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Periventricular nodular heterotopia (PVNH) is an X-linked disease caused by loss-of-function variants in the filamin A (

Apoptosis is a critical process for the maintenance of cell populations, and involves mitochondrial depolarization, the sequential cleavage of caspase-9 and -3, followed by the externalization of phosphatidylserine (PS) on the plasma membrane. The actin cytoskeleton and its accessory proteins are known regulators of apoptotic signaling in nucleated cells but their roles in platelet apoptosis are undefined. Filamin A (FLNA) is a ubiquitously expressed actin-crosslinking protein that also serves as an intracellular signaling scaffold. Here we used platelets from mice with a platelet-specific FLNA deficiency (Flnafl/Y, Pf4-cre/+, termed platelet-specific knockout) to test the role of FLNA in platelet apoptosis. Treatment with the BH3-mimetic drug ABT-737 induced caspase-3 cleavage and PS exposure in platelets from floxed mice (Flnafl/Y, termed control) but these effects were essentially abrogated in FLNA-null platelets (platelet-specific knockout). Protein kinase C (PKC), a known FLNA ligand, was also activated by ABT-737, and PKC's phosphorylation of its downstream substrates was attenuated in FLNA-null platelets. The PKC inhibitor bisindolylmaleimide (BIM) also reduced caspase-3 cleavage, thus essentially phenocopying the FLNA-null platelets. Notably, the caspase-3 cleavage defect in FLNA-null platelets was rescued by the PKC-activating phorbol ester PMA, suggesting that FLNA and PKC share a common pathway in regulating platelet apoptosis. Mitochondrial depolarization and caspase-9 cleavage were unaffected by BIM treatment, suggesting that PKC specifically controls the downstream caspase-3 point of the pro-apoptotic signaling pathway. These data point to a novel role for FLNA in the regulation of platelet apoptosis, thus providing an improved understanding of how circulating platelet counts are maintained.

TheWe identified rare, putativeSixty individuals (0.03%; median age 58 years [47-70 interquartile range], 43% male) harbored 27 unique

Small heat shock proteins (sHsps) play an important role in the maintenance of proteome stability and, particularly, in stabilization of the cytoskeleton and cell contractile apparatus. Cell exposure to different types of stress is accompanied by the translocation of sHsps onto actin filaments; therefore, it is commonly believed that the sHsps are true actin-binding proteins. Investigations of last years have shown that this assumption is incorrect. Stress-induced translocation of sHsp to actin filaments is not the result of direct interaction of these proteins with intact actin, but results from the chaperone-like activity of sHsps and their interaction with various actin-binding proteins. HspB1 and HspB5 interact with giant elastic proteins titin and filamin thus providing an integrity of the contractile apparatus and its proper localization in the cell. HspB6 binds to the universal adapter protein 14-3-3 and only indirectly affects the structure of actin filament. HspB7 interacts with filamin C and controls actin filament assembly. HspB8 forms tight complex with the universal regulatory and adapter protein Bag3 and participates in the chaperone-assisted selective autophagy (CASA) of actin-binding proteins (e.g., filamin), as well as in the actin-depending processes taking place in mitoses. Hence, the mechanisms of sHsp participation in the maintenance of the contractile apparatus and cytoskeleton are much more complicated and diverse than it has been postulated earlier and are not limited to direct interactions of sHsps with actin. The old hypothesis on the direct binding of sHsps to intact actin should be revised and further detailed investigation on the sHsp interaction with minor proteins participating in the formation and remodeling of actin filaments is required.

Pathogenic variants of the X-linked FLNA gene encoding filamin A protein have been associated with a wide spectrum of symptoms, including the recently described pulmonary phenotype with childhood-onset panlobular emphysema. We describe three female patients from two families with novel heterozygous FLNA variants c.5837_2del and c.508C > T. Analysis of immunofluorescence of peripheral blood smears and platelet function was performed for all patients. FLNA-negative platelets were observed, suggesting that these variants result in the loss of a functional protein product. All three patients also had periventricular nodular heterotopia and panlobular emphysema. However, they had considerably milder symptoms and later age of onset than in the previously reported cases. Therefore, patients with pathogenic FLNA variants should be studied actively for lung involvement even in the absence of pronounced respiratory symptoms. Conversely, any patient with unexplained panlobular emphysema should be analyzed for pathogenic FLNA variants. We also suggest that immunofluorescence analysis is a useful tool for investigating the pathogenicity of novel FLNA variants.

