The Netrin-1-Neogenin-1 signaling axis controls neuroblastoma cell migration via integrin-β1 and focal adhesion kinase activation.
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ABSTRACT:
SUBMITTER: Villanueva AA
PROVIDER: S-EPMC7971226 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Neogenin-1 (NEO1) is a transmembrane receptor involved in axonal guidance, angiogenesis, neuronal cell migration and cell death, during both embryonic development and adult homeostasis. It has been described as a dependence receptor, because it promotes cell death in the absence of its ligands (Netrin and Repulsive Guidance Molecule (RGM) families) and cell survival when they are present. Although NEO1 and its ligands are involved in tumor progression, their precise role in tumor cell survival and migration remain unclear. Public databases contain extensive information regarding the expression of NEO1 and its ligands Netrin-1 (NTN1) and Netrin-4 (NTN4) in primary neuroblastoma (NB) tumors. Analysis of this data revealed that patients with high expression levels of both NEO1 and NTN4 have a poor survival rate. Accordingly, our analyses in NB cell lines with different genetic backgrounds revealed that knocking-down NEO1 reduces cell migration, whereas silencing of endogenous NTN4 induced cell death. Conversely, overexpression of NEO1 resulted in higher cell migration in the presence of NTN4, and increased apoptosis in the absence of ligand. Increased apoptosis was prevented when utilizing physiological concentrations of exogenous Netrin-4. Likewise, cell death induced after NTN4 knock-down was rescued when NEO1 was transiently silenced, thus revealing an important role for NEO1 in NB cell survival. In vivo analysis, using the chicken embryo chorioallantoic membrane (CAM) model, showed that NEO1 and endogenous NTN4 are involved in tumor extravasation and metastasis. Our data collectively demonstrate that endogenous NTN4/NEO1 maintain NB growth via both pro-survival and pro-migratory molecular signaling.
| S-EPMC5354769 | biostudies-literature
Project description:SALL2/Sall2 is a transcription factor associated with development, neuronal differentiation, and cancer. Interestingly, SALL2/Sall2 deficiency leads to failure of the optic fissure closure and neurite outgrowth, suggesting a positive role for SALL2/Sall2 in cell migration. However, in some cancer cells, SALL2 deficiency is associated with increased cell migration. To further investigate the role of Sall2 in the cell migration process, we used immortalized Sall2 knockout (Sall2 -/- ) and Sall2 wild-type (Sall2 +/+ ) mouse embryonic fibroblasts (iMEFs). Our results indicated that Sall2 positively regulates cell migration, promoting cell detachment and focal adhesions turnover. Sall2 deficiency decreased cell motility and altered focal adhesion dynamics. Accordingly, restoring Sall2 expression in the Sall2 -/- iMEFs by using a doxycycline-inducible Tet-On system recovered cell migratory capabilities and focal adhesion dynamics. In addition, Sall2 promoted the autophosphorylation of Focal Adhesion Kinase (FAK) at Y397 and increased integrin β1 mRNA and its protein expression at the cell surface. We demonstrated that SALL2 increases ITGB1 promoter activity and binds to conserved SALL2-binding sites at the proximal region of the ITGB1 promoter, validated by ChIP experiments. Furthermore, the overexpression of integrin β1 or its blockade generates a cell migration phenotype similar to that of Sall2 +/+ or Sall2 -/- cells, respectively. Altogether, our data showed that Sall2 promotes cell migration by modulating focal adhesion dynamics, and this phenotype is associated with SALL2/Sall2-transcriptional regulation of integrin β1 expression and FAK autophosphorylation. Since deregulation of cell migration promotes congenital abnormalities, tumor formation, and spread to other tissues, our findings suggest that the SALL2/Sall2-integrin β1 axis could be relevant for those processes.
| S-EPMC9682130 | biostudies-literature
Project description:Neuroblastoma (NB) is the most common pediatric extracranial solid tumor. It arises during development of the sympathetic nervous system. Netrin-4 (NTN4), a laminin-related protein, has been proposed as a key factor to target NB metastasis, although there is controversy about its function. Here, we show that NTN4 is broadly expressed in tumor, stroma and blood vessels of NB patient samples. Furthermore, NTN4 was shown to act as a cell adhesion molecule required for the migration induced by Neogenin-1 (NEO1) in SK-N-SH neuroblastoma cells. Therefore, we propose that NTN4, by forming a ternary complex with Laminin γ1 (LMγ1) and NEO1, acts as an essential extracellular matrix component, which induces the migration of SK-N-SH cells.
