Search

Activation of the Wnt signalling pathway is vital in the anterior-posterior patterning during neural development. In a new study, Stephane Angers and colleagues leverage previously developed selective antibodies against Frizzled receptors of the Wnt pathway to stimulate midbrain progenitor differentiation in human pluripotent stem cells. We caught up with first author Andy Yang and corresponding author Stephane Angers, Professor at the University of Toronto, to learn more about the story behind the paper., (© 2024. Published by The Company of Biologists Ltd.)

As a family of G protein-coupled receptors (GPCRs) with a seven-span transmembrane structure, frizzled class receptors (FZDs) play crucial roles in regulating multiple biological functions. However, their transcriptional expression profile and prognostic significance in acute myeloid leukemia (AML) are unclear. In AML, the role of FZDs was explored by performing the comprehensive analysis on the relationship between clinical characteristics and mRNA expression profiles from public databases including cBioPortal for Cancer Genomics, Gene Expression Profile Interactive Analysis (GEPIA), and Cancer Cell Line Encyclopedia (CCLE). We identified that in the majority of 27 AML cell lines, frizzled class receptor 6 (FZD6) was high-expressed. A significantly higher expression of FZD6 in AML patients was observed when compared to normal controls ( P < 0.01 ). Compared with intermediate and poor/adverse risk group patients, FZD6 expressed much lower in cytogenetic favorable risk group patients ( P < 0.0001 ). Patients with higher-expressed FZD6 were associated with shorter overall survival (OS) ( P = 0.0089 ) rather than progression-free survival (PFS). However, the predictive effect of FZD6 on OS could be reversed by hematopoietic stem cell transplantation (HSCT). The data of gene set enrichment analysis (GSEA) demonstrated that 4 gene sets, including MYC targets, HEME metabolism, E2F targets, and UV response, were differentially enriched in the high-expression FZD6 group. To conclude, the study suggested that high expression of FZD6 might be a novel poor prognostic biomarker for AML treatment.

The next-generation antiandrogen drugs such as enzalutamide and abiraterone extend survival times and improve quality of life in patients with advanced prostate cancer. However, resistance to both drugs occurs frequently through mechanisms that are incompletely understood. Wnt signaling, particularly through Wnt5a, plays vital roles in promoting prostate cancer progression and induction of resistance to enzalutamide and abiraterone. Development of novel strategies targeting Wnt5a to overcome resistance is an urgent need. In this study, we demonstrated that Wnt5a/FZD2-mediated noncanonical Wnt pathway is overexpressed in enzalutamide-resistant prostate cancer. In patient databases, both the levels of Wnt5a and FZD2 expression are upregulated upon the development of enzalutamide resistance and correlate with higher Gleason score, biochemical recurrence, and metastatic status, and with shortened disease-free survival duration. Blocking Wnt5a/FZD2 signal transduction not only diminished the activation of noncanonical Wnt signaling pathway, but also suppressed the constitutively activated androgen receptor (AR) and AR variants. Furthermore, we developed a novel bioengineered BERA-Wnt5a siRNA construct and demonstrated that inhibition of Wnt5a expression by the BERA-Wnt5a siRNA significantly suppressed tumor growth and enhanced enzalutamide treatment in vivo. These results indicate that Wnt5a/FZD2 signal pathway plays a critical role in promoting enzalutamide resistance, and targeting this pathway by BERA-Wnt5a siRNA can be developed as a potential therapy to treat advanced prostate cancer.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease with unknown etiology, characterized by motor neuron degeneration, and there is no highly effective treatment. The canonical WNT/β-catenin signaling pathway has a critical role in the physiological and pathophysiological processes of the central nervous system. In this study, we investigated the regulatory mechanism of the WNT/β-catenin signaling pathway from the perspective of ligand-receptor binding and its relationship with the degeneration of ALS motor neurons. We used hSOD1-G93A mutant ALS transgenic mice and hSOD1-G93A mutant NSC34 cells combined with morphological and molecular biology techniques to determine the role of the WNT/β-catenin pathway in ALS. Our findings demonstrated that WNT5A regulates the WNT/β-catenin signaling pathway by binding to the FZD4 receptor in the pathogenesis of ALS and affects the proliferation and apoptosis of ALS motor neurons. Therefore, these findings may lead to the development of novel therapies to support the survival of ALS motor neurons.

Abstract Wnt5a triggers inflammatory responses and damage via NFkB/p65 in retinal pigment epithelial (RPE) cells undergoing uncompensated oxidative stress (UOS) and in experimental ischemic stroke. We found that Wnt5a-Clathrin-mediated uptake leads to NFkB/p65 activation and that Wnt5a is secreted in an exosome-independent fashion. We uncovered that docosahexaenoic acid (DHA) and its derivative, Neuroprotectin D1 (NPD1), upregulate c-Rel expression that, as a result, blunts Wnt5a abundance by competing with NFkB/p65 on the Wnt5a promoter A. Wnt5a increases in ischemic stroke penumbra and blood, while DHA reduces Wnt5a abundance with concomitant neuroprotection. Peptide inhibitor of Wnt5a binding, Box5, is also neuroprotective. DHA-decreased Wnt5a expression is concurrent with a drop in NFkB-driven inflammatory cytokine expression, revealing mechanisms after stroke, as in RPE cells exposed to UOS. Limiting the Wnt5a activity via Box5 reduces stroke size, suggesting neuroprotection pertinent to onset and progression of retinal degenerations and stroke consequences. Graphical Abstract NPD1 disrupts Wnt5a feedback loop at two sites: (1) decreasing FZD5, thus Wnt5a internalization, and (2) by enhancing cREL activity, which competes with p65/NFkB downstream endocytosis. As a result, Wnt5a expression is reduced, and so is its inflammatory signaling in RPE cells and neurons in ischemic stroke.

The identification of characteristic differences between cancer stem cells and their normal counterparts remains a key challenge for cancer treatment. Here, we investigated the role of immunoglobulin superfamily member 8 (Igsf8, also known as EWI-2, PGRL, and CD316) on normal and malignant hematopoietic stem cells, mainly using the conditional knockout model. Deletion of Igsf8 did not affect steady state hematopoiesis, but it led to a significant improvement of survival in mouse myeloid leukemia models. Deletion of Igsf8 significantly depletes leukemia stem cells (LSCs) through enhanced apoptosis and β-catenin degradation. At a molecular level, we found that activation of β-catenin in LSCs depends on Igsf8, which promotes the association of FZD4 with its co-receptor LRP6 in the presence of Igsf8. Similarly, IGSF8 inhibition blocks the colony-forming ability of LSCs and improves the survival of recipients in xenograft models of myeloid leukemia. Collectively, these data indicate strong genetic evidence identifying Igsf8 as a key regulator of myeloid leukemia and the possibility that targeting IGSF8 may serve as a new therapeutic approach against myeloid leukemia.