Tripartite motif (TRIM) proteins have been proved to contribute to cancer progression, while whether tripartite motif-containing 54 (TRIM54) could functionally influence gastric cancer (GC) progression remains elusive.The expression level of TRIM54 and filamin C (FLNC) in GC was determined by Western blot and online database. Cell Counting Kit-8 (CCK-8) assay, colony formation assay and Ethylenediurea (EdU) staining were performed to explore the effects of TRIM54 on GC cell proliferation. Transwell assay and wound healing assay were applied to detect the influence of TRIM54 on GC cell migration and invasion. Bioinformatics analysis and Co-immunoprecipitation assay (Co-Ip), Ubiquitination assay and Half-life assay were involved to explore the regulatory mechanism of TRIM54 on FLNC.TRIM54 was upregulated in GC tissues and cells, and a higher expression level of TRIM54 indicated a shorter overall survival of GC patients. The overexpression of TRIM54 significantly enhanced proliferation, migration, and invasion of GC cells, and inhibition of TRIM54 expression exerted reverse effects on GC cells. Mechanistically, TRIM54 was determined as a post-translational mediator of FLNC, and TRIM54 was co-immunoprecipitated with FLNC and degraded its protein level via K63-linked ubiquitination of FLNC. Notably, FLNC efficiently inhibited GC progression by TRIM54 overexpression.Collectively, our findings suggested that the TRIM54/FLNC axis could be considered as a potential prognostic biomarker for GC.

Tripartite motif-containing 44 (TRIM44) has recently been implicated in various pathological processes in numerous cancers, including lung adenocarcinoma (LUAD); however, its functional roles in chemoresistance are poorly understood. Herein, TRIM44 knockdown sensitized LUAD cells to cisplatin and enhanced cisplatin-induced apoptosis. Microarray analysis indicated that the "

The γ-aminobutyric acid type B (GABAB) receptor is a prototypical family C G protein-coupled receptor (GPCR) that plays a key role in the regulation of synaptic transmission. Although growing evidence suggests that GPCR signaling in neurons might be highly organized in time and space, limited information is available about the mechanisms controlling the nanoscale organization of GABAB receptors and other GPCRs on the neuronal plasma membrane. Using a combination of biochemical assays in vitro, single-particle tracking, and super-resolution microscopy, we provide evidence that the spatial organization and diffusion of GABAB receptors on the plasma membrane are governed by dynamic interactions with filamin A, which tethers the receptors to sub-cortical actin filaments. We further show that GABAB receptors are located together with filamin A in small nanodomains in hippocampal neurons. These interactions are mediated by the first intracellular loop of the GABAB1 subunit and modulate the kinetics of Gαi protein activation in response to GABA stimulation.

Pediatric restrictive cardiomyopathy (RCM) is the rarest in its group and accounts for only 2.5-5% of all the diagnosed cardiomyopathies in children. It is a relentless disease with poor prognosis, and heart transplantation is the only long-term treatment option. The aetiology of pediatric RCM varies and includes conditions such as endomyocardial fibrosis, storage disorder (Fabry's disease, MPS), drugs, radiation, post-cardiac transplantation and genetic. Genetic causes encompasses mutations in sarcomeric (troponin I and T, actin, myosin and titin) and nonsarcomeric protein-coding genes (Desmin, RSK2, lamin A/C and bcl-2-associated athanogene 3 (

Most proteins begin to fold during biosynthesis on the ribosome. It has been suggested that interactions between the emerging polypeptide and the ribosome surface might allow the ribosome itself to modulate co-translational folding. Here we combine protein engineering and NMR spectroscopy to characterize a series of interactions between the ribosome surface and unfolded nascent chains of the immunoglobulin-like FLN5 filamin domain. The strongest interactions are found for a C-terminal segment that is essential for folding, and we demonstrate quantitative agreement between the strength of this interaction and the energetics of the co-translational folding process itself. Mutations in this region that reduce the extent of binding result in a shift in the co-translational folding equilibrium towards the native state. Our results therefore demonstrate that a competition between folding and binding provides a simple, dynamic mechanism for the modulation of co-translational folding by the ribosome., During polypeptide biosynthesis, a strong interaction can occur between a segment of an emerged, disordered nascent protein and the ribosomal surface. Now, it has been shown that competition between this ribosomal binding and the folding energetics of an immunoglobulin-like domain modulates the mechanism of co-translational folding.

Rationale: Dominant heterozygous variants in filamin C ( FLNC ) cause diverse cardiomyopathies, although the underlying molecular mechanisms remain poorly understood. Objective: We aimed to define the molecular mechanisms by which FLNC variants altered human cardiomyocyte gene and protein expression, sarcomere structure, and contractile performance. Methods and Results: Using CRISPR/Cas9, we introduced FLNC variants into human induced pluripotent stem cell–derived cardiomyocytes (hiPSC-CMs). We compared isogenic hiPSC-CMs with normal (wild-type), ablated expression ( FLNC −/− ), or haploinsufficiency ( FLNC +/− ) that causes dilated cardiomyopathy. We also studied a heterozygous in-frame deletion ( FLNC +/Δ7aa ) which did not affect FLNC expression but caused aggregate formation, similar to FLNC variants associated with hypertrophic cardiomyopathy. FLNC −/− hiPSC-CMs demonstrated profound sarcomere misassembly and reduced contractility. Although sarcomere formation and function were unaffected in FLNC +/ − and FLNC +/Δ7aa hiPSC-CMs, these heterozygous variants caused increases in lysosome content, enhancement of autophagic flux, and accumulation of FLNC-binding partners and Z-disc proteins. Conclusions: FLNC expression is required for sarcomere organization and physiological function. Variants that produce misfolded FLNC proteins cause the accumulation of FLNC and FLNC-binding partners which leads to increased lysosome expression and activation of autophagic pathways. Surprisingly, similar pathways were activated in FLNC haploinsufficient hiPSC-CMs, likely initiated by the loss of stoichiometric FLNC protein interactions and impaired turnover of proteins at the Z-disc. These results indicate that both FLNC haploinsufficient variants and variants that produce misfolded FLNC protein cause disease by similar proteotoxic mechanisms and indicate the therapeutic potential for augmenting protein degradative pathways to treat a wide range of FLNC -related cardiomyopathies.