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Project description:Directional cell migration drives embryonic development, cancer metastasis, and tissue repair and regeneration. Here, we examine the role of intraflagellar transport (IFT) 20 (Ift20) during polarized migration of epidermal cells. IFT20 is implicated in regulating cell migration independently of the primary cilium, but how IFT proteins integrate with the cell migration machinery is poorly understood. We show that genetic ablation of IFT20 in vitro slows keratinocyte migration during wound healing. We find that this phenotype is independent of the primary cilium and instead can be attributed to alterations in integrin-mediated mechanotransduction and focal adhesion (FA) dynamics. Loss of Ift20 resulted in smaller and less numerous FAs and reduced the levels of activated FA kinase. Studies of FA dynamics during microtubule-induced FA turnover demonstrated that Ift20 loss specifically impaired the reformation, but not the disassembly, of FAs. In the absence of Ift20 function, β1 integrins endocytosed during FA disassembly are not transferred out of Rab5 (+) endosomes. This defective transit from the early endosome disrupts eventual recycling of β1 integrins back to the cell surface, resulting in defective FA reformation. In vivo, conditional ablation of Ift20 in hair follicle stem cells (HF-SCs) similarly impairs their ability to invade and migrate during epidermal wound healing. Using explant studies, lineage tracing, and clonal analysis, we demonstrate that Ift20 is required for HF-SC migration and their contribution to epidermal regeneration. This work identifies a new Ift20 mechanotrafficking mechanism required for polarized cell migration and stem cell-driven tissue repair.
| S-EPMC7525813 | biostudies-literature
Project description:To search for factors promoting bone fracture repair, we investigated the effects of extracorporeal shock wave (ESW) on the adhesion, spreading, and migration of osteoblasts and its specific underlying cellular mechanisms. After a single period of stimulation by 10 kV (500 impulses) of shock wave (SW), the adhesion rate was increased as compared with the vehicle control. The data from both wound healing and transwell tests confirmed an acceleration in the migration of osteoblasts by SW treatment. RT-PCR, flow cytometry, and Western blotting showed that SW rapidly increased the surface expression of α5 and β1 subunit integrins, indicating that integrin β1 acted as an early signal for ESW-induced osteoblast adhesion and migration. It has also been found that a significant elevation occurred in the expression of phosphorylated β-catenin and focal adhesion kinase (FAK) at the site of tyrosine 397 in response to SW stimulation after the increasing expression of the integrin β1 molecule. When siRNAs of integrin α5 and β1 subunit were added, the level of FAK phosphorylation elevated by SW declined. Interestingly, the adhesion and migration of osteoblasts were decreased when these siRNA reagents as well as the ERK1/2 signaling pathway inhibitors, U0126 and PD98059, were present. Further studies demonstrated that U0126 could inhibit the downstream integrin-dependent signaling pathways, such as the FAK signaling pathway, whereas it had no influence on the synthesis of integrin β1 molecule. In conclusion, these data suggest that ESW promotes the adhesion and migration of osteoblasts via integrin β1-mediated expression of phosphorylated FAK at the Tyr-397 site; in addition, ERK1/2 are also important for osteoblast adhesion, spreading, migration, and integrin expression.
| S-EPMC3406705 | biostudies-literature
Project description:The motility of blood monocytes is orchestrated by the activity of cell-surface integrins, which translate extracellular signals into cytoskeletal changes to mediate adhesion and migration. Toxoplasma gondii is an intracellular parasite that infects migratory cells and enhances their motility, but the mechanisms underlying T. gondii-induced hypermotility are incompletely understood. We investigated the molecular basis for the hypermotility of primary human peripheral blood monocytes and THP-1 cells infected with T. gondii Compared with uninfected monocytes, T. gondii infection of monocytes reduced cell spreading and the number of activated β1 integrin clusters in contact with fibronectin during settling, an effect not observed in monocytes treated with lipopolysaccharide (LPS) or Escherichia coli Furthermore, T. gondii infection disrupted the phosphorylation of focal adhesion kinase (FAK) at tyrosine 397 (Tyr-397) and Tyr-925 and of the related protein proline-rich tyrosine kinase (Pyk2) at Tyr-402. The localization of paxillin, FAK, and vinculin to focal adhesions and the colocalization of these proteins with activated β1 integrins were also impaired in T. gondii-infected monocytes. Using time-lapse confocal microscopy of THP-1 cells expressing enhanced GFP (eGFP)-FAK during settling on fibronectin, we found that T. gondii-induced monocyte hypermotility was characterized by a reduced number of enhanced GFP-FAK-containing clusters over time compared with uninfected cells. This study demonstrates an integrin conformation-independent regulation of the β1 integrin adhesion pathway, providing further insight into the molecular mechanism of T. gondii-induced monocyte hypermotility.
