Meflin is a marker of pancreatic stellate cells involved in fibrosis and epithelial regeneration in the pancreas.

Pancreatic stellate cells (PSCs) are stromal cells in the pancreas that play an important role in pancreatic pathology. In chronic pancreatitis (CP) and pancreatic ductal adenocarcinoma (PDAC), PSCs are known to get activated to form myofibroblasts or cancer-associated fibroblasts (CAFs) that promote stromal fibroinflammatory reactions. However, previous studies on PSCs were mainly based on the findings obtained using ex vivo expanded PSCs, with few studies that addressed the significance of in situ tissue-resident PSCs using animal models. Their contributions to fibrotic reactions in CP and PDAC are also lesser-known. These limitations in our understanding of PSC biology have been attributed to the lack of specific molecular markers of PSCs. Herein, we established Meflin (Islr), a glycosylphosphatidylinositol-anchored membrane protein, as a PSC-specific marker in both mouse and human by using human pancreatic tissue samples and Meflin reporter mice. Meflin-positive (Meflin+ ) cells contain lipid droplets and express the conventional PSC marker Desmin in normal mouse pancreas, with some cells also positive for Gli1, the marker of pancreatic tissue-resident fibroblasts. Three-dimensional analysis of the cleared pancreas of Meflin reporter mice showed that Meflin+ PSCs have long and thin cytoplasmic protrusions, and are localised on the abluminal side of vessels in the normal pancreas. Lineage tracing experiments revealed that Meflin+ PSCs constitute one of the origins of fibroblasts and CAFs in CP and PDAC, respectively. In these diseases, Meflin+ PSC-derived fibroblasts showed a distinctive morphology and distribution from Meflin+ PSCs in the normal pancreas. Furthermore, we showed that the genetic depletion of Meflin+ PSCs accelerated fibrosis and attenuated epithelial regeneration and stromal R-spondin 3 expression, thereby implying that Meflin+ PSCs and their lineage cells may support tissue recovery and Wnt/R-spondin signalling after pancreatic injury and PDAC development. Together, these data indicate that Meflin may be a marker specific to tissue-resident PSCs and useful for studying their biology in both health and disease. © 2023 The Pathological Society of Great Britain and Ireland.

Wnt activation disturbs cell competition and causes diffuse invasion of transformed cells through NF-κB-MMP21 pathway.

Normal epithelial cells exert their competitive advantage over RasV12-transformed cells and eliminate them into the apical lumen via cell competition. However, the internal or external factors that compromise cell competition and provoke carcinogenesis remain elusive. In this study, we examine the effect of sequential accumulation of gene mutations, mimicking multi-sequential carcinogenesis on RasV12-induced cell competition in intestinal epithelial tissues. Consequently, we find that the directionality of RasV12-cell extrusion in Wnt-activated epithelia is reversed, and transformed cells are delaminated into the basal lamina via non-cell autonomous MMP21 upregulation. Subsequently, diffusively infiltrating, transformed cells develop into highly invasive carcinomas. The elevated production of MMP21 is elicited partly through NF-κB signaling, blockage of which restores apical elimination of RasV12 cells. We further demonstrate that the NF-κB-MMP21 axis is significantly bolstered in early colorectal carcinoma in humans. Collectively, this study shows that cells with high mutational burdens exploit cell competition for their benefit by behaving as unfit cells, endowing them with an invasion advantage.

Significance of expression of CD109 in osteosarcoma and its involvement in tumor progression via BMP signaling.