Fibroblast migration is closely regulated by the mechanical characteristics in surrounding microenvironment. While increased interstitial hydrostatic pressure (HP) is a hallmark in many pathological and physiological conditions, little is known about how the HP affects fibroblast motility. Using cell-culture chips with elevated HP conditions, we showed that 20 cmH2O HP significantly accelerated fibroblast migration. The HP-induced migration acceleration was dependent on the augmentation of transforming growth factor-β1, and correlated with the activation of filamin A via the phosphorylation of p38 mitogen-activated protein kinase. Our results suggest that interstitial HP elevation associated with various pathological states could significantly regulate fibroblast migration.

Sinus of Valsalva aneurysm (SVA) is a rare cardiovascular disease with male predominance. Recently, an association with aortic aneurysm and SVA has been revealed in periventricular nodular heterotopia patients with loss-of-function Filamin A (FLNA) mutations, which were located on chromosome X and almost exclusively affect females.Among patients hospitalized for aortic surgery with aortic root diameter over 4.0 cm, next-generation sequencing was performed to investigate 30 candidate genes related to inherited aortic aneurysm syndromes and familial thoracic aortic aneurysm and dissection. The present report reviewed an electronic case database and identified two female cases of unruptured SVA with heterozygous FLNA truncating mutations.Case 1 displaying a rare SVA phenotype involving left and noncoronary sinus harbored a nonsense variant p.Tyr1720Ter/c.5160CG. Case 2 displayed right and noncoronary SVA with predominantly enlarged right coronary sinus, posterior mitral valve prolapse, and harbored a frameshift variant p.Val1724fs*68/c.5171_5172delTG. Both novel mutations resulted in the premature termination of filamin A with the loss of functional Rod 2 and dimerization region.The present report raised the possibility of the presence of a cardiovascular onset form in the spectrum of FLNA hereditary diseases. The association between SVA and loss-of-function FLNA mutations indicates a unique etiology and pathogenesis among female patients, which requires further investigation to establish the linkage between FLNA variants and a wide spectrum of phenotypes.

Filamins are large proteins with actin-binding properties. Mutations in FLNC, one of the three filamin genes in humans, have recently been implicated in dominant cardiomyopathies, but the underlying mechanisms are not well understood. Here, we aimed to use Drosophila melanogaster as a new in vivo model to study these diseases. First, we show that adult-specific cardiac RNAi-induced depletion of Drosophila Filamin (dFil) induced cardiac dilatation, impaired systolic function and sarcomeric alterations, highlighting its requirement for cardiac function and maintenance of sarcomere integrity in the adult stage. Next, we introduced in the cheerio gene, using CRISPR/Cas9 gene editing, three missense variants, previously identified in patients with hypertrophic cardiomyopathy. Flies carrying these variants did not exhibit cardiac defects or increased propensity to form filamin aggregates, arguing against their pathogenicity. Finally, we show that deletions of the C-term part of dFil carrying the last four Ig-like domains are dispensable for cardiac function. Collectively, these results highlight the relevance of this model to explore the cardiac function of filamins and increase our understanding of physio-pathological mechanisms involved in FLNC-related cardiomyopathies. This article has an associated First Person interview with the first author of the paper.

INTRODUCTION: Filamin C (FLNC) gene variants associated with atrial cardiomyopathies have not been reported so far. The aim of this study was to assess the genetics of two siblings presenting with recurrent right atrial arrhythmias, severe right atrial dilatation, and skeletal myopathy. METHODS: A family with subjects affected by recurrent atrial arrhythmias and skeletal myopathy was extensively evaluated by the means of electrocardiographic recordings, magnetic resonance, intracardiac high-density mapping, and genetic testing. RESULTS: Two siblings with right atrial arrhythmias and severe right atrial disease were found to be heterozygous carriers of the variant FLNC-c.925G>A p.(Glu309Lys), previously reported as a variant of uncertain significance. Despite the presence of a severe dilatation of the right atrium in both patients, one presented with skeletal muscle myopathy and an atrial arrhythmia refractory to pharmacological and invasive treatment, while the other one did not have any myopathy, and rhythm control was easily achieved by drugs. CONCLUSION: Filamin C missense variant c.925G>A p.(Glu309Lys) is associated with the severe right atrial disease. Considering cosegregation with the disease (PP1 supporting), this variant should be classified as likely pathogenic.