Prostate cancer (PCa) causes significant mortality and morbidity, with advanced metastasis. WNT signaling is a promising therapeutic target for metastatic PCa. GIPC2 is a GIPC1 paralog involved in WNT signaling pathways associated with tumor progression, but its role in PCa metastasis remains unclear. Herein, we demonstrated that high GIPC2 expression in PCa tissues was significantly associated with distant metastasis and poor prognosis. Functional studies demonstrated that high GIPC2 expression due to CpG-island demethylation promoted increased metastatic capabilities of PCa cells. Conversely, silencing GIPC2 expression significantly inhibited PCa metastasis in vitro and in vivo. Furthermore, GIPC2 directly bound the WNT co-receptor Fzd7 through its PDZ domain, which enabled activation of WNT-β-catenin cascades, thereby stimulating PCa metastasis. Interestingly, GIPC2 protein was also identified as a component of exosomes and that it robustly stimulated PCa adhesion, invasion, and migration. The presence of GIPC2 in tumor-derived exosomes and ability to impact the behavior of tumor cells suggest that GIPC2 is a novel epigenetic oncogene involved in PCa metastasis. Our findings identified GIPC2 as a novel exosomal molecule associated with WNT signaling and may represent a potential therapeutic target and biomarker for metastatic PCa.

We aimed to detect the expression pattern of long non-coding RNA (lncRNA) FGD5-AS1 in gastric cancer (GC) samples and its impact on driving the development of GC. FGD5-AS1 levels in 66 cases of GC tissues and paracancerous ones were detected. Its influences on clinical features and prognosis in GC patients were analyzed. In AGS and SGC-7901 cells with FGD5-AS1 knockdown, phenotype changes were assessed through cell counting kit-8 (CCK-8), Transwell and wound healing assay. The downstream target of FGD5-AS1 was searched by a bioinformatics tool and confirmed by dual-luciferase reporter assay. Their interaction in regulating the malignant development of GC was finally explored. FGD5-AS1 was upregulated in GC tissues compared to paracancerous ones. GC patients expressing a high level of FGD5-AS1 had higher risk of lymphatic metastasis or distant metastasis and worse prognosis than those with a low level. Knockdown of FGD5-AS1 weakened proliferative and metastatic abilities in AGS and SGC-7901 cells. FZD3 was the downstream target of FGD5-AS1. Protein levels of FZD3 and FZD5 were upregulated, while b-catenin, TGF-b and MMP9 were downregulated in GC cells with FGD5-AS1 knockdown. Knockdown of FZD3 abolished the regulatory effects of FGD5-AS1 on malignant phenotypes of GC cells. FGD5-AS1 is upregulated in GC samples, which is linked to metastasis and prognosis in GC. It drives proliferative and metastatic abilities in GC cells via negatively interacting with FZD3.

Background Retinopathy of prematurity (ROP) is a multifactorial retinal disease, involving both environmental and genetic factors; The purpose of this study is to evaluate the clinical presentations and genetic variants in Chinese patients with ROP. Methods A total of 36 patients diagnosed with ROP were enrolled in this study, their medical and ophthalmic histories were obtained, and comprehensive clinical examinations were performed. Genomic DNA was isolated from peripheral blood of ROP patients, polymerase chain reaction and direct sequencing of the associated pathogenic genes (FZD4, TSPAN12, and NDP) were performed. Results All patients exhibited the clinical manifestations of ROP. No mutations were detected in the TSPAN12 and NDP genes in all patients; Interestingly, three novel missense mutations were identified in the FZD4 gene (p.A2P, p.L79M, and p.Y378C) in four patients, for a detection rate of 11.1% (4/36). Conclusions This study expands the genotypic spectrum of FZD4 gene in ROP patients, and our findings underscore the importance of obtaining molecular analyses and comprehensive health screening for this retinal disease.

Background Invasive malignant pleomorphic adenoma (IMPA) is a highly malignant neoplasm of the oral salivary glands with a poor prognosis and a considerable risk of recurrence. Many disease-causing genes of IMPA have been identified in recent decades (e.g., P53, PCNA and HMGA2), but many of these genes remain to be explored. Weighted gene coexpression network analysis (WGCNA) is a newly emerged algorithm that can cluster genes and form modules based on similar gene expression patterns. This study constructed a gene coexpression network of IMPA via WGCNA and then carried out multifaceted analysis to identify novel disease-causing genes. Methods RNA sequencing (RNA-seq) was performed for 10 pairs of IMPA and normal tissues to acquire the gene expression profiles. Differentially expressed genes (DEGs) were screened out with the cutoff criteria of |log2 Fold change (FC)|> 1 and adjusted p value Results In this research, a total of 1970 DEGs were screened out in IMPA tissues, including 1056 upregulated DEGs and 914 downregulated DEGs. Functional enrichment analysis was performed for identified DEGs and revealed an enrichment of tumor-associated GO terms and KEGG pathways. We used WGCNA to identify gene module most relevant with the histological grade of IMPA. The gene FZD2 was then recognized as the hub gene of the selected module with the highest module membership (MM) value and intramodule connectivity in protein–protein interaction (PPI) network. According to immunohistochemistry (IHC) staining, the expression level of FZD2 was higher in low-grade IMPA than in high-grade IMPA. Conclusion FZD2 shows an expression dynamic that is negatively correlated with the clinical malignancy of IMPA and it plays a central role in the transcription network of IMPA. Thus, FZD2 serves as a promising histological indicator for the precise prediction of IMPA histological stages.

The organismal body axes that are formed during embryogenesis are intimately linked to intrinsic asymmetries established at the cellular scale in oocytes (1). However, the mechanisms that generate cellular asymmetries within the oocyte, and then transduce that polarity to organismal scale body axes are poorly understood outside of select model organisms. Here, we report an axis-defining event in meiotic oocytes of the sea star Patiria miniata. Dishevelled is a cytoplasmic Wnt pathway effector required for axis development in diverse species (2–4), but the mechanisms governing its function and distribution remain poorly defined. Using time-lapse imaging, we find that Dishevelled localizes uniformly to puncta throughout the cell cortex in Prophase I-arrested oocytes, but becomes enriched at the vegetal pole following meiotic resumption through a dissolution-reassembly mechanism. This process is driven by an initial disassembly phase of Dvl puncta, followed by selective reformation of Dvl assemblies at the vegetal pole. Rather than being driven by Wnt signaling, this localization behavior is coupled to meiotic cell cycle progression and influenced by Lamp1+ endosome association and Frizzled receptors pre-localized within the oocyte cortex. Our results reveal a cell cycle-linked mechanism by which maternal cellular polarity is transduced to the embryo through spatially-regulated Dishevelled dynamics.