Osteosarcoma, the most common primary malignant bone tumor, is defined by the formation of neoplastic osteoid and/or bone. This sarcoma is a highly heterogeneous disease with a wide range of patient outcomes. CD109 is a glycosylphosphatidylinositol-anchored glycoprotein that is highly expressed in various types of malignant tumors. We previously reported that CD109 is expressed in osteoblasts and osteoclasts in normal human tissues and plays a role in bone metabolism in vivo. While CD109 has been shown to promote various carcinomas through the downregulation of TGF-ß signaling, the role and mechanism of CD109 in sarcomas remain largely unknown. In this study, we investigated the molecular function of CD109 in sarcomas using osteosarcoma cell lines and tissue. Semi-quantitative immunohistochemical analysis using human osteosarcoma tissue revealed a significantly worse prognosis in the CD109-high group compared with the CD109-low group. We found no association between CD109 expression and TGF-ß signaling in osteosarcoma cells. However, enhancement of SMAD1/5/9 phosphorylation was observed in CD109 knockdown cells under bone morphogenetic protein-2 (BMP-2) stimulation. We also performed immunohistochemical analysis for phospho-SMAD1/5/9 using human osteosarcoma tissue and found a negative correlation between CD109 expression and SMAD1/5/9 phosphorylation. In vitro wound healing assay showed that osteosarcoma cell migration was significantly attenuated in CD109-knockdown cells compared with control cells in the presence of BMP. These results suggest that CD109 is a poor prognostic factor in osteosarcoma and affects tumor cell migration via BMP signaling.

Novel Therapeutic Potentials of Taxifolin for Obesity-Induced Hepatic Steatosis, Fibrogenesis, and Tumorigenesis.

The molecular pathogenesis of nonalcoholic steatohepatitis (NASH) includes a complex interaction of metabolic stress and inflammatory stimuli. Considering the therapeutic goals of NASH, it is important to determine whether the treatment can prevent the progression from NASH to hepatocellular carcinoma. Taxifolin, also known as dihydroquercetin, is a natural bioactive flavonoid with antioxidant and anti-inflammatory properties commonly found in various foods and health supplement products. In this study, we demonstrated that Taxifolin treatment markedly prevented the development of hepatic steatosis, chronic inflammation, and liver fibrosis in a murine model of NASH. Its mechanisms include a direct action on hepatocytes to inhibit lipid accumulation. Taxifolin also increased brown adipose tissue activity and suppressed body weight gain through at least two distinct pathways: direct action on brown adipocytes and indirect action via fibroblast growth factor 21 production in the liver. Notably, the Taxifolin treatment after NASH development could effectively prevent the development of liver tumors. Collectively, this study provides evidence that Taxifolin shows pleiotropic effects for the treatment of the NASH continuum. Our data also provide insight into the novel mechanisms of action of Taxifolin, which has been widely used as a health supplement with high safety.

Possible disease-protective roles of fibroblasts in cancer and fibrosis and their therapeutic application.

Cancer and fibrotic diseases are characterized by continuous inflammation, tissue wounds, and injuries. Cancer is a "wound that does not heal," and the uncontrolled proliferation of cancer cells disrupts normal tissue integrity and induces stromal fibroinflammatory reactions. Fibroblasts proliferate extensively in the stroma, playing a major role in the development of these diseases. There has been considerable evidence that fibroblasts contribute to fibrosis and tissue stiffening and promote disease progression via multiple mechanisms. However, recent emerging findings, mainly derived from single-cell transcriptomic analysis, indicated that fibroblasts are functionally heterogeneous, leading to the hypothesis that both disease-promoting and -restraining fibroblasts exist. We recently showed that a fibroblast population, defined by the expression of the glycosylphosphatidylinositol-anchored membrane protein Meflin may suppress but not promote fibrotic response and disease progression in cancer and fibrotic diseases. Although currently hypothetical, the primary function of Meflin-positive fibroblasts may be tissue repair after injury and cancer initiation occurred. This observation has led to the proposal of a potential therapy that converts the phenotype of fibroblasts from pro-tumor to anti-tumor. In this short review, we summarize our recent findings on the function of Meflin in the context of cancer and fibrotic diseases and discuss how we can utilize this knowledge on fibroblasts in translational medicine. We also discuss several aspects of the interpretation of survival analysis data, such as Kaplan-Meier analysis, to address the function of specific genes expressed in fibroblasts.