Dilated cardiomyopathy (DCM) is a heterogenous group of disorders characterised by left ventricular dilatation and dysfunction, in the absence of factors affecting loading conditions such as hypertension or valvular disease, or significant coronary artery disease. The prevalence of idiopathic DCM is estimated between 1:250 and 1:500 individuals. Determining the aetiology of DCM can be challenging, particularly when evaluating an individual and index case with no classical history or investigations pointing towards an obvious acquired cause, or no clinical clues in the family history to suggest a genetic cause. We present a family affected by DCM associated with Filamin C variant, causing sudden cardiac death at a young age and heart failure due to severe left ventricular impairment and myocardial scarring. We review the diagnosis and treatment of DCM, its genetic associations and potential acquired causes. Thorough assessment is mandatory to risk stratify and identify patients who may benefit from primary prevention implantable cardioverter defibrillator therapy according to international guidelines. Genetic testing has some limitations, and is positive in only 20%–35% of DCM, but should be considered in specific cases to identify families who may benefit from cascade screening after appropriate counselling. The management of often complex familial cardiomyopathy requires specialist input for every case, and the appropriate infrastructure to coordinate investigations.

Cancer cells migrating in confined microenvironments exhibit plasticity of migration modes. Confinement of contractile cells in a nonadhesive environment drives “leader bleb–based migration” (LBBM), morphologically characterized by a long bleb that points in the direction of movement separated from a cell body by a contractile neck. Although cells undergoing LBBM have been visualized within tumors, the organization of organelles and actin regulatory proteins mediating LBBM is unknown. We analyzed the localization of fluorescent organelle-specific markers and actin-associated proteins in human melanoma and osteosarcoma cells undergoing LBBM. We found that organelles from the endolysosomal, secretory, and metabolic systems as well as the vimentin and microtubule cytoskeletons localized primarily in the cell body, with some endoplasmic reticulum, microtubules, and mitochondria extending into the leader bleb. Overexpression of fluorescently tagged actin regulatory proteins showed that actin assembly factors localized toward the leader bleb tip, contractility regulators and cross-linkers in the cell body cortex and neck, and cross-linkers additionally throughout the leader bleb. Quantitative analysis showed that excess filamin-A and fascin-1 increased migration speed and persistence, while their depletion by small interfering RNA indicates a requirement in promoting cortical tension and pressure to drive LBBM. This indicates a critical role of specific actin crosslinkers in LBBM.

Creating in vitro models of diseases of the pancreatic ductal compartment requires a comprehensive understanding of the developmental trajectories of pancreas-specific cell types. Here we report the single-cell characterization of the differentiation of pancreatic duct-like organoids (PDLOs) from human induced pluripotent stem cells (hiPSCs) on a microwell chip that facilitates the uniform aggregation and chemical induction of hiPSC-derived pancreatic progenitors. Using time-resolved single-cell transcriptional profiling and immunofluorescence imaging of the forming PDLOs, we identified differentiation routes from pancreatic progenitors through ductal intermediates to two types of mature duct-like cells and a few non-ductal cell types. PDLO subpopulations expressed either mucins or the cystic fibrosis transmembrane conductance regulator, and resembled human adult duct cells. We also used the chip to uncover ductal markers relevant to pancreatic carcinogenesis, and to establish PDLO co-cultures with stellate cells, which allowed for the study of epithelial-mesenchymal signalling. The PDLO microsystem could be used to establish patient-specific pancreatic duct models.

Cancer cells show increases in protein degradation pathways, including autophagy, during progression to meet the increased protein degradation demand and support cell survival. On the other hand, reduced autophagy activity during aging is associated with a reduced DNA damage response and increased genomic instability. Therefore, it is a puzzling how DNA repair can be increased in cancer cells that are resistant to chemotherapies or during progression when autophagy activity is intact or increased. We discovered that tripartite motif containing 44 (TRIM44) is a pivotal element regulating the DNA damage response in cancer cells with intact autophagy. TRIM44 deubiquitinates p62, an autophagy substrate, which leads to its oligomerization. This prevents p62 localization to the nucleus upon irradiation. Increased cytoplasmic retention of p62 by TRIM44 prevents the degradation of FLNA and 53BP1, which increases DNA damage repair. Together, our data support TRIM44 a potential therapeutic target for therapy-resistant tumor cells with intact autophagy.

Endothelial cells (ECs) respond to blood shear stress by changing their morphology is important for maintaining vascular homeostasis. Studies have documented a relationship between endothelial cell shape and the stress flow, and however, the mechanism underlying this cytoskeletal rearrangement due to shear stress remains uncertain. In this paper, we demonstrate that laminar shear stress (LSS) significantly reduces latexin (LXN) expression in ECs. By using siRNA and cell imaging, we demonstrated that LXN knockdown results in the morphologic change and F‐actin remodelling just like what LSS does in ECs. We further demonstrate that LXN interacts with Filamin A (FLNA) and regulates FLNA proteolytic cleavage and nuclei translocation. By constructing LXN‐/‐ mice and ApoE‐/‐LXN‐/‐ double knockout mice, we evaluated the effect of LXN knockout on aortic endothelium damage in mice. We found that LXN deficiency significantly improves vascular permeability, vasodilation and atherosclerosis in mice. Our findings provide confident evidence, for the first time, that LXN is a novel regulator for morphological maintenance of ECs, and LXN deficiency has a protective effect on vascular homeostasis. This provides new strategies and drug targets for the treatment of vascular diseases.

