Search

Pancreatic ductal adenocarcinoma (PDAC) is one of the most refractory cancers with the worst prognosis. Although several molecules are known to be associated with the progression of PDAC, the molecular mechanisms underlying the progression of PDAC remain largely elusive. The Ror-family receptors, Ror1 and Ror2, which act as a receptor(s) for Wnt-family ligands, particularly Wnt5a, are involved in the progression of various types of cancers. Here, we show that higher expression of Ror1 and Wnt5b, but not Ror2, are associated with poorer prognosis of PDAC patients, and that Ror1 and Wnt5b are expressed highly in a type of PDAC cell lines, PANC-1 cells. Knockdown of either Ror1 or Wnt5b in PANC-1 cells inhibited their proliferation significantly in vitro, and knockout of Ror1 in PANC-1 cells resulted in a significant inhibition of tumor growth in vivo. Furthermore, we show that Wnt5b-Ror1 signaling in PANC-1 cells promotes their proliferation in a cell-autonomous manner by modulating our experimental setting in vitro. Collectively, these findings indicate that Wnt5b-Ror1 signaling might play an important role in the progression of some if not all of PDAC by promoting proliferation., (© 2024 Molecular Biology Society of Japan and John Wiley & Sons Australia, Ltd.)

Ameloblastoma is an odontogenic neoplasm characterized by slow intraosseous growth with progressive jaw resorption. Recent reports have revealed that ameloblastoma harbours an oncogenic BRAFV600E mutation with mitogen-activated protein kinase (MAPK) pathway activation and described cases of ameloblastoma harbouring a BRAFV600E mutation in which patients were successfully treated with a BRAF inhibitor. Therefore, the MAPK pathway may be involved in the development of ameloblastoma; however, the precise mechanism by which it induces ameloblastoma is unclear. The expression of ADP-ribosylation factor (ARF)-like 4c (ARL4C), induced by a combination of the EGF-MAPK pathway and Wnt/β-catenin signalling, has been shown to induce epithelial morphogenesis. It was also reported that the overexpression of ARL4C, due to alterations in the EGF/RAS-MAPK pathway and Wnt/β-catenin signalling, promotes tumourigenesis. However, the roles of ARL4C in ameloblastoma are unknown. We investigated the involvement of ARL4C in the development of ameloblastoma. In immunohistochemical analyses of tissue specimens obtained from 38 ameloblastoma patients, ARL4C was hardly detected in non-tumour regions but tumours frequently showed strong expression of ARL4C, along with the expression of both BRAFV600E and RAF1 (also known as C-RAF). Loss-of-function experiments using inhibitors or siRNAs revealed that ARL4C elevation depended on the RAF1-MEK/ERK pathway in ameloblastoma cells. It was also shown that the RAF1-ARL4C and BRAFV600E-MEK/ERK pathways promoted cell proliferation independently. ARL4C-depleted tumour cells (generated by knockdown or knockout) exhibited decreased proliferation and migration capabilities. Finally, when ameloblastoma cells were co-cultured with mouse bone marrow cells and primary osteoblasts, ameloblastoma cells induced osteoclast formation. ARL4C elevation in ameloblastoma further promoted its formation capabilities through the increased RANKL expression of mouse bone marrow cells and/or primary osteoblasts. These results suggest that the RAF1-MEK/ERK-ARL4C axis, which may function in cooperation with the BRAFV600E-MEK/ERK pathway, promotes ameloblastoma development. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd., (© 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.)

There are currently no treatments for salivary gland diseases, making it vital to understand signaling mechanisms operating in acinar and ductal cells so as to develop regenerative therapies. To date, little work has focused on elucidating the signaling cascades controlling the differentiation of these cell types in adult mammals. To analyze the function of the Hippo-TAZ/YAP1 pathway in adult mouse salivary glands, we generated adMOB1DKO mice in which both MOB1A and MOB1B were TAM-inducibly deleted when the animals were adults. Three weeks after TAM treatment, adMOB1DKO mice exhibited smaller submandibular glands (SMGs) than controls with a decreased number of acinar cells and an increased number of immature dysplastic ductal cells. The mutants suffered from reduced saliva production accompanied by mild inflammatory cell infiltration and fibrosis in SMGs, similar to the Sjogren's syndrome. MOB1-deficient acinar cells showed normal proliferation and apoptosis but decreased differentiation, leading to an increase in acinar/ductal bilineage progenitor cells. These changes were TAZ-dependent but YAP1-independent. Biochemically, MOB1-deficient salivary epithelial cells showed activation of the TAZ/YAP1 and β-catenin in ductal cells, but reduced SOX2 and SOX10 expression in acinar cells. Thus, Hippo-TAZ signaling is critical for proper ductal and acinar cell differentiation and function in adult mice., (© 2021 Molecular Biology Society of Japan and John Wiley & Sons Australia, Ltd.)

