking, promotes Wnt/ß-catenin signaling and bone formation in a cell type dependent manner.

Wnt/ß-catenin signaling is critical for various cellular processes in multiple cell types, including osteoblast (OB) differentiation and function. Exactly how Wnt/ß-catenin signaling is regulated in OBs remain elusive. ATP6AP2, an accessory subunit of V-ATPase, plays important roles in multiple cell types/organs and multiple signaling pathways. However, little is known whether and how ATP6AP2 in OBs regulates Wnt/ß-catenin signaling and bone formation. Here we provide evidence for ATP6AP2 in the OB-lineage cells to promote OB-mediated bone formation and bone homeostasis selectively in the trabecular bone regions. Conditionally knocking out (CKO) ATP6AP2 in the OB-lineage cells (Atp6ap2Ocn-Cre) reduced trabecular, but not cortical, bone formation and bone mass. Proteomic and cellular biochemical studies revealed that LRP6 and N-cadherin were reduced in ATP6AP2-KO BMSCs and OBs, but not osteocytes. Additional in vitro and in vivo studies revealed impaired ß-catenin signaling in ATP6AP2-KO BMSCs and OBs, but not osteocytes, under both basal and Wnt stimulated conditions, although LRP5 was decreased in ATP6AP2-KO osteocytes, but not BMSCs. Further cell biological studies uncovered that osteoblastic ATP6AP2 is not required for Wnt3a suppression of ß-catenin phosphorylation, but necessary for LRP6/ß-catenin and N-cadherin/ß-catenin protein complex distribution at the cell membrane, thus preventing their degradation. Expression of active ß-catenin diminished the OB differentiation deficit in ATP6AP2-KO BMSCs. Taken together, these results support the view for ATP6AP2 as a critical regulator of both LRP6 and N-cadherin protein trafficking and stability, and thus regulating ß-catenin levels, demonstrating an un-recognized function of osteoblastic ATP6AP2 in promoting Wnt/LRP6/ß-catenin signaling and trabecular bone formation.

Sclerostin blockade inhibits bone resorption through PDGF receptor signaling in osteoblast lineage cells.

While sclerostin-neutralizing antibodies (Scl-Abs) transiently stimulate bone formation by activating Wnt signaling in osteoblast lineage cells, they exert sustained inhibition of bone resorption, suggesting an alternate signaling pathway by which Scl-Abs control osteoclast activity. Since sclerostin can activate platelet-derived growth factor receptors (PDGFRs) in osteoblast lineage cells in vitro and PDGFR signaling in these cells induces bone resorption through M-CSF secretion, we hypothesized that the prolonged anticatabolic effect of Scl-Abs could result from PDGFR inhibition. We show here that inhibition of PDGFR signaling in osteoblast lineage cells is sufficient and necessary to mediate prolonged Scl-Ab effects on M-CSF secretion and osteoclast activity in mice. Indeed, sclerostin coactivates PDGFRs independently of Wnt/ß-catenin signaling inhibition, by forming a ternary complex with LRP6 and PDGFRs in preosteoblasts. In turn, Scl-Ab prevents sclerostin-mediated coactivation of PDGFR signaling and consequent M-CSF upregulation in preosteoblast cultures, thereby inhibiting osteoclast activity in preosteoblast/osteoclast coculture assays. These results provide a potential mechanism explaining the dissociation between anabolic and antiresorptive effects of long-term Scl-Ab.

DKK1 Activates the PI3K/AKT Pathway via CKAP4 to Balance the Inhibitory Effect on Wnt/ß-Catenin Signaling and Regulates Wnt3a-Induced MSC Migration.