Background Wnt genes code for ligands that activate signaling pathways during development in Metazoa. Through the canonical Wnt (cWnt) signaling pathway, these genes regulate important processes in bilaterian development, such as establishing the anteroposterior axis and posterior growth. In Arthropoda, Wnt ligands also regulate segment polarity, and outgrowth and patterning of developing appendages. Arthropods are part of a lineage called Panarthropoda that includes Onychophora and Tardigrada. Previous studies revealed potential roles of Wnt genes in regulating posterior growth, segment polarity, and growth and patterning of legs in Onychophora. Unlike most other panarthropods, tardigrades lack posterior growth, but retain segmentation and appendages. Here, we investigated Wnt genes in tardigrades to gain insight into potential roles that these genes play during development of the highly compact and miniaturized tardigrade body plan. Results We analyzed published genomes for two representatives of Tardigrada, Hypsibius exemplaris and Ramazzottius varieornatus. We identified single orthologs of Wnt4, Wnt5, Wnt9, Wnt11, and WntA, as well as two Wnt16 paralogs in both tardigrade genomes. We only found a Wnt2 ortholog in H. exemplaris. We could not identify orthologs of Wnt1, Wnt6, Wnt7, Wnt8, or Wnt10. We identified most other components of cWnt signaling in both tardigrade genomes. However, we were unable to identify an ortholog of arrow/Lrp5/6, a gene that codes for a Frizzled co-receptor of Wnt ligands. Additionally, we found that some other animals that have lost several Wnt genes and are secondarily miniaturized, like tardigrades, are also missing an ortholog of arrow/Lrp5/6. We analyzed the embryonic expression patterns of Wnt genes in H. exemplaris during developmental stages that span the establishment of the AP axis through segmentation and leg development. We detected expression of all Wnt genes in H. exemplaris besides one of the Wnt16 paralogs. During embryo elongation, expression of several Wnt genes was restricted to the posterior pole or a region between the anterior and posterior poles. Wnt genes were expressed in distinct patterns during segmentation and development of legs in H. exemplaris, rather than in broadly overlapping patterns. Conclusions Our results indicate that Wnt signaling has been highly modified in Tardigrada. While most components of cWnt signaling are conserved in tardigrades, we conclude that tardigrades have lost Wnt1, Wnt6, Wnt7, Wnt8, and Wnt10, along with arrow/Lrp5/6. Our expression data may indicate a conserved role of Wnt genes in specifying posterior identities during establishment of the AP axis. However, the loss of several Wnt genes and the distinct expression patterns of Wnt genes during segmentation and leg development may indicate that combinatorial interactions among Wnt genes are less important during tardigrade development compared to many other animals. Based on our results, and comparisons to previous studies, we speculate that the loss of several Wnt genes in Tardigrada may be related to a reduced number of cells and simplified development that accompanied miniaturization and anatomical simplification in this lineage.

Frizzled 7 (FZD7) is abnormally expressed and activated in a variety of cancers. In ovarian cancer, overexpression of FZD7 reduces the sensitivity of platinum-resistant ovarian cancer cells to ferroptosis, thereby allowing cancer cells to survive. However, whether FZD7 inhibits ferroptosis in ovarian cancer cells and its mechanisms are remain unclear. This study aims to explore the effects of FZD7 and its upstream regulator miR-1-3p on ferroptosis in ovarian cancer cells are evaluated to clarify the molecular mechanism for miR-1-3p and FZD7's involvement in ferroptosis in ovarian cancer cells.Human ovarian cancer cell lines HO8910 and SKOV3 were used as the research subjects. In the first part of the experiment, human ovarian cancer cells were transfected with blank plasmid and FZD7 overexpression plasmid, respectively; in the second and third parts, human ovarian cancer cells were transfected with miR-1-3p mimics negative control, miR-1-3p mimics, miR-1-3p inhibitors negative control, and miR-1-3p inhibitors, respectively; in the fourth part of the experiment, human ovarian cancer cells were transfected with miR-1-3p mimics and miR-1-3p mimics+FZD7 overexpression plasmid, respectively, and normal cultured cells were set as the control group. The human ovarian cancer cell ferroptosis model was established by incubating human ovarian cancer cells with different treatments with ferroptosis inducer Erastin or RSL3. Real-time RT-PCR was used to detect the mRNA expression levels of FZD7 and miR-1-3p; Western blotting was used to detect the protein expression levels of FZD7; CCK-8 assay was used to detect the cell viability; lipid peroxidation colorimetric assay kit was used to detect the level of intracellular MDA; and iron assay kit was used to detect the level of intracellular Fe2+. Dual-luciferase assay was used to detect the targeting relationship between miR-1-3p and FZD7.Overexpression of FZD7 increased the cell viability of human ovarian cancer cell lines HO8910 or SKOV3 (P0.05, P0.01, or P0.001) and decreased the intracellular MDA levels (P0.01) in Erastin-treated or RSL3-treated ovarian cancer cells. FZD7 was a direct target of miR-1-3p, which inhibited the expression of FZD7 (P0.01) by binding to the 3'-untranslated region (3'UTR) site of FZD7. MiR-1-3p mimics decreased the cell viability of human ovarian cancer cell lines HO8910 or SKOV3 (P0.05, P0.01, or P0.001) and increased the intracellular MDA levels (P0.01) in Erastin-treated or RSL3-treated ovarian cancer cells; while miR-1-3p inhibitors significantly increased the cell viability of human ovarian cancer cell lines HO8910 or SKOV3 (P0.05, P0.01, or P0.001) and decreased the intracellular MDA levels (P0.01) in Erastin-treated or RSL3-treated ovarian cancer cells. The effect of miR-1-3p mimics on enhancing the sensitivity of human ovarian cancer cells to Erastin-induced or RSL3-induced ferroptosis was abrogated by overexpression of FZD7(P0.05 or P0.01).MiR-1-3p enhances the sensitivity of ovarian cancer cells to ferroptosis by targeting FZD7.

Introduction Accumulating evidence highlights the critical roles of circular RNAs (circRNAs) in the malignant progression of cancers. In this study, we investigated the expression pattern of a newly identified circRNA (hsa_circ_0017109) in non–small cell lung cancer (NSCLC), and examined its downstream molecular targets. Methods Quantitative real-time PCR (qRT-PCR) and Western blotting (WB) were conducted to quantify gene and protein expression. In vitro functional assays such as colony formation assay, cell counting kit-8 (CCK-8) and flow cytometry were used to study cell proliferation and apoptosis. RNA pull-down assay, luciferase reporter assay and RNA immunoprecipitation were performed to validate molecular interaction. Mouse xenograft model of NSCLC cells was used to assess the role of circ_0017109 in tumorigenesis. Results Circ_0017109 was upregulated in NSCLC tumor samples and cells. Silencing circ_0017109 impaired cell proliferation and promoted apoptosis in NSCLC cells, and circ_0017109 knockdown suppressed in vivo tumorigenesis of NSCLC cells in mouse xenograft model. MiR-671-5p was identified as a target of circ_0017109, and circ_0017109 negatively impacted on miR-671-5p expression. MiR-671-5p downregulated FZD4 and dampened the activity of Wnt/β-catenin signaling pathway. Circ_0017109 modulated FZD4 expression by suppressing miR-671-5p activity. Conclusions Elevated circ_0017109 expression promotes tumor progression of NSCLC by modulating miR-671-5p/FZD4/β-catenin axis.