Dickkopf1 (DKK1) is overexpressed in various cancers and promotes cancer cell proliferation by binding to cytoskeleton-associated protein 4 (CKAP4). However, the mechanisms underlying DKK1 expression are poorly understood. RNA sequence analysis revealed that expression of the transcription factor forkhead box M1 (FOXM1) and its target genes concordantly fluctuated with expression of DKK1 in pancreatic ductal adenocarcinoma (PDAC) cells. DKK1 knockdown decreased FOXM1 expression and vice versa in PDAC and esophageal squamous cell carcinoma (ESCC) cells. Inhibition of either the DKK1-CKAP4-AKT pathway or the ERK pathway suppressed FOXM1 expression, and simultaneous inhibition of both pathways showed synergistic effects. A FOXM1 binding site was identified in the 5'-untranslated region of the DKK1 gene, and its depletion decreased DKK1 expression and cancer cell proliferation. Clinicopathological and database analysis revealed that PDAC and ESCC patients who simultaneously express DKK1 and FOXM1 have a poorer prognosis. Multivariate analysis demonstrated that expression of both DKK1 and FOXM1 is the independent prognostic factor in ESCC patients. Although it has been reported that FOXM1 enhances Wnt signaling, FOXM1 induced DKK1 expression independently of Wnt signaling in PDAC and ESCC cells. These results suggest that DKK1 and FOXM1 create a positive feedback loop to promote cancer cell proliferation.

Most cancer cells are exposed to altered extracellular environments, such as an increase in extracellular matrix (ECM) stiffness and soluble signals consisting of growth factors and cytokines. It is therefore conceivable that changes in tumor extracellular environments affect tumor cell behavior. The Hippo pathway reportedly responds to the extracellular environment and regulates the nuclear localization of the transcription co-activator, yes-associated protein (YAP)/transcriptional co-activator with PDZ-binding motif (TAZ). Inactivation of the Hippo pathway with nuclear translocation of YAP/TAZ stimulates cell proliferation. Its pathway also regulates gene expression, but the precise molecule(s) meditating the cell-proliferating effect of YAP signaling on oral squamous cell carcinoma (OSCC) is unclear. First, we examined the effects of YAP signaling on OSCC tumorigenesis. Loss-of-function experiments using siRNA or an inhibitor, and immunohistochemical analyses of tissue specimens obtained from OSCC patients demonstrated that YAP signaling was involved in OSCC cell proliferation. Second, we identified Piezo-type mechanosensitive ion channel component 1 (PIEZO1), a Ca 2+ channel, as a transcriptional target of YAP signaling and showed that elevated PIEZO1 was required for PIEZO1 agonist-dependent Ca 2+ entry and cell proliferation in OSCC cells. Experiments using three-dimensional and suspension culture revealed that PIEZO1 was involved in OSCC cellular growth. Finally, YAP overexpression in the nucleus and/or cytoplasm was immunohistochemically detected in tumor lesions with frequent expression of both PIEZO1 and Ki-67, but not in non-tumor regions of OSCC specimens. These results suggest that the YAP/PIEZO1 axis promotes OSCC cell growth. © 2020 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd., (© 2020 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.)

ADP-ribosylation factor-like 4c (ARL4C) is identified as a small GTP-binding protein, which is expressed by Wnt and EGF signaling and plays an important role in tubulogenesis of cultured cells and the ureters. ARL4C is little expressed in adult tissues, but it is highly expressed in lung cancer and colorectal cancer and shown to represent a molecular target for cancer therapy based on siRNA experiments. This study revealed that ARL4C is highly expressed in primary hepatocellular carcinoma (HCC) tumors and colorectal cancer liver metastases, and that ARL4C expression is associated with poor prognosis for these cancers. Chemically modified antisense oligonucleotides (ASO) against ARL4C effectively reduced ARL4C expression in both HCC and colorectal cancer cells and inhibited proliferation and migration of these cancer cells in vitro ARL4C ASOs decreased the PIK3CD mRNA levels and inhibited the activity of AKT in HCC cells, suggesting that the downstream signaling of ARL4C in HCC cells is different from that in lung and colon cancer cells. In addition, subcutaneous injection of ARL4C ASO was effective in reducing the growth of primary HCC and metastatic colorectal cancer in the liver of immunodeficient mice. ARL4C ASO accumulated in cancer cells more efficiently than the surrounding normal cells in the liver and decreased ARL4C expression in the tumor. These results suggest that ARL4C ASO represents a novel targeted nucleic acid medicine for the treatment of primary and metastatic liver cancers., (©2019 American Association for Cancer Research.)