BRCA1 haploinsufficiency promotes chromosomal amplification under Fenton reaction-based carcinogenesis through ferroptosis-resistance.

Germline-mutation in BRCA1 tumor suppressor gene is an established risk for carcinogenesis not only in females but also in males. Deficiency in the repair of DNA double-strand breaks is hypothesized as a responsible mechanism for carcinogenesis. However, supporting data is insufficient both in the mutation spectra of cancers in the patients with BRCA1 germline-mutation and in murine knockout/knock-in models of Brca1 haploinsufficiency. Furthermore, information on the driving force toward carcinogenesis in BRCA1 mutation carriers is lacking. Here we applied Fenton reaction-based renal carcinogenesis to a rat heterozygously knockout model of BRCA1 haploinsufficiency (mutant [MUT] model; L63X/+). Rat MUT model revealed significant promotion of renal cell carcinoma (RCC) induced by ferric nitrilotriacetate (Fe-NTA). Array-based comparative genome hybridization of the RCCs identified significant increase in chromosomal amplification, syntenic to those in breast cancers of BRCA1 mutation carriers, including c-Myc, in comparison to those in the wild-type. Subacute-phase analysis of the kidney after repeated Fe-NTA treatment in the MUT model revealed dysregulated iron metabolism with mitochondrial malfunction assessed by expression microarray and electron microscopy, leading to renal tubular proliferation with iron overload. In conclusion, we for the first time demonstrate that biallelic wild-type BRCA1 provides more robust protection for mitochondrial metabolism under iron-catalyzed oxidative stress, preventing the emergence of neoplastic cells with chromosomal amplification. Our results suggest that oxidative stress via excess iron is a major driving force for carcinogenesis in BRCA1 haploinsufficiency, which can be a target for cancer prevention and therapeutics.

BMP3b Is a Novel Antifibrotic Molecule Regulated by Meflin in Lung Fibroblasts.

Fibroblasts play a central role in the lung fibrotic process. Our recent study identified a novel subpopulation of lung fibroblasts expressing meflin (mesenchymal stromal cell- and fibroblast-expressing Linx paralogue), antifibrotic properties of which were confirmed by murine lung fibrosis model. Meflin-expressing fibroblasts were resistant to fibrogenesis induced by TGF-ß (transforming growth factor-ß), but its underlying mechanisms remain unknown. In this study, evaluation of a silica-nanoparticle-induced lung fibrosis model confirmed the antifibrotic effect of meflin via the regulation of TGF-ß signaling. We conducted comparative gene expression profiling in lung fibroblasts, which identified growth differentiation factor 10 (Gdf10) encoding bone morphogenic protein 3b (BMP3b) as the most downregulated gene in meflin-deficient cells under the profibrotic condition with TGF-ß. We hypothesized that BMP3b can be an effector molecule playing an antifibrotic role downstream of meflin. As suggested by single-cell transcriptomic data, restricted expressions of Gdf10 (Bmp3b) in stromal cells including fibroblasts were confirmed. We examined possible antifibrotic properties of BMP3b in lung fibroblasts and demonstrated that Bmp3b-null fibroblasts were more susceptible to TGF-ß-induced fibrogenic changes. Furthermore, Bmp3b-null mice exhibited exaggerated lung fibrosis induced by silica-nanoparticles in vivo. We also demonstrated that treatment with recombinant BMP3B was effective against TGF-ß-induced fibrogenesis in fibroblasts, especially in the suppression of excessive extracellular matrix production. These lines of evidence suggested that BMP3b is a novel humoral effector molecule regulated by meflin which exerts antifibrotic properties in lung fibroblasts. Supplementation of BMP3B could be a novel therapeutic strategy for fibrotic lung diseases.

Pharmacologic conversion of cancer-associated fibroblasts from a protumor phenotype to an antitumor phenotype improves the sensitivity of pancreatic cancer to chemotherapeutics.