OBJECTIVE: To investigate the effect of exosomal FZD10 derived from non-small cell lung cancer (NSCLC) cells on angiogenesis of human umbilical venous endothelial cells (HUVECs) and explore the possible mechanism. METHODS: We analyzed the expression of FZD10 in two NSCLC cell lines (95D and H1299 cells), normal human bronchial epithelial cells (BEAS-2B cells) and their exosomes isolated by ultracentrifugation. Cultured HUVECs were treated with the exosomes derived from NSCLC cells or NSCLC cells transfected with FZD10-siRNA, and the changes in tube formation ability of the cells were analyzed using an in vitro angiogenesis assay. ELISA was performed to determine the concentration of VEGFA and Ang-1 in the conditioned media of HUVECs, and RT-qPCR was used to analyze the mRNA levels of VEGFA and Ang-1 in the HUVECs. The effects of exosomal FZD10 on the activation of PI3K, Erk1/2 and YAP/TAZ signaling pathways were evaluated using Western blotting. RESULTS: Compared with BEAS-2B cells and their exosomes, 95D and H1299 cells and their exosomes all expressed high levels of FZD10 (P < 0.01). The exosomes derived from 95D and H1299 cells significantly enhanced tube formation ability and increased the expressions of VEGFA and Ang-1 protein and mRNA in HUVECs (P < 0.01), but FZD10 knockdown in 95D and H1299 cells obviously inhibited these effects of the exosomes. Exosomal FZD10 knockdown suppressed the activation of PI3K and Erk1/2 signaling pathways, but had no obvious effect on the activation of YAP/TAZ signaling pathway. CONCLUSION: Exosomal FZD10 derived from NSCLC cells promotes HUVEC angiogenesis in vitro, the mechanism of which may involve the activation of PI3K and Erk1/2 signaling pathways.

The bacteriocin, nisin, produced by Lactococcus and Streptococcus species during fermentation, is widely used for bio preservatives in a wide variety of foods. Liver cancer has a high mortality rate and is the fourth leading cause of cancer-related deaths worldwide. Recently, researchers have shown the anti-cancer effects of nisin through in vitro and in vivo studies. This study aimed to investigate the effect of nisin on liver cancer cell lines, which represented two subgroups of the disease model. Nisin exhibited significant growth inhibition and apoptosis in both cell lines, HuH-7, and SNU182. Drug resistance is the main problem in liver cancer and the epithelial-to-mesenchymal transition has a role in the development of drug resistance in hepatocellular carcinoma. The expression of EMT transcription factors ZEB1, SNAI1, and TWIST1 were analyzed depending on nisin treatment, TWIST1 expression was down-regulated after nisin treatment compared to the untreated SNU182 and HuH-7 cell lines. Besides, due to the reported correlation between the overexpression of Frizzled (FZD) proteins, specifically FZD7, in primary hepatocellular carcinomas (HCCs), molecular docking was assessed for Nisin A in the binding site of FZD7. Results confirmed that Nisin A was able to form important hydrogen bonding with key residues. This research not only determined the role of nisin in different liver cancer cell models but it also provided the first result of FZD7 and nisin interaction.

Signal peptide (SP) mutations are an infrequent cause of inherited retinal diseases (IRDs). We report the genes currently associated with an IRD that possess an SP sequence and assess the prevalence of these variants in a multi-institutional retrospective review of clinical genetic testing records. The online databases, RetNet and UniProt, were used to determine which IRD genes possess a SP. A multicenter retrospective review was performed to retrieve cases of patients with a confirmed diagnosis of an IRD and a concurrent SP variant. In silico evaluations were performed with MutPred, MutationTaster, and the signal peptide prediction tool, SignalP 6.0. SignalP 6.0 was further used to determine the locations of the three SP regions in each gene: the N-terminal region, hydrophobic core, and C-terminal region. Fifty-six (56) genes currently associated with an IRD possess a SP sequence. Based on the records review, a total of 505 variants were present in the 56 SP-possessing genes. Six (1.18%) of these variants were within the SP sequence and likely associated with the patients’ disease based on in silico predictions and clinical correlation. These six SP variants were in the CRB1 (early-onset retinal dystrophy), NDP (familial exudative vitreoretinopathy) (FEVR), FZD4 (FEVR), EYS (retinitis pigmentosa), and RS1 (X-linked juvenile retinoschisis) genes. It is important to be aware of SP mutations as an exceedingly rare cause of IRDs. Future studies will help refine our understanding of their role in each disease process and assess therapeutic approaches.

Wnt1 is the first mammalian Wnt gene, which is discovered as proto-oncogene and in human the gene is located on the chromosome 12q13. Mutations in Wnt1 are reported to be associated with various cancers and other human diseases. The structural and functional consequences of most of the non-synonymous SNPs (nsSNPs), present in the human Wnt1 gene, are not known. In the present work, extensive bioinformatics analyses are used to screen 292 nsSNPs of Wnt1 for predicting pathogenic and harmless polymorphisms. We have identified 10 highly deleterious nsSNPs among which 7 are located within the highly conserved areas. These 10 nsSNPs are also predicted to affect the post-translational modifications of Wnt1. Further, structure based stability analyses of these 10 highly deleterious nsSNPs revealed 8 variants as highly destabilizing. These 8 highly destabilizing variants were shown to have high BC score and high RMSIP score from normal mode analyses. Based on the deformation energies, obtained from the normal mode analyses, variants like G169A, G169S, G331R and G331S were found to be unstable. Molecular Dynamics (MD) simulations revealed structural stability and fluctuation of WT Wnt1 and its prioritized variants. RMSD remained fluctuating mostly between 4 and 5 Å and occasionally between 3.5 and 5.5 Å ranges. RMSF in the CTD region (residues 330–360) of the binding pocket were lower compared to that of WT. Studying the impacts of nsSNPs on the binding interface of Wnt1 and seven Frizzled receptors have predicted substitutions which can stabilize or destabilize the binding interface. We have found that Wnt1 and FZD8-CRD is the best docked complex in our study. MD simulation based analyses of wild type Wnt1-FZD8-CRD complex and the 8 prioritized variants revealed that RMSF was higher in the unstructured regions and RMSD remained fluctuating in the region of 5 Å ± 1 Å. We have also observed differential Wnt1 gene expression pattern in normal, tumor and metastatic conditions across different tissues. Wnt1 gene expression was significantly higher in metastatic tissues of lungs, colon and skin; and was significantly lower in metastatic tissues of breast, esophagus and kidney. We have also found that Wnt1 deregulation is associated with survival outcome in patients with gastric and breast cancer. Furthermore, these computationally screened highly deleterious nsSNPs of Wnt1 can be analyzed in population based genetic studies and may help understand the Wnt1 associated diseases.