Dilated cardiomyopathy (DCM) is a heterogeneous cardiac disorder characterized by left ventricular dilatation and dysfunction. Mutations in dozens of cardiac genes have been connected to the development of DCM including the filamin C gene (FLNC). We generated two induced pluripotent stem cell (iPSCs) lines from DCM patients carrying single missense heterozygote FLNC mutations (c.6689G A and c.3745G A). Both lines expressed high levels of pluripotency markers, differentiated into derivatives of the three germ layers and possessed normal karyotypes. The derived iPSC lines can serve as powerful tools to model DCM in vitro and as a platform for therapeutic development.

Filaminopathies A are rare disorders affecting the brain, intestine, or skeleton, characterized by dominant X-linked filamin A (FLNA) gene mutations. Macrothrombocytopenia with functionally defective platelets is frequent. We have described a filaminopathy A male patient, exhibiting a C-terminal frame-shift FLNa mutation (Berrou et al., Arterioscler Thromb Vasc Biol. 2017;37:1087-1097). Contrasting with female patients, this male patient exhibited gain of platelet functions, including increased platelet aggregation, integrin αIIbβ3 activation, and secretion at low agonist concentration, raising the issue of thrombosis risk.Our goal is to assess the thrombotic potential of the patient FLNa mutation in an in vivo model.We have established a mutant FlnA knock-in mouse model.The mutant FlnA mouse platelets phenocopied patient platelets, showing normal platelet count, lower expression level of mutant FlnA, and gain of platelet functions: increased platelet aggregation, secretion, and αIIbβ3 activation, as well as increased spreading and clot retraction. Surprisingly, mutant FlnA mice exhibited a normal bleeding time, but with increased re-bleeding (77%) compared to wild type (WT) FlnA mice (27%), reflecting hemostatic plug instability. Again, in an in vivo thrombosis model, the occlusion time was not altered by the FlnA mutation, but arteriolar embolies were increased (7-fold more frequent in mutant FlnA mice versus WT mice), confirming thrombus instability.This study shows that the FlnA mutation found in the male patient induced gain of platelet functions in vitro, but thrombus instability in vivo. Implications for the role of FLNa in physiology of thrombus formation are discussed.

Patient-derived organoids and cellular spheroids recapitulate tissue physiology with remarkable fidelity. We investigated how engagement with a reconstituted basement membrane in three dimensions (3D) supports the polarized, stress resilient tissue phenotype of mammary epithelial spheroids. Cells interacting with reconstituted basement membrane in 3D had reduced levels of total and actin-associated filamin and decreased cortical actin tension that increased plasma membrane protrusions to promote negative plasma membrane curvature and plasma membrane protein associations linked to protein secretion. By contrast, cells engaging a reconstituted basement membrane in 2D had high cortical actin tension that forced filamin unfolding and endoplasmic reticulum (ER) associations. Enhanced filamin-ER interactions increased levels of PKR-like ER kinase effectors and ER-plasma membrane contact sites that compromised calcium homeostasis and diminished cell viability. Consequently, cells with decreased cortical actin tension had reduced ER stress and survived better. Consistently, cortical actin tension in cellular spheroids regulated polarized basement membrane membrane deposition and sensitivity to exogenous stress. The findings implicate cortical actin tension-mediated filamin unfolding in ER function and underscore the importance of tissue mechanics in organoid homeostasis.

Lung cancer is still a main cause of cancer-related death worldwide. Non-small-cell lung cancer (NSCLC) accounts for the majority of lung cancers, and gefitinib is an effective targeted drug for NSCLC. It is important to explore the underlying molecular mechanisms of gefitinib resistance to provide new treatment strategies and to improve the prognosis of gefitinib-resistant NSCLC patients. This study aimed to examine the role of filamin A (FLNA) in acquired resistance to gefitinib in NSCLC, and identify ANXA2 (annexin A2), one of calcium-dependent phospholipid-binding proteins, as its corresponding regulatory factor. First, we established resistant cells via long-term exposure to gefitinib to analyse the association between FLNA and gefitinib resistance. Through quantitative real-time polymerase chain reaction (qRT-PCR), Cell Counting Kit-8 (CCK-8), western blotting (WB), and flow cytometry assays, we evaluated the role of FLNA. The effect of FLNA knockdown or overexpression was analysed not only in cell lines but also in mouse models. We verified the FLNA-interacting protein through coimmunoprecipitation (CoIP) experiments and found that the downstream signalling pathway was regulated by FLNA and its interacting protein. Finally, the upstream transcription factor was identified by chromatin immunoprecipitation (ChIP). Increased FLNA expression induced gefitinib resistance. Knockdown of FLNA restored gefitinib sensitivity and induced apoptosis in vivo and in vitro. FLNA and ANXA2 cooperatively led to the activation of the Wnt pathway, which was closely linked to gefitinib resistance. Subsequently, SP1 promoted transcriptional activation of FLNA to regulate gefitinib resistance. We determined that FLNA serves as a regulator of gefitinib resistance in NSCLC and found that FLNA and ANXA2 together induced gefitinib resistance by activating the Wnt pathway.