Previous therapeutic attempts to deplete cancer-associated fibroblasts (CAFs) or inhibit their proliferation in pancreatic ductal adenocarcinoma (PDAC) were not successful in mice or patients. Thus, CAFs may be tumor suppressive or heterogeneous, with distinct cancer-restraining and -promoting CAFs (rCAFs and pCAFs, respectively). Here, we showed that induced expression of the glycosylphosphatidylinositol-anchored protein Meflin, a rCAF-specific marker, in CAFs by genetic and pharmacological approaches improved the chemosensitivity of mouse PDAC. A chemical library screen identified Am80, a synthetic, nonnatural retinoid, as a reagent that effectively induced Meflin expression in CAFs. Am80 administration improved the sensitivity of PDAC to chemotherapeutics, accompanied by increases in tumor vessel area and intratumoral drug delivery. Mechanistically, Meflin was involved in the suppression of tissue stiffening by interacting with lysyl oxidase to inhibit its collagen crosslinking activity. These data suggested that modulation of CAF heterogeneity may represent a strategy for PDAC treatment.

A novel renal perivascular mesenchymal cell subset gives rise to fibroblasts distinct from classic myofibroblasts.

Perivascular mesenchymal cells (PMCs), which include pericytes, give rise to myofibroblasts that contribute to chronic kidney disease progression. Several PMC markers have been identified; however, PMC heterogeneity and functions are not fully understood. Here, we describe a novel subset of renal PMCs that express Meflin, a glycosylphosphatidylinositol-anchored protein that was recently identified as a marker of fibroblasts essential for cardiac tissue repair. Tracing the lineage of Meflin+ PMCs, which are found in perivascular and periglomerular areas and exhibit renin-producing potential, showed that they detach from the vasculature and proliferate under disease conditions. Although the contribution of Meflin+ PMCs to conventional α-SMA+ myofibroblasts is low, they give rise to fibroblasts with heterogeneous α-SMA expression patterns. Genetic ablation of Meflin+ PMCs in a renal fibrosis mouse model revealed their essential role in collagen production. Consistent with this, human biopsy samples showed that progressive renal diseases exhibit high Meflin expression. Furthermore, Meflin overexpression in kidney fibroblasts promoted bone morphogenetic protein 7 signals and suppressed myofibroblastic differentiation, implicating the roles of Meflin in suppressing tissue fibrosis. These findings demonstrate that Meflin marks a PMC subset that is functionally distinct from classic pericytes and myofibroblasts, highlighting the importance of elucidating PMC heterogeneity.

Meflin-positive cancer-associated fibroblasts enhance tumor response to immune checkpoint blockade.

Cancer-associated fibroblasts (CAFs) are an integral component of the tumor microenvironment (TME). Most CAFs shape the TME toward an immunosuppressive milieu and attenuate the efficacy of immune checkpoint blockade (ICB) therapy. However, the detailed mechanism of how heterogeneous CAFs regulate tumor response to ICB therapy has not been defined. Here, we show that a recently defined CAF subset characterized by the expression of Meflin, a glycosylphosphatidylinositol-anchored protein marker of mesenchymal stromal/stem cells, is associated with survival and favorable therapeutic response to ICB monotherapy in patients with non-small cell lung cancer (NSCLC). The prevalence of Meflin-positive CAFs was positively correlated with CD4-positive T-cell infiltration and vascularization within non-small cell lung cancer tumors. Meflin deficiency and CAF-specific Meflin overexpression resulted in defective and enhanced ICB therapy responses in syngeneic tumors in mice, respectively. These findings suggest the presence of a CAF subset that promotes ICB therapy efficacy, which adds to our understanding of CAF functions and heterogeneity.

Matrix remodeling-associated protein 8 is a marker of a subset of cancer-associated fibroblasts in pancreatic cancer.