In the Wnt–β-catenin pathway, Wnt binding to Frizzled (Fzd) and LRP5 or LRP6 (LRP5/6) co-receptors inhibits the degradation of the transcriptional coactivator β-catenin by recruiting the cytosolic effector Dishevelled (Dvl). Polymerization of Dvl at the plasma membrane recruits the β-catenin destruction complex, enabling the phosphorylation of LRP5/6, a key step in inhibiting β-catenin degradation. Using purified Fzd proteins reconstituted in lipid nanodiscs, we investigated the factors that promote the recruitment of Dvl to the plasma membrane. We found that the affinity of Fzd for Dvl was not affected by Wnt ligands, in contrast to other members of the GPCR superfamily for which the binding of extracellular ligands affects the affinity for downstream transducers. Instead, Fzd-Dvl binding was enhanced by increased concentration of the lipid PI(4,5)P 2 , which is generated by Dvl-associated lipid kinases in response to Wnt and which is required for LRP5/6 phosphorylation. Moreover, binding to Fzd did not promote Dvl DEP domain dimerization, which has been proposed to be required for signaling downstream of Fzd. Our findings suggest a positive feedback loop in which Wnt-stimulated local PI(4,5)P 2 production enhances Dvl recruitment and further PI(4,5)P 2 production to support Dvl polymerization, LRP5/6 phosphorylation, and β-catenin stabilization.

Female breast cancer (BC) is the leading cause of cancer-related deaths worldwide with higher mortality rates and early onset in developing countries. The molecular basis of early disease onset is still elusive. We recruited 472 female breast cancer from two sub-Saharan African countries (Cameroon and Congo) between 2007 and 2018 and collected clinical data from these patients. To investigate the molecular drivers of early disease onset, we analyzed publicly available breast cancer molecular data from the cancer genome atlas (TCGA) and the gene expression omnibus (GEO) for copy number alteration, mutation and gene expression. Early BC onset (EOBRCA) (diagnosis before 45 years) was higher in African women compared with the TCGA cohort (51.7% vs 15.6%). The tumor grade, mitotic index, HER2 + phenotype, basal-like phenotype and ki67 were higher in EOBRCA for all cohorts. BC risk factors such as parity, breastfeeding early onset of menarche and use of hormonal contraceptives were significantly associated with EOBRCA (p CDH6 and FOXM1 and tumor suppressor including TGM3 and DMBT1 as well as higher TP53 mutation rates (OR: 2.93, p TGM3 deletions, which was associated with high expression of all SMAD transcription factors as well as WNT ligands. The Frizzled receptors FZD1, FZD4 and FZD6 were significantly upregulated in EOBRCA, suggesting activation of non-canonical WNT signaling. Our data, suggest the implication of TGM3 deletion in early breast cancer onset. Further molecular investigations are warranted in African patients.

ImportanceFamilial exudative vitreoretinopathy (FEVR) is a nonsyndromic autosomal dominant retinal disorder commonly caused by variants in the FZD4 gene. This study investigates the potential role beyond ocular abnormalities for FZD4 gene variants in patients with FEVR.ObjectiveTo evaluate the role of FZD4 in symptoms beyond those associated with FEVR through a patient with biallelic variants in FZD4.Design, Setting, and ParticipantsThis case series included the DNA testing and phenotyping of 1 patient proband and her parents, combined with signaling assays, to determine the association of patient-derived compound heterozygous variants on FZD4 signaling and biologic function.Main Outcomes and MeasuresFZD4 genes were tested using next-generation sequencing and Sanger sequencing. Cell-based assays measured the effect of the variants on FZD4 signaling.ResultsThe proband presented with absent red reflexes from complete tractional retinal detachments diagnosed at 3 days of age and failed the newborn screening hearing test. Auditory brainstem response at 6 months of age showed bilateral mild to moderate high-frequency sensorineural hearing loss. The patient manifested developmental delays in speech and walking. Intravenous fluorescein angiography (IVFA) of the patient’s parents detected stage 1 FEVR. Genetic testing revealed 2 FZD4 variants in the patient, each variant found in 1 parent. Signaling assays confirmed that the presence of both variants was associated with significantly worse signaling activity compared with the heterozygous state.Conclusions and RelevanceResults of this case series suggest that extraocular syndromic FEVR was associated with FZD4 variants. The decrease in FZD4 signaling owing to the biallelic nature of the disease resulted in hearing deficits, developmental delays, and a more severe retinal phenotype.

To expand the mutation spectrum of patients with familial exudative vitreoretinopathy (FEVR) disease.74 probands (53 families and 21 sporadic probands) with familial exudative vitreoretinopathy (FEVR) disease and their available family members (n = 188) were recruited for sequencing.Panel-based targeted screening was performed on all subjects. Before sanger sequencing, variants of LRP5, NDP, FZD4, TSPAN12, ZNF408, KIF11, RCBTB1, JAG1, and CTNNA1 genes were verified by a series of bioinformatics tools and genotype-phenotype co-segregation analysis.40.54% (30/74) of the probands were sighted to possess at least one etiological mutation of the nine FEVR-causative genes. The etiological mutation detection rate was 37.74% (20/53) in family-attainable probands while 47.62% (10/21) in sporadic cases. The diagnosis rate of patients in the early-onset subgroup (≤5 years old, 45.4%) is higher than that of the children or adolescence-onset subgroup (6-16 years old, 42.1%) and the late-onset subgroup (≥17 years old, 39.4%). A total of 36 etiological mutations were identified in this study, comprising 26 novel mutations and 10 reported mutations. LRP5 was the most prevalent mutant gene among the 36 mutation types with a percentage of 41.67% (15/36). Followed by FZD4 (10/36, 27.78%), TSPAN12 (5/36, 13.89%), NDP (4/36, 11.11%), KIF11 (1/36, 2.78%), and RCBTB1 (1/36, 2.78%). Among these mutations, 63.89% (23/36) were missense mutations, 25.00% (9/36) were frameshift mutations, 5.56% (2/36) were splicing mutations, 5.56% (2/36) were nonsense mutations. Moreover, the clinical pathogenicity of these variants was defined according to American College of Medical Genetics (ACMG) and genomics guidelines: 41.67% (15/36) were likely pathogenic variants, 27.78% (10/36) pathogenic variants, 30.55% (11/36) variants of uncertain significance. No etiological mutations discovered in the ZNF408, JAG1, and CTNNA1 genes in this FEVR cohort.We systematically screened nine FEVR disease-associated genes in a cohort of 74 Chinese probands with FEVR disease. With a detection rate of 40.54%, 36 etiological mutations of six genes were authenticated in 30 probands, including 26 novel mutations and 10 reported mutations. The most prevalent mutated gene is LRP5, followed by FZD4, TSPAN12, NDP, KIF11, and RCBTB1. In total, a de novo mutation was confirmed. Our study significantly clarified the mutation spectrum of variants bounded up to FEVR disease.