Wnt/ß-catenin signaling plays a crucial role in the migration of mesenchymal stem cells (MSCs). However, our study has revealed an intriguing phenomenon where Dickkopf-1 (DKK1), an inhibitor of Wnt/ß-catenin signaling, promotes MSC migration at certain concentrations ranging from 25 to 100 ng/mL while inhibiting Wnt3a-induced MSC migration at a higher concentration (400 ng/mL). Interestingly, DKK1 consistently inhibited Wnt3a-induced phosphorylation of LRP6 at all concentrations. We further identified cytoskeleton-associated protein 4 (CKAP4), another DKK1 receptor, to be localized on the cell membrane of MSCs. Overexpressing the CRD2 deletion mutant of DKK1 (ΔCRD2), which selectively binds to CKAP4, promoted the accumulation of active ß-catenin (ABC), the phosphorylation of AKT (Ser473) and the migration of MSCs, suggesting that DKK1 may activate Wnt/ß-catenin signaling via the CKAP4/PI3K/AKT cascade. We also investigated the effect of the CKAP4 intracellular domain mutant (CKAP4-P/A) that failed to activate the PI3K/AKT pathway and found that CKAP4-P/A suppressed DKK1 (100 ng/mL)-induced AKT activation, ABC accumulation, and MSC migration. Moreover, CKAP4-P/A significantly weakened the inhibitory effects of DKK1 (400 ng/mL) on Wnt3a-induced MSC migration and Wnt/ß-catenin signaling. Based on these findings, we propose that DKK1 may activate the PI3K/AKT pathway via CKAP4 to balance the inhibitory effect on Wnt/ß-catenin signaling and thus regulate Wnt3a-induced migration of MSCs. Our study reveals a previously unrecognized role of DKK1 in regulating MSC migration, highlighting the importance of CKAP4 and PI3K/AKT pathways in this process.

Low-density lipoprotein receptor-related protein 5/6 promotes endometrial cancer progression and cancer cell immune escape.

The study investigated the potential association of the low-density lipoprotein (LDL) genome with endometrial cancer progression based on the Gene Expression Omnibus data set and The Cancer Genome Atlas data set. Differential and weighted gene coexpression network analysis was performed on endometrial cancer transcriptome datasets GSE9750 and GSE106191. The protein-protein interaction network was built using LDL-receptor proteins and the top 50 tumor-associated genes. Low-density lipoprotein-related receptors 5/6 (LRP5/6) in endometrial cancer tissues were correlated with oncogenes, cell cycle-related genes, and immunological checkpoints using Spearman correlation. MethPrimer predicted the LRP5/6 promoter CpG island. LRP2, LRP6, LRP8, LRP12, low-density lipoprotein receptor-related protein-associated protein, and LRP5 were major LDL-receptor-related genes associated with endometrial cancer. LRP5/6 was enriched in various cancer-related pathways and may be a key LDL-receptor-related gene in cancer progression. LRP5/6 may be involved in the proliferation process of endometrial cancer cells by promoting the expression of cell cycle-related genes. LRP5/6 may be involved in the proliferation of endometrial cancer cells by promoting the expression of cell cycle-related genes. LRP5/6 may promote the immune escape of cancer cells by promoting the expression of immune checkpoints, promoting endometrial cancer progression. The MethPrimer database predicted that the LRP5/6 promoter region contained many CpG islands, suggesting that DNA methylation can occur in the LRP5/6 promoter region. LRP5/6 may aggravate endometrial cancer by activating the phosphoinositide 3-kinase/protein kinase B pathway.

Novel variant in LRP6 associated with unusual and severe clinical presentation: Case report.

Low-density lipoprotein receptor-related protein 6 (LRP6) is a co-receptor of the Wnt signaling pathway, which plays an essential role in various biological activities during embryonic and postnatal development. LRP6 is exceptionally associated with rare diseases and always with autosomal dominant inheritance. Here we report a familial phenotype of high bone mass associated with skeletal anomalies and oligodontia but also persistent left superior vena cava, inguinal hernia, hepatic cysts, abnormal posterior fossa and genital malformations. Molecular analysis revealed a novel heterozygous variant, NM_002336.2: c.724T>C, p.(Trp242Arg), in affected individuals. This variant is located in the first ß-propellant motif of LRP6, to which sclerostin (SOST) and dickkopf1 (DKK1), two LRP6 co-receptor inhibitors and various Wnt ligands bind. According to the literature and integrating data from structural analysis, this variant distorts the binding of SOST and DKK1, thus leading to overactivation of Wnt signaling pathways involved in osteoblast differentiation. This novel heterozygous variant in LRP6 underlies the role of LRP6 in skeletal and dental disorders as well as, probably, cardiac, cerebral and genital developments.

Uptake of ox-LDL by binding to LRP6 mediates oxidative stress-induced BMSCs senescence promoting obesity-related bone loss.