Objective Tetralogy of Fallot (TOF) is one of the most common congenital abnormalities that need early intervention. Here, for the first time, we report a nonsyndromic form of TOF caused by a novel variant in the FLNA gene in 2 siblings of an Iranian family. Methods The family underwent a complete workup, including karyotyping, sequencing of 6 common genes in congenital heart diseases (GATA4, NKX2-5, ZIC3, FOXH1, NODAL, and GJA1), array comparative genomic hybridization, multiplex ligation-dependent probe amplification, and whole-exome sequencing. Segregation and in silico analysis were also conducted for the identified variant. Results A variant, c.3415C>T, in the FLNA gene was found in both affected brothers in this family; this variant was heterozygous in their mother. Bioinformatics tools predicted the variant as a pathogenic one. Conclusion Many allelic disorders have been reported for FLNA mutations. Mutations in this gene may cause a nonsyndromic congenital form of TOF.

Background Filamin A (FLNA) is an intracellular actin-binding protein, encoded by the FLNA gene, with a wide tissue expression. It is involved in several cellular functions, and extracellular matrix structuring. FLNA gene alterations lead to diseases with a wide phenotypic spectrum, such as brain periventricular nodular heterotopia (PVNH), cardiovascular abnormalities, skeletal dysplasia, and lung involvement. Clinical findings We present the case of a female infant who showed at birth aortic valve stenosis and PVNH, and subsequently developed interstitial lung disease with severe pulmonary hypertension. Primary diagnosis The association of aortic valve dysplasia, left ventricular outflow obstruction, persistent patent ductus arteriosus, and brain heterotopic gray matter suggested a possible FLNA gene alteration. A novel heterozygous intronic variant in the FLNA gene (NM_001110556.1), c.4304-1G >A, was detected. Interventions In consideration of valve morphology and severity of stenosis, the neonate was scheduled for a transcatheter aortic valvuloplasty. At 3 months of life, she developed hypoxemic respiratory failure with evidence of severe pulmonary hypertension. Inhaled nitric oxide (iNO) and milrinone on continuous infusion were started. Because of a partial response to iNO, an intravenous continuous infusion of sildenafil was introduced. Outcomes In consideration of severe clinical course and fatal outcome, the new FLNA gene mutation described in our patient seems to be associated with a loss of function of FLNA. Practice recommendations Lung and brain involvement, in association with left ventricular outflow obstruction and persistent patency of ductus arteriosus, should be considered highly suggestive of FLNA gene alterations, in a female newborn.

Background: Interstitial lung disease (ILD) has been recently reported in a few patients with pathogenic variants in the Filamin A (FLNA) gene with variable presentation and prognosis. This study evaluated the respiratory manifestations and clinical features in children with FLNA disease. Methods: We conducted a retrospective review of pediatric patients with variants in FLNA in a tertiary children's hospital. The clinical features, genotype, management, and outcomes were analyzed. Results: We identified 9 patients with variants in FLNA aged 15 months to 24 years, 4 females and 5 males. Six patients had abnormal chest imaging ranging from mild interstitial prominence to atelectasis, interstitial densities, and hyperinflation. Three patients with ILD presented during the neonatal period or early infancy with respiratory distress or respiratory failure requiring supplemental oxygen or assisted ventilation via tracheostomy. We report male twins with the same FLNA variant and lung disease, but different ages and clinical features at presentation eventually culminating in respiratory failure requiring assisted ventilation. All patients had FLNA variants identified by FLNA sequencing, had abnormal echocardiograms, and none of the patients underwent lung biopsy or lung transplantation. The outcomes were variable and could be as severe as chronic respiratory failure. Conclusion: The wide spectrum of respiratory manifestations and abnormal chest imaging in our study highlights the importance of evaluation for lung disease in patients with variants in FLNA. FLNA sequencing in suspected cases with ILD may obviate the need for a lung biopsy, prompt surveillance for progressive lung disease, and evaluation for associated clinical features.

Significantly high-expressed circFLNA has been found in various cancer cell lines, but not in lung cancer. Therefore, this study aimed to explore the role of circFLNA in the progression of lung cancer. The target gene of circFLNA was determined by bioinformatics and luciferase reporter assay. Viability, proliferation, migration, and invasion of the transfected cells were detected by CCK-8, colony formation, wound-healing, and transwell assays, respectively. A mouse subcutaneous xenotransplanted tumor model was established, and the expressions of circFLNA, miR-486-3p, XRCC1, CYP1A1, and related genes in the cancer cells and tissues were detected by RT-qPCR, Western blot, or immunohistochemistry. The current study found that miR-486-3p was low-expressed in lung cancer. MiR-486-3p, which has been found to target XRCC1 and CYP1A1, was regulated by circFLNA. CircFLNA was located in the cytoplasm and had a high expression in lung cancer cells. Cancer cell viability, proliferation, migration, and invasion were promoted by overexpressed circFLNA, XRCC1, and CYP1A1 but inhibited by miR-486-3p mimic and circFLNA knockdown. The weight of the xenotransplanted tumor was increased by circFLNA overexpression yet reduced by miR-486-3p mimic. Furthermore, miR-486-3p mimic reversed the effect of circFLNA overexpression on promoting lung cancer cells and tumors and regulating the expressions of miR-486-3p, XRCC1, CYP1A1, and metastasis/apoptosis/proliferation-related factors. However, overexpressed XRCC1 and CYP1A1 reversed the inhibitory effect of miR-486-3p mimic on cancer cells and tumors. In conclusion, circFLNA acted as a sponge of miR-486-3p to promote the proliferation, migration, and invasion of lung cancer cells in vitro and in vivo by regulating XRCC1 and CYP1A1.