Dishevelled is a cytoplasmic hub that transduces Wnt signals to cytoplasmic effectors, which can be broadly characterised as canonical (β-catenin dependent) and noncanonical, to specify cell fates and behaviours during development. To transduce canonical Wnt signals, Dishevelled binds to the intracellular face of Frizzled through its DEP domain and polymerises through its DIX domain to assemble dynamic signalosomes. Dishevelled also contains a PDZ domain, whose function remains controversial. Here, we use genome editing to delete the PDZ domain-encoding region from Drosophila dishevelled. Canonical Wingless signalling is entirely normal in these deletion mutants; however, they show defects in multiple contexts controlled by noncanonical Wnt signalling, such as planar polarity. We use nuclear magnetic resonance spectroscopy to identify bona fide PDZ-binding motifs at the C termini of different polarity proteins. Although deletions of these motifs proved aphenotypic in adults, we detected changes in the proximodistal distribution of the polarity protein Flamingo (also known as Starry night) in pupal wings that suggest a modulatory role of these motifs in polarity signalling. We also provide new genetic evidence that planar polarity relies on the DEP-dependent recruitment of Dishevelled to the plasma membrane by Frizzled.

RING finger protein 43 (RNF43) encodes the transmembrane E3 ubiquitin ligase, which targets the Wnt receptor Frizzled (FZD). RNF43 mutations have been discovered in various human cancers including colon, pancreatic, stomach, ovarian, and liver cancers. Functional studies on RNF43 missense mutations have shown that they negatively regulate Wnt signaling; however, there are few functional studies on RNF43 frameshift mutations. In this study, we showed that R117fs and P441fs mutants enhanced Wnt/β-catenin signaling, whereas Q409fs and G659fs mutants retained the ability to suppress Wnt/β-catenin signaling. Specifically, R117fs was unable to ubiquitinate FZD5 due to lack of the RING domain, although it was able to interact with FZD5. Immunofluorescence showed that R117fs failed to internalize FZD5 expressed on the cell surface. We also showed that LGK974, a potent Wnt inhibitor, decreased the Wnt/β-catenin activity by R117fs and P441fs mutations. Together, these results demonstrate that RNF43 frameshift mutations retain normal functionality; thus, targeted anti-cancer therapy can be developed according to the mutation type of RNF43.

The purpose of this study was to establish a genotype-phenotype correlation of familial exudative vitreoretinopathy (FEVR) caused by FZD4 gene mutations.Six hundred fifty-one probands and their family members were recruited based on a clinical diagnosis of FEVR between 2015 and 2021 at Zhongshan Ophthalmic Center. Ocular examinations were performed in all participants. Targeted gene panel sequencing and whole-exome sequencing were performed in the probands, and Sanger sequencing was used to verify the mutations and segregation analysis was performed in the family members.Fifty-one FZD4 mutations (24 novels and 27 known) were detected in 84 families. Of these 168 eyes with FEVR, the eyes at stages 1, 2, 3, 4, and 5 were 29 (17.3%), 15 (8.9%), 19 (11.3%), 55 (32.7%), and 12 (7.1%), respectively. Exact stage of 38 (22.6%) eyes could not be determined. The FEVR phenotypes were more severe in the probands than the phenotypes in the family members (P0.001). The families were divided into two groups, probands that inherited the variant from the mother, and probands that inherited the variant from the father. In addition, the FEVR stage differences between these two groups were different (P0.05). Despite the mutations being located in different domains of FZD4, no significant differences were identified among the domains in terms of FEVR staging, retinal folds, retinal detachment, temporal midperipheral vitreoretinal interface abnormality, and foveal hypoplasia.The FZD4 probands had severer phenotype than the family members, and the FEVR stage difference was greater between the probands and mothers than that between the probands and fathers.

Genes associated with specific neurocognitive phenotypes in Williams–Beuren syndrome are still controversially discussed. This study identified nine patients with atypical deletions out of 111 patients with Williams–Beuren syndrome; these deletions included seven smaller deletions and two larger deletions. One patient had normal neurodevelopment with a deletion of genes on the distal side of the Williams–Beuren syndrome chromosomal region, including GTF2I and GTF2IRD1. However, another patient retained these genes but showed neurodevelopmental abnormalities. By comparing the genotypes and phenotypes of patients with typical and atypical deletions and previous reports in the literature, we hypothesize that the BAZ1B, FZD9, and STX1A genes may play an important role in the neurodevelopment of patients with WBS.

SummaryPolycystic ovarian syndrome (PCOS) is a chronic hormonal turmoil that is demonstrated in 2.2−27% of women of pre-menopausal age. This disease is a complex multigenic disorder that results from the interaction between excess androgen expression, genetic susceptibility and environmental influences. PCOS is associated with 40% of female infertility and endometrial cancer. The WNT/β-catenin signalling transduction pathway regulates aspects of cell proliferation, migration and cell fate determination in the tissue along with early embryonic development and controls the proper activation of the female reproductive system, along with regulating hormonal activity in ovarian granulosa cells. In the current study, we investigated the expression profiles of WNT/β-catenin signalling pathway genes (AXIN2, FZD4, TCF4, WNT3, WNT4, WNT5A, WNT7A, WNT1, APC, GSK3B and β-catenin) in a total of 13 oocyte samples. Seven of these samples were from polycystic women and six were from healthy women. The results of this study displayed the absence of expression of AXIN2, FZD4, TCF4, WNT5A, WNT3, WNT4 and WNT7A genes in ovaries from women with PCOS and from healthy women. While APC and β-catenin expression levels were similar in the oocytes of both patients and controls, conversely, WNT1 and GSK3β genes both showed elevated expression in the oocytes of patients with PCOS, therefore suggesting an association between aberrant expression of WNT1 and GSK3β and the pathogenesis of PCOS. The observations of the current study could be helpful to provide evidence regarding the pathogenesis of PCOS and its treatment.

The function and pathological significance of circular RNAs (circRNAs) in autoimmune diseases, such as rheumatoid arthritis (RA), are barely known. Here, we explored the role of circ_0088036 in RA progression and its associated mechanism.The synovial lining layer tissues of RA patients and non-RA control patients were collected for clinical studyCirc_0088036 expression was elevated in the synovial lining layer tissues of RA patients and TNF-α-treated RA-FLSs. Circ_0088036 interference largely reversed TNF-α-induced proliferation and inflammation in RA-FLSs. The interaction between circ_0088036 and miR-326 was verified, and miR-326 silencing largely reversed circ_0088036 knockdown-mediated effects in TNF-α-treated RA-FLSs. MiR-326 bound to the 3' untranslated region (3'UTR) of FZD4 in RA-FLSs. FZD4 overexpression largely diminished miR-326 accumulation-mediated influences in TNF-α-treated RA-FLSs. Circ_0088036 could up-regulate FZD4 by sponging miR-326 in RA-FLSs.Circ_0088036 contributed to TNF-α-induced RA progression partly by targeting miR-326/FZD4 signalling.