| S-EPMC5836128 | biostudies-literature
Project description:BackgroundRecycling of integrin via endosomal vesicles is critical for the migration of cancer cells, which leads to the metastasis of pancreatic cancer and devastating cancer-related death. So, new diagnostic and therapeutic molecules which target the recycling of endosomal vesicles need to be developed.MethodsPublic databases including TCGA, ICGC, GSE21501, GSE28735, and GENT are analyzed to derive diagnostic and therapeutic targets. To reveal biological roles and underlying mechanisms of molecular targets, various molecular biological experiments were conducted.ResultsFirst, we identified UNC13D's overexpression in patients with pancreatic cancer (n = 824) and its prognostic significance and high hazard ratio (HR) in four independent pancreatic cancer cohorts (TCGA, n = 178, p = 0.014, HR = 3.629; ICGC, n = 91, p = 0.000, HR = 4.362; GSE21501, n = 102, p = 0.002, HR = 2.339; GSE28735, n = 45, p = 0.022, HR = 2.681). Additionally, its expression is associated with the clinicopathological progression of pancreatic cancer. Further biological studies have shown that UNC13D regulates the migration of pancreatic cancer cells by coupling the exocytosis of recycling endosomes with focal adhesion turnover via the regulation of FAK phosphorylation. Immunoprecipitation and immunocytochemistry showed the formation of the RAB11-UNC13D-FAK axis in endosomes during integrin recycling. We observed that UNC13D directly interacted with the FERM domain of FAK and regulated FAK phosphorylation in a calcium-dependent manner. Finally, we found co-expression of UNC13D and FAK showed the poorest survival (TCGA, p = 0.000; ICGC, p = 0.036; GSE28735, p = 0.006).ConclusionsWe highlight that UNC13D, a novel prognostic factor, promotes pancreatic cancer progression by coupling integrin recycling with focal adhesion turnover via the RAB11-UNC13D-FAK axis for the migration of pancreatic cancer cells.
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Project description:The fibronectin (FN)-binding integrins alpha4beta1 and alpha5beta1 confer different cell adhesive properties, particularly with respect to focal adhesion formation and migration. After analyses of alpha4+/alpha5+ A375-SM melanoma cell adhesion to fragments of FN that interact selectively with alpha4beta1 and alpha5beta1, we now report two differences in the signals transduced by each receptor that underpin their specific adhesive properties. First, alpha5beta1 and alpha4beta1 have a differential requirement for cell surface proteoglycan engagement for focal adhesion formation and migration; alpha5beta1 requires a proteoglycan coreceptor (syndecan-4), and alpha4beta1 does not. Second, adhesion via alpha5beta1 caused an eightfold increase in protein kinase Calpha (PKCalpha) activation, but only basal PKCalpha activity was observed after adhesion via alpha4beta1. Pharmacological inhibition of PKCalpha and transient expression of dominant-negative PKCalpha, but not dominant-negative PKCdelta or PKCzeta constructs, suppressed focal adhesion formation and cell migration mediated by alpha5beta1, but had no effect on alpha4beta1. These findings demonstrate that different integrins can signal to induce focal adhesion formation and migration by different mechanisms, and they identify PKCalpha signaling as central to the functional differences between alpha4beta1 and alpha5beta1.
| S-EPMC2172880 | biostudies-other
Project description:Integrins are membrane bound receptors that regulate several cellular processes, such as cell adhesion, migration, survival and proliferation, and may contribute to tumor initiation/progression in cells exposed to genotoxic stress. The extent of integrin activation and its role in cell survival upon intoxication with bacterial genotoxins are still poorly characterized. These toxins induce DNA strand breaks in the target cells and activate the DNA damage response (DDR), coordinated by the Ataxia Telangectasia Mutated (ATM) kinase. In the present study, we demonstrate that induction of DNA damage by two bacterial genotoxins promotes activation of integrin β1, leading to enhanced assembly of focal adhesions and cell spreading on fibronectin, but not on vitronectin. This phenotype is mediated by an ATM-dependent inside-out integrin signaling, and requires the actin cytoskeleton remodeler NET1. The toxin-mediated cell spreading and anchorage-independent survival further relies on ALIX and TSG101, two components of the endosomal sorting complex required for transport (ESCRT), known to regulate integrin intracellular trafficking. These data reveal a novel aspect of the cellular response to bacterial genotoxins, and provide new tools to understand the carcinogenic potential of these effectors in the context of chronic intoxication and infection.
| S-EPMC4395369 | biostudies-literature
Project description:Given that integrin β1 is an important component of the connection to maintain glomerular structural integrity, by binding with multiple extracellular matrix proteins and mediating intracellular signaling. Focal adhesion kinase (FAK) is the most essential intracellular integrator in the integrin β1-FAK signalling pathway. Here, we investigated the changes of the two molecules and visualized the possible interaction between them under various hemodynamic conditions in podocytes. Mice kidney tissues were prepared using in vivo cryotechnique (IVCT) and then were stained and observed using light microscopy, confocal laser scanning microscopy and immunoelectron microscopy. The expression of these molecules were examined by western blot. Under the normal condition, integrin β1 stained continually and evenly at the membrane, and FAK was located in the cytoplasm and nuclei of the podocytes. There were significant colocalized plaques of two molecules. But under acute hypertensive and cardiac arrest conditions, integrin β1 decreased and stained intermittently. Similarly, FAK decreased and appeared uneven. Additionally, FAK translocated to the nuclei of the podocytes. As a result, the colocalization of integrin β1 and FAK reduced obviously under these conditions. Western blot assay showed a consistent result with the immunostaining. Collectively, the abnormal redistribution and decreased expressions of integrin β1 and FAK are important molecular events in regulating the functions of podocytes under abnormal hemodynamic conditions. IVCT could offer considerable advantages for morphological analysis when researching renal diseases.
| S-EPMC3976409 | biostudies-literature