Pancreatic ductal adenocarcinoma (PDAC) has a dismal prognosis largely owing to its inefficient diagnosis, rapid progress, and tenacious drug resistance. Here, we aimed to analyze the expressive patterns of proteins and phosphorylation in PDAC tissue samples and compare them to normal pancreatic tissue to investigate the underlying mechanisms and to reveal potential protein targets for diagnosis and drug development. Liquid chromatography coupled to mass spectrometry (LC-MS) based proteomics and phosphoproteomics analyses were performed using 20 pairs of patient-derived PDAC tissue and normal pancreatic tissue samples. Protein identification and quantification were conducted using MaxQuant software. Bioinformatics analysis was used to retrieve PDAC-relevant pathways and gene ontology (GO) terms. 4985 proteins and 3643 phosphoproteins were identified with high confidence; of these, 322 proteins and 235 phosphoproteins were dysregulated in PDAC. Several pathways, including several extracellular matrix-related pathways, were found to be strongly associated with PDAC. Further, the expression levels of filamin A (FLNA), integrin alpha-V (ITGAV), thymidine phosphorylase (TYMP), medium-chain specific acyl-CoA dehydrogenase, mitochondrial (ACADM), short-chain specific acyl-CoA dehydrogenase, mitochondrial (ACADS), and acetyl-CoA acetyltransferase, mitochondrial (ACAT1) were examined through western blot and immunohistochemistry analysis, and the results confirmed the validity of the proteomics analysis results. These findings provide comprehensive insight into the dysregulated regulative networks in PDAC tissue samples at the protein and phosphorylation levels, and they provide clues for further pathological studies and drug-target development.

Head and neck squamous cell carcinoma (HNSCC) is a malignant tumor of the upper aerodigestive tract. The loss and gain of miRNA function promote cancer development through various mechanisms. RNA sequencing (RNA-seq) and miRNAs sequencing data from the Cancer Genome Atlas (TCGA) was used to show the dysfunctional miRNAs microenvironment and to provide useful biomarkers for miRNAs therapy. Seven miRNAs were found to be independent prognostic factors of HNSCC patients in the training cohort. A total of 60 target genes for these miRNAs were predicted. Nine target genes (CDCA4, CXCL14, FLNC, KLF7, NBEAL2, P4HA1, PFKM, PFN2 and SEPPINE1) were correlated with patient's overall survival (OS) outcomes. We identified novel miRNAs markers for the prognosis of head and neck squamous cell carcinoma.

We report a case of a 38-year-old female with an

Multivesicular bodies (MVBs) contain intralumenal vesicles that are delivered to lysosomes for degradation or released extracellularly for intercellular signaling. Here, we identified Caenorhabditis elegans filamin FLN-2 as a novel regulator of MVB biogenesis. FLN-2 co-localizes with V-ATPase subunits on MVBs, and the loss of FLN-2 affects MVB biogenesis, reducing the number of MVBs in C. elegans hypodermis. FLN-2 associates with actin filaments and is required for F-actin organization. Like fln-2(lf) mutation, inactivation of the V0 or V1 sector of V-ATPase or inhibition of actin polymerization impairs MVB biogenesis. Super-resolution imaging shows that FLN-2 docks V-ATPase-decorated MVBs onto actin filaments. FLN-2 interacts via its calponin-homology domains with F-actin and the V1-E subunit, VHA-8. Our data suggest that FLN-2 mediates the docking of MVBs on the actin cytoskeleton, which is required for MVB biogenesis.

The process of GPCR dimerization can have profound effects on GPCR activation, signaling, and intracellular trafficking. Somatostatin receptors (SSTs) are class A GPCRs abundantly expressed in pituitary tumors where they represent the main pharmacological targets of somatostatin analogs (SSAs), thanks to their antisecretory and antiproliferative actions. The cytoskeletal protein filamin A (FLNA) directly interacts with both somatostatin receptor type 2 (SST