Background Metastasis of malignant tumors accelerates systemic failure and hastens the deaths of pancreatic cancer patients. During the metastatic process, the physical translocation of cancer cells from the primary lesion to distant organs and is crucial. CSCs properties, such as self-renewal and multiple-direction differentiation capacity are essential for colonization in the microenvironment of distant organs and metastatic lesion formation. It is widely believed that EMT can cause cancer cells to penetrate blood vessels by undergoing phenotypic and cytoskeletal changes, so that they can infiltrate surrounding tissue and disseminate from the primary tumor to the blood circulation, where they are termed circulating tumor cells (CTCs), while CTCs often exhibit stemness properties. Accumulating evidence demonstrates that some EMT-related transcription factors are essential for CSCs self-renewal, so cancer cells that have undergone EMT typically acquire increased stemness properties. Abnormal activation of the WNT signaling pathway can drive a series of gene transcripts to promote EMT in multiple types of cancer, and among different Frizzled receptors of WNT signaling pathway, FZD7 expression is associated with distant organ metastasis, advanced clinical stages, and poor clinical prognosis. Objective of this study is to demonstrate that high FZD7 expression in pancreatic cancer can accelerate hepatic metastases and elucidate the related molecular mechanisms. Methods The expression of Frrizled receptor 7 (FZD7) in pancreatic ductal adenocarcinoma (PDAC) and relating survival rate were analyzed by bioinformatics, histochemistry assay and follow-up study. In vitro, FZD7 expression was silenced by lentiviral vectors carrying short hair RNA (shRNA) or upregulated by overexpression plasmid. Then, Wound-healing and Transwell experiment was used to analyze the abilities of migration and invasion; the levels of epithelial-to-mesenchymal transition (EMT) relating phenotype proteins, stemness relating phenotype proteins, and signaling molecular proteins were measured by Western-blot; cell stemness was evaluated by sphere forming ability of cells in suspension culture and detecting the proportion of CD24+CD44+ cells with flow cytometry. TGF-β1 was used to induce EMT, and observe the effect of shRNA silencing FZD7 on which. Results High level of FZD7 expression in pancreatic cancer samples was associated with earlier hepatic metastasis. In vitro upregulation FZD7 can enable pancreatic cancer cells to obtain stronger migration and invasion ability and higher mesenchymal phenotype, and vice versa; the proportion of cancer stem cell (CSC) was also positively correlated with the level of FZD7; cells forming spheres in suspension culture showed stronger migration and invasion ability and higher level of mesenchymal phenotype than normal adherent cultured cells; the level of FZD7 was positively correlated with the level of activated β-catenin. Silencing FZD7 expression can attenuate EMT induced by TGF-β1 stimulating, and TGF-β1 stimulating can also upregulate stemness phenotype expression, such as ABCG2, CD24, and CD44 by mediating of FZD7. Conclusions High FZD7 expression in pancreatic cancer can accelerates hepatic metastases by promoting EMT and strengthening cell stemness, and FZD7 can work through the canonical Wingless-type (WNT) signaling pathway and participate in TGF-β/SMAD3 signaling pathway also.

This study aimed to report the frequency ofGenetic data collected from 696 FEVR families were reviewed. The ocular phenotypes in patients withIn a cohort of 696 FEVR families, disease-causingThe frequency of the

Precise Norrin and β-catenin (Norrin/β-catenin; encoded by NDP and CTNNB1, respectively) signaling is critical for proper angiogenesis. Dysregulation of this signaling leads to various diseases, of which retinal exudative vitreoretinopathy is the most prevalent. Here, we used a global knockout mouse model to show that limb development membrane protein 1 like (LMBR1L), a transmembrane protein of unknown function in angiogenesis, is essential for retinal vascular development. In vitro experiments revealed that LMBR1L depletion results in aberrant activation of the Norrin/β-catenin signaling pathway via decreased ubiquitylation of FZD4 and increased Norrin co-receptor LRP5 and p-GSK3β-Ser9 expression levels, which cause accumulation of β-catenin. Moreover, inhibition of LMBR1L in human retinal microvascular endothelial cells (HRECs) caused increased proliferation ability and defective cell migration, which might have occurred as a result of upregulated expression levels of the apical junction components. Treatment with p-GSK3β-Ser9 inhibitor AR-A014418 restored the phenotypes in LMBR1L-null HRECs, which further demonstrated the important regulatory role of LMBR1L in the Norrin/β-catenin signaling pathway. Taken together, our data reveal an essential role for LMBR1L in angiogenesis. This article has an associated First Person interview with the first author of the paper.

While Inherited Retinal Diseases (IRDs) are typically considered rare diseases, Familial Exudative Vitreo-Retinopathy (FEVR) and Norrie Disease (ND) are more rare than retinitis pigmentosa. We wanted to determine if multigenic protein-altering variants are common in FEVR subjects within a set of FEVR-related genes. The potential occurrence of protein-altering variants in two different genes has been documented in a very small percentage of patients, but potential multigenic contributions to FEVR remain unclear. Genes involved in these orphan pediatric retinal diseases are not universally included in available IRD targeted-sequencing panels, and cost is also a factor limiting multigenic-sequence-based testing for these rare conditions. To provide an accurate solution at lower cost, we developed a targeted-sequencing protocol that includes seven genes involved in Familial Exudative Vitreo-Retinopathy (FEVR) and Norrie disease. Seventy-six DNA samples from persons refered to clinic with possible FEVR and some close relatives were sequenced using a novel Oakland-ERI orphan pediatric retinal disease panel (version 2) providing 900 times average read coverage. The seven genes involved in FEVR/ND were: NDP (ChrX), CTNNB1 (Chr3); TSPAN12 (Chr7); KIF11 (Chr10), FZD4 (Chr11), LRP5 (Chr11), ZNF408 (Chr11). A total of 33 variants were found that alter protein sequence, with the following relative distribution: LRP5 13/33 (40%), FZD4 9/33 (27%), ZNF408 6/33 (18%), (KIF11 3/33 (9%), NDP 1/33 (3%), CTNNB1 1/33 (3%). Most protein-altering variants, 85%, were found in three genes: FZD4, LRP5, and ZNF408. Four previously known pathogenic variants were detected in five families and two unrelated individuals. Two novel, likely pathogenic variants were detected in one family (FZD4: Cys450ter), and a likely pathogenic frame shift termination variant was detected in one unrelated individual (LRP5: Ala919CysfsTer67). The average number of genes with protein-altering variants was greater in subjects with confirmed FEVR (1.46, n = 30) compared to subjects confirmed unaffected by FEVR (0.95, n = 20), (p = 0.009). Thirty-four percent of persons sequenced had digenic and trigenic protein-altering variants within this set of FEVR genes, which was much greater than expected in the general population (3.6%), as derived from GnomAD data. While the potential contributions to FEVR are not known for most of the variants in a multigenic context, the high multigenic frequency suggests that potential multigenic contributions to FEVR severity warrant future investigation. The targeted-sequencing format developed will support such exploration by reducing the testing cost to $250 (US) for seven genes and facilitating greater access to genetic testing for families with this very rare inherited retinal disease.