Cancer-associated fibroblasts (CAFs), a compartment of the tumor microenvironment, were previously thought to be a uniform cell population that promotes cancer progression. However, recent studies have shown that CAFs are heterogeneous and that there are at least two types of CAFs, that is, cancer-promoting and -restraining CAFs. We previously identified Meflin as a candidate marker of cancer-restraining CAFs (rCAFs) in pancreatic ductal adenocarcinoma (PDAC). The precise nature of rCAFs, however, has remained elusive owing to a lack of understanding of their comprehensive gene signatures. Here, we screened genes whose expression correlated with Meflin in single-cell transcriptomic analyses of human cancers. Among the identified genes, we identified matrix remodeling-associated protein 8 (MXRA8), which encodes a type I transmembrane protein with unknown molecular function. Analysis of MXRA8 expression in human PDAC samples showed that MXRA8 was differentially co-expressed with other CAF markers. Moreover, in patients with PDAC or syngeneic tumors developed in MXRA8-knockout mice, MXRA8 expression did not affect the roles of CAFs in cancer progression, and the biological importance of MXRA8+ CAFs is still unclear. Overall, we identified MXRA8 as a new CAF marker; further studies are needed to determine the relevance of this marker.

Newly established patient-derived organoid model of intracranial meningioma.

BACKGROUND: Recent comprehensive studies have revealed several molecular alterations that are frequently found in meningiomas. However, effective treatment reagents targeting specific molecular alterations have not yet been identified because of the limited number of representative research models of meningiomas. METHODS: We performed organoid cultures using meningioma cells and meningioma tumor tissues. Using immunohistochemistry and molecular analyses consisting of whole-exome sequencing, RNA-seq, and DNA methylation analyses, we compared the histological findings and molecular profiling of organoid models with those of parental tumors. Further, using these organoid models together with a public database of meningiomas, we explored molecular alterations, which are a potent treatment target for meningioma. RESULTS: We established 18 organoid models comprising of two malignant meningioma cells (HKBMM and IOMM-Lee), 10 benign meningiomas, four malignant meningiomas, and two solitary fibrous tumors (SFTs). The organoids exhibited consistent histological features and molecular profiles with those of the parental tumors. Using a public database, we identified that upregulated forkhead box M1 (FOXM1) was correlated with increased tumor proliferation. Overexpression of FOXM1 in benign meningioma organoids increased organoid proliferation; depletion of FOXM1 in malignant organoids decreased proliferation. Additionally, thiostrepton, a FOXM1 inhibitor combined with radiation therapy, significantly inhibited the proliferation of malignant meningioma organoid models. CONCLUSIONS: An organoid model for meningioma enabled us to elucidate the tumor biology of meningioma along with potent treatment targets for meningioma.

Meflin defines mesenchymal stem cells and/or their early progenitors with multilineage differentiation capacity.

Mesenchymal stem cells (MSCs) are the likely precursors of multiple lines of mesenchymal cells. The existence of bona fide MSCs with self-renewal capacity and differentiation potential into all mesenchymal lineages, however, has been unclear because of the lack of MSC-specific marker(s) that are not expressed by the terminally differentiated progeny. Meflin, a glycosylphosphatidylinositol-anchored protein, is an MSC marker candidate that is specifically expressed in rare stromal cells in all tissues. Our previous report showed that Meflin expression becomes down-regulated in bone marrow-derived MSCs cultured on plastic, making it difficult to examine the self-renewal and differentiation of Meflin-positive cells at the single-cell level. Here, we traced the lineage of Meflin-positive cells in postnatal and adult mice, showing that those cells differentiated into white and brown adipocytes, osteocytes, chondrocytes and skeletal myocytes. Interestingly, cells derived from Meflin-positive cells formed clusters of differentiated cells, implying the in situ proliferation of Meflin-positive cells or their lineage-committed progenitors. These results, taken together with previous findings that Meflin expression in cultured MSCs was lost upon their multilineage differentiation, suggest that Meflin is a useful potential marker to localize MSCs and/or their immature progenitors in multiple tissues.

The sodium-glucose cotransporter-2 inhibitor Tofogliflozin prevents the progression of nonalcoholic steatohepatitis-associated liver tumors in a novel murine model.