Obesity has long been thought to be a main cause of hyperlipidemia. As a systemic disease, the impact of obesity on organs, tissues and cells is almost entirely negative. However, the relationship between obesity and bone loss is highly controversial. On the one hand, obesity has long been thought to have a positive effect on bone due to increased mechanical loading on the skeleton, conducive to increasing bone mass to accommodate the extra weight. On the other hand, obesity-related metabolic oxidative modification of low-density lipoprotein (LDL) in vivo causes a gradual increase of oxidized LDL (ox-LDL) in the bone marrow microenvironment. We have reported that low-density lipoprotein receptor-related protein 6 (LRP6) acts as a receptor of ox-LDL and mediates the bone marrow stromal cells (BMSCs) uptake of ox-LDL. We detected elevated serum ox-LDL in obese mice. We found that ox-LDL uptake by LRP6 led to an increase of intracellular reactive oxygen species (ROS) in BMSCs, and N-acetyl-L-cysteine (NAC) alleviated the cellular senescence and impairment of osteogenesis induced by ox-LDL. Moreover, LRP6 is a co-receptor of Wnt signaling. We found that LRP6 preferentially binds to ox-LDL rather than dickkopf-related protein 1 (DKK1), both inhibiting Wnt signaling and promoting BMSCs senescence. Mesoderm development LRP chaperone (MESD) overexpression inhibits ox-LDL binding to LRP6, attenuating oxidative stress and BMSCs senescence, eventually rescuing bone phenotype.

The inhibition of YTHDF3/m6A/LRP6 reprograms fatty acid metabolism and suppresses lymph node metastasis in cervical cancer.

The lipid synthesis of fatty acid (FA) represents a significant hallmark in the occurrence and progression of malignant tumor, which are associated with lymph node (LN) metastasis. Elucidation of the molecular mechanisms underlying LN metastasis could provide therapeutic strategies for cervical cancer (CCa). N6-Methyladenosine (m6A), the most prevalent and abundant RNA modification, exerts specific regulatory control over a series of oncogene expressions. This study demonstrated a clinical correlation between the upregulation of the m6A reader YTHDF3 and LN metastasis, thereby contributing to poor overall survival probability (OS) among CCa patients. The mechanistic investigation revealed that SREBF1 transcriptionally activated YTHDF3 expression by binding to its promoter. Functional experiments demonstrated that the upregulation of YTHDF3 significantly enhanced the in vitro proliferative, migratory, and invasive capacities of CCa cells, while also promoting lymphangiogenesis and facilitating LN metastasis in vivo. Mechanistically, the upregulation of LRP6 through YTHDF3-mediated m6A modification resulted in increased expression of FASN and ACC1, leading to both lipolysis of lipid droplets and synthesis of free fatty acid. Ultimately, this promoted fatty acid metabolism and enhanced LN metastasis by activating the LRP6-YAP-VEGF-C axis, which could induce lymphangiogenesis in CCa. Our study highlighted that YTHDF3 can serve as a promising therapeutic target and predictive biomarker for CCa patients with LN metastasis.

LRP6 is a potential biomarker of kidney clear cell carcinoma related to prognosis and immune infiltration.

Renal cell carcinoma is the most common and most lethal genitourinary tumor. The causes of renal clear cell carcinoma are complex and the heterogeneity of the tumor tissue is high, so patient outcomes are not very satisfactory. Exploring biomarkers in the progression of renal clear cell carcinoma is crucial to improve the diagnosis and guide the treatment of renal clear cell carcinoma. LRP6 is a co-receptor of the Wnt/ß-catenin signaling pathway, which is involved in cell growth, inflammation and cell transformation through activation of the Wnt/ß-catenin signaling pathway. Abnormal expression of LRP6 is associated with the malignant phenotype, metastatic potential and poor prognosis of various tumors. In this study, we found that LRP6 was abnormally highly expressed in a variety of tumors and significantly correlated with microsatellite instability, tumor mutation burden, and immune cell infiltration and immune checkpoint expression in a variety of tumors. Moreover, we found that LRP6 was significantly associated with the prognosis of renal clear cell carcinoma. Further we found a significant correlation between LRP6 and the expression of m6A-related genes and ferroptosis-related genes. Finally, we also found a significant correlation between the expression of LRP6 and the sensitivity to common drugs used in kidney clear cell carcinoma treatment. These results suggest that LRP6 is likely to be a potential target for kidney clear cell carcinoma treatment.

Low-density lipoprotein receptor-related protein 6 ablation in macrophages differentially inhibits lung injury-mediated inflammation and metastasis.