The blood-brain barrier (BBB) protects the central nervous system (CNS) from harmful blood-borne factors. Although BBB dysfunction is a hallmark of several neurological disorders, therapies to restore BBB function are lacking. An attractive strategy is to repurpose developmental BBB regulators, such as Wnt7a, into BBB-protective agents. However, safe therapeutic use of Wnt ligands is complicated by their pleiotropic Frizzled signaling activities. Taking advantage of the Wnt7a/b-specific Gpr124/Reck co-receptor complex, we genetically engineered Wnt7a ligands into BBB-specific Wnt activators. In a “hit-and-run” adeno-associated virus–assisted CNS gene delivery setting, these new Gpr124/Reck-specific agonists protected BBB function, thereby mitigating glioblastoma expansion and ischemic stroke infarction. This work reveals that the signaling specificity of Wnt ligands is adjustable and defines a modality to treat CNS disorders by normalizing the BBB.

Expression of Frizzled 9 (FZD9) is critical to the activity of the lung cancer chemoprevention agent and prostacyclin analogue, iloprost. FZD9 is required in lung epithelial cells for iloprost to activate peroxisome proliferator activated receptor gamma (PPARG) and related anti-tumor signaling. We aimed to investigate which miRNA regulate FZD9 in the context of cigarette smoke exposure and iloprost treatment. We found that miR-520a-5p binds the FZD9 3’UTR in lung cell lines and alters activity and expression of FZD9 downstream targets. Cigarette smoke condensate (CSC) increases expression of miR-520a-5p, while iloprost decreases expression. Cancer promoting effects of a miR-520a-5p mimic were rescued with iloprost treatment, and effects of cigarette smoke were partially rescued with a miR-520a-5p inhibitor. Here we confirm miR-520a-5p as a regulator of FZD9 activity and a mediator of CSC and iloprost effects in the lung. Targeting miR-520a-5p could be an approach to restoring FZD9 expression and improving response to iloprost lung cancer chemoprevention.

Wnt signaling is an evolutionary cell-to-cell coordination mechanism and it is highly critical for a variety of physiological processes of an organism's body, including stem cell regeneration, proliferation, division, migration, polarity of a cell, determining fate of the cell and specification of neural crest, neural symmetry and morphogenesis. Wnts are extracellular secreted glycol proteins, consisted of a family of 19 human proteins that represent the complex nature of the regulatory structure and physiological efficiency of signaling. Moreover, a Wnt/β-catenin-dependent pathway and the β-catenin-independent pathway that is further classified into the Planar Cell Polarity and Wnt/Ca

The mammalian tongue is a highly specialized muscular organ. The Wnt5a ligand regulates muscle development by mediating the activation of several noncanonical Wnt signaling pathways in a receptor context-dependent fashion. However, there is poor information on the expression and behavior of Wnt5a proteins during muscle development of the embryonic tongue.The spatiotemporal distribution profiles of the Wnt5a ligand and its receptors, receptor tyrosine kinase-like orphan receptor 2 (Ror2), Frizzled2 (Fzd2), and Frizzled5 (Fzd5), in the developing tongue muscles of prenatal mice from embryonic day 12.5-18.5 were analyzed using immunofluorescence (IF) double staining of a target protein and desmin, a marker protein of myogenic cells. Immunolabeling images were subjected to digital detection analysis using the WinROOF 2018 version 4.19.0 image processing software when needed.IF signals of the Wnt5a ligand protein and its receptors Ror2 and Fzd2 were detected in developing myoblasts and myotubes of the embryonic tongue, but they were undetectable in mature myofibers equipped with sarcomere structures. Fzd2 expression was specific for desmin-positive developing muscle cells, whereas those of Ror2 and the Wnt5a ligand were widespread and nonselective for desmin-positive cells and that of Fzd5 was predominant in desmin-negative cells of the epithelium and subepithelial mesenchyme.Developing muscle cells but not mature myofibers of the mouse embryonic tongue express the Wnt5a ligand and its receptors Ror2 and Fzd2, which may mediate Wnt5a signaling in the development processes of tongue muscle fibers.

To explore the regulatory function of RNA binding motif protein 38 (RBM38) in human acute myeloid leukemia cells HL-60 and its mechanism.The lentivirus carriers of overexpressed and knockdown RBM38 were constructed. After HL-60 cells were transfected, Western blot was used to analyze the expression level of RBM38 in HL-60 cells. The cell proliferation and cycle of HL-60 were detected by CCK-8 assay and flow cytometry assay, respectively. RNA immunoprecipitation coupled real-time PCR (RIP-qPCR) was used to detect the combination of RBM38 with mRNAs. Actinomycin D treatment followed by real-time PCR (AcD-qPCR) was used to detect the effect of RBM38 on the stability of target mRNAs.RBM38 in HL-60 cells was overexpressed or inhibited by lentivirus transduction. Overexpressed RBM38 promoted the cell cycle and proliferation of HL-60, while RBM38 knockdown repressed the two processes. RBM38 showed an interaction with FZD1 mRNA and enhancement of its stability.RBM38 can regulate cell proliferation of HL-60 by improving the stability of FZD1 mRNA.RBM38调节FZD1 mRNA稳定性介导急性髓系白血病细胞HL-60的增殖.研究RNA结合基序蛋白38（RBM38）对人急性髓系白血病细胞株HL-60增殖的调控功能和作用机制。.分别构建过表达、敲低RBM38的慢病毒载体，使用慢病毒感染HL-60细胞后，Western blot方法检测细胞内RBM38的表达水平，分别用CCK-8法和PI染色结合流式细胞术检测HL-60细胞的增殖和周期，通过RNA免疫共沉淀结合实时定量PCR方法检测RBM38与靶mRNA的结合情况，放线菌素D处理结合实时定量PCR方法检测RBM38对靶mRNA稳定性的影响。.通过慢病毒介导的基因转移可成功实现HL-60细胞内RBM38的过表达或表达抑制。过表达RBM38可显著促进HL-60细胞的增殖活性和细胞周期运行；相反，抑制内源性RBM38的表达可抑制HL-60细胞增殖和细胞周期。RBM38可与FZD1 mRNA结合，促进其mRNA的稳定。.RBM38通过促进FZD1 mRNA的稳定调节HL-60细胞的增殖。.