Epithelial cell rests of Malassez (ERM) are involved in the maintenance and homeostasis of the periodontal ligament. The objective of this study was to investigate the effect of mechanical stretching on cell growth, cell death and differentiation in the ERM. Cultured porcine ERM were stretched for 24 hr in cycles of 18% elongation for 1 sec followed by 1 sec relaxation. The numbers of cells and TUNEL-positive cells were then counted. The expression of mRNAs encoding gap junction protein α1 (Gja1), ameloblastin, bone morphogenetic protein 2 (BMP2), bone morphogenetic protein 4 (BMP4) and noggin were evaluated using quantitative real-time PCR. The number of cells in the stretching group was approximately 1.3-fold higher than that in the non-stretching controls at 24 hr (p<0.01). Apoptotic cells ranged from 1.9-2.5% in the stretching group at 24 hr, but were only 0.6% in the control group (p<0.01). The expression of Gja1, ameloblastin and noggin mRNAs in the stretching group was decreased at 24 hr compared with in the non-stretching group (p<0.01), whereas the expression of BMP2 and BMP4 mRNAs in the stretching group was significantly higher than that in the control group (p<0.01). Incorporation of 18 α-glycyrrhetinic acid (18GA, a gap junction inhibitor) promoted proliferation and apoptosis and confirmed both the increase of BMP2 and BMP4 and the decline of Gja1, ameloblastin and noggin in ERM. Thus, the ERM modulate cell proliferation and apoptosis, and inhibit differentiation by reducing expression of Gja1 under mechanical stretching.

External genitalia development in mice involves multiple developmental processes under the regulation of various signaling pathways. Wnt5a, one of the major Wnt ligands, is a crucial developmental regulator of outgrowing organs such as the limb, the mandible, and the external genitalia. Defects in Wnt5a signaling have been linked to Robinow syndrome, a genetic disorder in which male patients manifest a micropenis and defective urethral tube formation. Whereas Wnt5a is required for cell proliferation during embryonic external genitalia outgrowth, its role for urethral tube formation has yet to be understood. Here, we show that Wnt5a contributes to urethral tube formation as well as external genitalia outgrowth. Wnt5a is expressed in the embryonic external genitalia mesenchyme, and mesenchymal‐specific conditional Wnt5a knockout mice resulted in hypospadias‐like urethral defects. Early deletion of Wnt5a at E10.5 showed severe defects in both external genitalia outgrowth and urethral tube formation, along with reduced cell proliferation. The severe urethral tube defect persisted during later timing deletion of Wnt5a (E13.5). Further analyses revealed that loss of Wnt5a disrupted cell polarity and led to a reduction of the phosphorylated myosin light chain and the focal adhesion protein, vinculin. Altogether, these results suggest that Wnt5a coordinates cell proliferation and directed cell migration in a stage‐dependent manner during male external genitalia development. Furthermore, Wnt5a may regulate cell polarity, focal adhesion formation, and cell contractility, leading to directed cell migration during male‐type urethral formation in a manner that has not been reported in other organ fusion events. [ABSTRACT FROM AUTHOR]

Growth factor-dependent epithelial morphological changes and proliferation are essential for the formation of tubular structures, but the underlying molecular mechanisms are poorly understood. Co-stimulation with Wnt3a and epidermal growth factor (Wnt3a/ EGF) induced development of tubes consisting of intestinal epithelial cells by inducing expression of Arl4c, an Arf-like small GTP-binding protein, in three-dimensional culture, while stimulation with Wnt3a or EGF alone did not. Arl4c expression resulted in rearrangement of the cytoskeleton through activation of Rac and inactivation of Rho properly, which promoted cell growth by inducing nuclear translocation of Yes-associated protein and transcriptional co-activator with PDZ-binding motif ( YAP/ TAZ) in leading cells. Arl4c was expressed in ureteric bud tips and pretubular structures in the embryonic kidney. In an organoid culture assay, Wnt and fibroblast growth factor signaling simultaneously induced elongation and budding of kidney ureteric buds through Arl4c expression. YAP/ TAZ was observed in the nucleus of extending ureteric bud tips. Thus, Arl4c expression induced by a combination of growth factor signaling mechanisms is involved in tube formation. [ABSTRACT FROM AUTHOR]