BACKGROUND: Diabetes and obesity contribute to the pathogenesis of nonalcoholic steatohepatitis (NASH) and hepatocellular carcinoma (HCC). However, how diabetes and obesity accelerate liver tumorigenesis remains to be fully understood. Moreover, to verify the therapeutic potential of anti-diabetic drugs, there exists a strong need for appropriate animal models that recapitulate human pathophysiology of NASH and HCC. METHODS: We established a novel murine model of NASH-associated liver tumors using genetically obese melanocortin 4 receptor-deficient mice fed on Western diet in combination with a chemical procarcinogen, and verified the validity of our model in evaluating drug efficacy. FINDINGS: Our model developed multiple liver tumors together with obesity, diabetes, and NASH within a relatively short period (approximately 3 months). In this model, sodium glucose cotransporter 2 inhibitor Tofogliflozin prevented the development of NASH-like liver phenotypes and the progression of liver tumors. Tofogliflozin attenuated p21 expression of hepatocytes in non-tumorous lesions in the liver. INTERPRETATION: Tofogliflozin treatment attenuates cellular senescence of hepatocytes under obese and diabetic conditions. This study provides a unique animal model of NASH-associated liver tumors, which is applicable for assessing drug efficacy to prevent or treat NASH-associated HCC.

Urinary MicroRNA-Based Diagnostic Model for Central Nervous System Tumors Using Nanowire Scaffolds.

There are no accurate mass screening methods for early detection of central nervous system (CNS) tumors. Recently, liquid biopsy has received a lot of attention for less-invasive cancer screening. Unlike other cancers, CNS tumors require efforts to find biomarkers due to the blood-brain barrier, which restricts molecular exchange between the parenchyma and blood. Additionally, because a satisfactory way to collect urinary biomarkers is lacking, urine-based liquid biopsy has not been fully investigated despite the fact that it has some advantages compared to blood or cerebrospinal fluid-based biopsy. Here, we have developed a mass-producible and sterilizable nanowire-based device that can extract urinary microRNAs efficiently. Urinary microRNAs from patients with CNS tumors (n = 119) and noncancer individuals (n = 100) were analyzed using a microarray to yield comprehensive microRNA expression profiles. To clarify the origin of urinary microRNAs of patients with CNS tumors, glioblastoma organoids were generated. Glioblastoma organoid-derived differentially expressed microRNAs (DEMs) included 73.4% of the DEMs in urine of patients with parental tumors but included only 3.9% of those in urine of noncancer individuals, which suggested that many CNS tumor-derived microRNAs could be identified in urine directly. We constructed the diagnostic model based on the expression of the selected microRNAs and found that it was able to differentiate patients and noncancer individuals at a sensitivity and specificity of 100 and 97%, respectively, in an independent dataset. Our findings demonstrate that urinary microRNAs extracted with the nanowire device offer a well-fitted strategy for mass screening of CNS tumors.

The Balance of Stromal BMP Signaling Mediated by GREM1 and ISLR Drives Colorectal Carcinogenesis.

BACKGROUND & AIMS: Cancer-associated fibroblasts (CAFs), key constituents of the tumor microenvironment, either promote or restrain tumor growth. Attempts to therapeutically target CAFs have been hampered by our incomplete understanding of these functionally heterogeneous cells. Key growth factors in the intestinal epithelial niche, bone morphogenetic proteins (BMPs), also play a critical role in colorectal cancer (CRC) progression. However, the crucial proteins regulating stromal BMP balance and the potential application of BMP signaling to manage CRC remain largely unexplored. METHODS: Using human CRC RNA expression data, we identified CAF-specific factors involved in BMP signaling, then verified and characterized their expression in the CRC stroma by in situ hybridization. CRC tumoroids and a mouse model of CRC hepatic metastasis were used to test approaches to modify BMP signaling and treat CRC. RESULTS: We identified Grem1 and Islr as CAF-specific genes involved in BMP signaling. Functionally, GREM1 and ISLR acted to inhibit and promote BMP signaling, respectively. Grem1 and Islr marked distinct fibroblast subpopulations and were differentially regulated by transforming growth factor ß and FOXL1, providing an underlying mechanism to explain fibroblast biological dichotomy. In patients with CRC, high GREM1 and ISLR expression levels were associated with poor and favorable survival, respectively. A GREM1-neutralizing antibody or fibroblast Islr overexpression reduced CRC tumoroid growth and promoted Lgr5+ intestinal stem cell differentiation. Finally, adeno-associated virus 8 (AAV8)-mediated delivery of Islr to hepatocytes increased BMP signaling and improved survival in our mouse model of hepatic metastasis. CONCLUSIONS: Stromal BMP signaling predicts and modifies CRC progression and survival, and it can be therapeutically targeted by novel AAV-directed gene delivery to the liver.