Low-density lipoprotein receptor-related protein 6 (LRP6) is a receptor protein for Wnt ligands. Yet, their role in immune cell regulation remains elusive. Here we demonstrated that genetic deletion of LRP6 in macrophages using LysM-cre Lrp6fl/fl (Lrp6MKO) mice showed differential inhibition of inflammation in the bleomycin (BLM)-induced lung injury model and B16F10 melanoma lung metastasis model. Lrp6MKO mice showed normal immune cell populations in the lung and circulating blood in homeostatic conditions. In the BLM-induced lung injury model, Lrp6MKO mice showed a decreased number of monocyte-derived alveolar macrophages, reduced collagen deposition and alpha-smooth muscle actin (αSMA) protein levels in the lung. In B16F10 lung metastasis model, Lrp6MKO mice reduced lung tumor foci. Monocytic and granulocytic-derived myeloid-derived suppressor cells (M-MDSCs and G-MDSCs) were increased in the lung. In G-MDSCs, hypoxia-inducible factor 1α (HIF1α)+ PDL1+ population was markedly decreased but not in M-MDSCs. Taken together, our results show that the role of LRP6 in macrophages is differential depending on the inflammation microenvironment in the lung.

Kallistatin as an inhibitory protein against colorectal cancer cells through binding to LRP6.

Kallistatin (KL) is a member of the serine proteinase inhibitor (serpin) family regulating oxidative stress, vascular relaxation, inflammation, angiogenesis, cell proliferation, and invasion. The heparin-binding site of Kallistatin has an important role in the interaction with LRP6 leading to the blockade of the Wnt signaling pathway. In this study, we aimed to explore the structural basis of the Kallistatin-LRP6E1E4 complex using in silico approaches and evaluating the anti-proliferative, apoptotic, and cell cycle arrest activities of Kallistatin in colon cancer lines. The molecular docking showed Kallistatin could bind to the LRP6E3E4 much stronger than LRP6E1E2. The Kallistatin-LRP6E1E2 and Kallistatin-LRP6E3E4 complexes were stable during Molecular Dynamics (MD) simulation. The Molecular Mechanics/Poisson-Boltzmann Surface Area (MM/PBSA) showed that the Kallistatin-LRP6E3E4 has a higher binding affinity compared to Kallistatin-LRP6E1E2. Kallistatin induced higher cytotoxicity and apoptosis in HCT116 compared to the SW480 cell line. This protein-induced cell-cycle arrest in both cell lines at the G1 phase. The B-catenin, cyclin D1, and c-Myc expression levels were decreased in response to treatment with Kallistatin in both cell lines while the LRP6 expression level was decreased in the HCT116 cell line. Kallistatin has a greater effect on the HCT116 cell line compared to the SW480 cell line. Kallistatin can be used as a cytotoxic and apoptotic-inducing agent in colorectal cancer cell lines.

The USP46 complex deubiquitylates LRP6 to promote Wnt/ß-catenin signaling.

The relative abundance of Wnt receptors plays a crucial role in controlling Wnt signaling in tissue homeostasis and human disease. While the ubiquitin ligases that ubiquitylate Wnt receptors are well-characterized, the deubiquitylase that reverses these reactions remains unclear. Herein, we identify USP46, UAF1, and WDR20 (USP46 complex) as positive regulators of Wnt signaling in cultured human cells. We find that the USP46 complex is similarly required for Wnt signaling in Xenopus and zebrafish embryos. We demonstrate that Wnt signaling promotes the association between the USP46 complex and cell surface Wnt coreceptor, LRP6. Knockdown of USP46 decreases steady-state levels of LRP6 and increases the level of ubiquitylated LRP6. In contrast, overexpression of the USP46 complex blocks ubiquitylation of LRP6 by the ubiquitin ligases RNF43 and ZNFR3. Size exclusion chromatography studies suggest that the size of the USP46 cytoplasmic complex increases upon Wnt stimulation. Finally, we show that USP46 is essential for Wnt-dependent intestinal organoid viability, likely via its role in LRP6 receptor homeostasis. We propose a model in which the USP46 complex increases the steady-state level of cell surface LRP6 and facilitates the assembly of LRP6 into signalosomes via a pruning mechanism that removes sterically hindering ubiquitin chains.

Circ_0011373 promotes papillary thyroid carcinoma progression by regulating miR-1271/LRP6 axis.