Wnts compromise a large family of secreted and hydrophobic glycoproteins that control a variety of developmental and adult processes in all metazoan organisms. Recent advances in the field of Wnt signalling have revealed that Wnt activates multiple intracellular cascades, resulting in the regulation of cellular proliferation, differentiation, migration and polarity. However, it is not clear how Wnt activates these pathways after it binds to the receptors. It has been shown that Wnt and its antagonist Dickkopf are internalized with their receptors. This review highlights distinct endocytic pathways correlate with specificity of Wnt signalling events. [ABSTRACT FROM PUBLISHER]

Glycogen synthase kinase-3 (GSK-3) mediates epidermal growth factor, insulin and Wnt signals to various downstream events such as glycogen metabolism, gene expression, proliferation and differentiation. We have isolated here a GSK-3β-interacting protein from a rat brain cDNA library using a yeast twohybrid method. This protein consists of 832 amino acids and possesses Regulators of G protein Signaling (RGS) and dishevelled (Dsh) homologous domains in its N- and C-terminal regions, respectively. The predicted amino acid sequence of this GSK-3β-interacting protein shows 94% identity with mouse Axin, which recently has been identified as a negative regulator of the Wnt signaling pathway; therefore, we termed this protein rAxin (rat Axin). rAxin interacted directly with, and was phosphorylated by, GSK-3β. rAxin also interacted directly with the armadillo repeats of β-catenin. The binding site of rAxin for GSK-3β was distinct from the β-catenin-binding site, and these three proteins formed a ternary complex. Furthermore, rAxin promoted GSK-3β-dependent phosphorylation of β-catenin. These results suggest that rAxin negatively regulates the Wnt signaling pathway by interacting with GSK-3β and β-catenin and mediating the signal from GSK-3β to β-catenin. [ABSTRACT FROM AUTHOR]

The β-catenin mutation is frequently observed in hepatoblastoma (HB), but the underlying mechanism by which Wnt/β-catenin signaling induces HB tumor formation is unknown. Here we show that expression of growth regulation by estrogen in breast cancer 1 (GREB1) depends on Wnt/β-catenin signaling in HB patients. GREB1 is localized to the nucleus where it binds Smad2/3 in a competitive manner with p300 and inhibits TGFβ signaling, thereby promoting HepG2 HB cell proliferation. Forced expression of β-catenin, YAP, and c-Met induces HB-like mouse liver tumor (BYM mice), with an increase in GREB1 expression and HB markers. Depletion of GREB1 strongly suppresses marker gene expression and HB-like liver tumorigenesis, and instead enhances TGFβ signaling in BYM mice. Furthermore, antisense oligonucleotides for GREB1 suppress the formation of HepG2 cell-induced tumors and HB-like tumors in vivo. We propose that GREB1 is a target molecule of Wnt/β-catenin signaling and required for HB progression. The mechanisms promoting hepatoblastoma (HB) progression through Wnt/β-catenin signaling are unclear. Here, the authors show that the Wnt/ β-catenin axis induces GREB1 expression and nuclear localization, and suppresses TGFβ pathway, and propose GREB1 as a therapeutic target in HB. [ABSTRACT FROM AUTHOR]

Wnt5a activates the Wnt/β-catenin-independent pathway and its overexpression is associated with tumor aggressiveness enhancing invasive activity. For this action, Wnt5a-induced receptor endocytosis with clathrin is required. Wnt5a expression was previously believed to be associated with cancer cell motility but not proliferation. Recently, it was reported that Wnt5a is also implicated in cancer cell proliferation, but the mechanism was not clear. In this study, we generated a neutralizing anti-Wnt5a monoclonal antibody (mAb5A16) to investigate the mechanism by which Wnt5a regulates cancer cell proliferation. Wnt5a stimulated both invasion and proliferation of certain types of cancer cells, including HeLaS3 cervical cancer cells and A549 lung cancer cells although Wnt5a promoted invasion but not proliferation in other cancer cells such as KKLS gastric cancer cells. mAb5A16 did not affect the binding of Wnt5a to its receptor, but it suppressed Wnt5a-induced receptor-mediated endocytosis. mAb5A16 inhibited invasion but not proliferation of HeLaS3 and A549 cells. Wnt5a activated Src family kinases (SFKs) and Wnt5a-dependent cancer cell proliferation was dependent on SFKs, yet blockade of receptor-mediated endocytosis did not affect cancer cell proliferation and SFK activity. These results suggest that Wnt5a promotes invasion and proliferation of certain types of cancer cells through receptor-mediated endocytosis-dependent and -independent mechanisms, respectively. [ABSTRACT FROM AUTHOR]