CD109 regulates in vivo tumor invasion in lung adenocarcinoma through TGF-ß signaling.

Stromal invasion is considered an important prognostic factor in patients with lung adenocarcinoma. The mechanisms underlying the formation of tumor stroma and stromal invasion have been studied in the lung; however, they are still unclear. CD109 is a glycosylphosphatidylinositol-anchored glycoprotein highly expressed in several types of human malignant tumors including lung cancers. In this study, we investigated the in vivo functions of CD109 protein in malignant lung tumors. Initially, we identified an association between higher expression of CD109 protein in human lung adenocarcinoma and a significantly worse prognosis, according to immunohistochemical analysis. We also showed that CD109 deficiency significantly reduced the area of stromal invasive lesions in a genetically engineered CD109-deficient lung adenocarcinoma mouse model, which correlated with the results observed in human lung adenocarcinoma. Furthermore, we identified latent TGF-ß binding protein-1 (LTBP1) as a CD109-interacting protein using mass spectrometry and confirmed their interaction by co-immunoprecipitation. Importantly, increased CD109 expression enhanced stromal TGF-ß activation in the presence of LTBP1. Therefore, these data suggest the significance of the regulation of TGF-ß signaling through CD109 and LTBP1 interaction in tumor stroma and also reveal the importance of CD109 expression levels in promoting lung cancer cell proliferation, migration, and invasion, and thus predicting the outcome of patients suffering from lung adenocarcinoma. Therefore, CD109 protein could be a potential therapeutic target for this disease.

The Daple-CK1ε complex regulates Dvl2 phosphorylation and canonical Wnt signaling.

The canonical Wnt signaling pathway plays a crucial role in embryonic development, tissue homeostasis and cancer progression. The binding of Wnt ligands to their cognate receptors, the Frizzled (Fzd) family of proteins, recruits Dishevelled segment polarity protein (Dvl) to the plasma membrane and induces its phosphorylation via casein kinase 1 (CK1), which leads to the activation of ß-catenin. Previous studies showed that Dishevelled-associating protein with a high frequency of leucine residues (Daple) is an important component of the Wnt signaling pathway and essential for Dvl phosphorylation. However, the mechanism by which Daple promotes CK1-mediated phosphorylation of Dvl is not fully understood. In this study, we found that Daple overexpression induced CK1ε-mediated Dvl2 phosphorylation at threonine 224 (Thr224). A Daple mutant (Daple ΔGCV) that lacks a carboxyl-terminal motif to associate with Dvl, retained the ability to interact with CK1ε, but did not induce Dvl phosphorylation, suggesting the importance of the Daple/Dvl/CK1ε trimeric protein complex. We further found that Thr224 phosphorylation of Dvl was required for full activation of ß-catenin transcriptional activity. Consistent with this, wild-type Daple promoted ß-catenin transcriptional activity, following dissociation of ß-catenin and axin. Finally, Wnt3a stimulation increased the membrane localization of Daple and its association with Dvl, and Daple knockdown attenuated Wnt3a-mediated ß-catenin transcriptional activity. Collectively, these data suggested a essential role of spatial Daple localization in CK1ε-mediated activation of Dvl in the canonical Wnt signaling pathway.