PURPOSE: This research aimed to explore the regulatory molecular mechanism among circular RNA (circ)_0011373, microRNA (miR)-1271, and lipoprotein receptor-related protein 6 (LRP6) in papillary thyroid carcinoma (PTC). METHODS: Quantitative real-time PCR (qRT-PCR) assay was adopted to measure the expression of circ_0011373, miR-1271, and LRP6 mRNA. Furthermore, cell cycle distribution, apoptosis, migration and invasion were investigated by flow cytometry and transwell assay, respectively. The target relationship between miR-1271 and circ_0011373 or LRP6 was predicted by using the Starbase website and DIANA TOOL and verified by dual-luciferase reporter and RIP assay. Protein expression levels of LRP6, p-mTOR, mTOR, p-AKT, AKT, p-PI3K, and PI3K were tested by Western blot. The function of circ_0011373 on PTC tumor growth was validated by the xenograft tumor model in vivo. RESULTS: Circ_0011373 and LRP6 were upregulated, while miR-1271 was downregulated in PTC tissues and cell lines. Moreover, knockdown of circ_0011373 inhibited cell cycle, migration, and invasion and promoted apoptosis. Of particular importance was the fact that circ_0011373 directly interacted with miR-1271 and miR-1271 inhibitor was able to reverse the effect of circ_0011373 knockdown on PTC cell progression. Meanwhile, LRP6 was directly targeted by miR-1271, and its expression was positively regulated by circ_0011373. We further confirmed that miR-1271 overexpression suppressed cell cycle, migration, and invasion and enhanced apoptosis by regulating LRP6. In addition, circ_0011373 knockdown restrained PTC tumor growth in vivo. CONCLUSION: Circ_0011373 might be able to regulate PTC cell cycle, migration, invasion, and apoptosis by regulating the miR-1271/LRP6 axis.

Screening and Identification of ssDNA Aptamers for Low-Density Lipoprotein (LDL) Receptor-Related Protein 6.

Low-density lipoprotein receptor-related protein 6 (LRP6), a member of the low-density lipoprotein receptor (LDLR) family, displays a unique structure and ligand-binding function. As a co-receptor of the Wnt/ß-catenin signaling pathway, LRP6 is a novel therapeutic target that plays an important role in the regulation of cardiovascular disease, lipid metabolism, tumorigenesis, and some classical signals. By using capillary electrophoresis-systematic evolution of ligands by exponential enrichment (CE-SELEX), with recombinant human LRP-6 as the target, four candidate aptamers with a stem-loop structure were selected from an ssDNA library-AptLRP6-A1, AptLRP6-A2, AptLRP6-A3, and AptLRP6-A4. The equilibrium dissociation constant KD values between these aptamers and the LRP6 protein were in the range of 0.105 to 1.279 µmol/L, as determined by CE-LIF analysis. Their affinities and specificities were further determined by the gold nanoparticle (AuNP) colorimetric method. Among them, AptLRP6-A3 showed the highest affinity with LRP6-overexpressed human breast cancer cells. Therefore, the LRP6 aptamer identified in this study constitutes a promising modality for the rapid diagnosis and treatment of LRP6-related diseases.

Salinomycin sodium exerts anti diffuse large B-cell lymphoma activity through inhibition of LRP6-mediated Wnt/ß-catenin and mTORC1 signaling.

Low-density lipoprotein receptor-related protein-6 (LRP6) is overexpressed in various cancers. The small molecule salinomycin sodium inhibits LRP6. We observed a higher proportion of subjects with non-germinal center B (non-GCB) subtypes having high LRP6 expression than those with GCB subtypes by immunohistochemistry. The PCR and Western blot assays demonstrated increased LRP6 expression in non-GCB subtype cells. In addition, CCK-8 assays and transwell cell migration assays revealed that salinomycin sodium exhibited dose- and time-dependent inhibition of proliferation and migration in non-GCB subtype cells. Furthermore, Western blot assays showed that salinomycin sodium decreased the expression of Bcl2, while increasing the expression of Bax. Additionally, salinomycin sodium suppressed LRP6 expression, blocked LRP6 phosphorylation, and inhibited the Wnt/ß-catenin and mTORC1 signaling pathways. Our results suggest that LRP6 is highly expressed in non-GCB subtype. Furthermore, salinomycin sodium inhibited LRP6 expression and the Wnt/ß-catenin and mTORC1 signaling in non-GCB subtype cells, and displayed potent anticancer activity.