Background FLNC is one of the few genes associated with all types of cardiomyopathies, but it also underlies neuromuscular phenotype. The combination of concomitant neuromuscular and cardiac involvement is not often observed in filaminopathies and the impact of this on the disease prognosis has hitherto not been analyzed. Results Here we provide a detailed clinical, genetic, and structural prediction analysis of distinct FLNC-associated phenotypes based on twelve pediatric cases. They include early-onset restrictive cardiomyopathy (RCM) in association with congenital myopathy. In all patients the initial diagnosis was established during the first year of life and in five out of twelve (41.7%) patients the first symptoms were observed at birth. RCM was present in all patients, often in combination with septal defects. No ventricular arrhythmias were noted in any of the patients presented here. Myopathy was confirmed by neurological examination, electromyography, and morphological studies. Arthrogryposes was diagnosed in six patients and remained clinically meaningful with increasing age in three of them. One patient underwent successful heart transplantation at the age of 18 years and two patients are currently included in the waiting list for heart transplantation. Two died due to congestive heart failure. One patient had ICD instally as primary prevention of SCD. In ten out of twelve patients the disease was associated with missense variants and only in two cases loss of function variants were detected. In half of the described cases, an amino acid substitution A1186V, altering the structure of IgFLNc10, was found. Conclusions The present description of twelve cases of early-onset restrictive cardiomyopathy with congenital myopathy and FLNC mutation, underlines a distinct unique phenotype that can be suggested as a separate clinical form of filaminopathies. Amino acid substitution A1186V, which was observed in half of the cases, defines a mutational hotspot for the reported combination of myopathy and cardiomyopathy. Several independent molecular mechanisms of FLNC mutations linked to filamin structure and function can explain the broad spectrum of FLNC-associated phenotypes. Early disease presentation and unfavorable prognosis of heart failure demanding heart transplantation make awareness of this clinical form of filaminopathy of great clinical importance.

Relato de Caso Apresentamos o relato de caso de um paciente masculino de 50 anos de idade que procurou o departamento de pronto-socorro de nossa instituicao com queixa de dispneia progressiva ha 1 semana. O paciente relatava historia previa de fibromialgia e negava demais comorbidades, como infarto miocardico ou acidente vascular cerebral. Negava tabagismo atual e fazia uso eventual de alcool. Como medicamentos de uso continuo, utilizava duloxetina, pregabalina e zolpidem. Mae e primos com historia de insuficiencia cardiaca por [...]

Actomyosin networks are organized in space, direction, size, and connectivity to produce coordinated contractions across cells. We use the C. elegans spermatheca, a tube composed of contractile myoepithelial cells, to study how actomyosin structures are organized. FLN-1/filamin is required for the formation and stabilization of a regular array of parallel, contractile, actomyosin fibers in this tissue. Loss of fln-1 results in the detachment of actin fibers from the basal surface, which then accumulate along the cell junctions and are stabilized by spectrin. In addition, actin and myosin are captured at the nucleus by the linker of nucleoskeleton and cytoskeleton complex (LINC) complex, where they form large foci. Nuclear positioning and morphology, distribution of the endoplasmic reticulum and the mitochondrial network are also disrupted. These results demonstrate that filamin is required to prevent large actin bundle formation and detachment, to prevent excess nuclear localization of actin and myosin, and to ensure correct positioning of organelles.

Background Filamin C is a cytoskeletal protein expressed in cardiac cells. Nonsense variations in the filamin C gene (FLNC) were associated with dilated and arrhythmogenic cardiomyopathies. Methods and results We identified an intronic variation in FLNC gene (c.3791-1G > C) in three unrelated Ashkenazi Jewish families with variable expression of arrhythmia and cardiomyopathy. cDNA was prepared from a mutation carrier's cultured skin fibroblasts. Quantitative PCR demonstrated a reduction in total FLNC transcript, and no other FLNC splice variants were found. Single-nucleotide polymorphism (SNP) analysis revealed heterozygous variations in the genomic DNA that were not expressed in the messenger RNA. Immunohistochemical analysis of cardiac sections detected a normal distribution of filamin C protein in the heart ventricles. Conclusion The transcript that included the FLNC variant was degraded. Haploinsufficiency in filamin C underlies arrhythmogenic cardiomyopathy with variable symptoms.

BackgroundAdolescent idiopathic scoliosis (AIS) is a genetically heterogeneous disease characterised by three-dimensional deformity of the spine in the absence of a congenital spinal anomaly or neurological musculoskeletal disorder. The clinical variability and incomplete penetrance of some genes linked with AIS indicate that this disease constitutes an oligogenic trait.ObjectiveWe aimed to explore the oligogenic nature of this disease and identify novel AIS genes.MethodsWe analysed rare damaging variants within AIS-associated genes by using exome sequencing in 40 AIS trios and 183 sporadic patients.ResultsMultiple variants within AIS-associated genes were identified in eight AIS trios, and five individuals harboured rare damaging variants in theFLNBgene. The patients showed more frequent oligogenicity than the controls. In the gene-based burden test, the top signal resided inFLNB. In functional studies, we found that the AIS-associatedFLNBvariants altered the protein’s conformation and subcellular localisation and its interaction with other proteins (TTC26 and OFD1) involved in AIS. The most compelling evidence of an oligogenic basis was that the number of rare damaging variants was recognised as an independent prognostic factor for curve progression in Cox regression analysis.ConclusionOur data indicate that AIS is an oligogenic disease and identifyFLNBas a susceptibility gene for AIS.

The discovery of antibodies against Coxiella burnetii in cattery‐confined breeding cats indicating prior or current exposure (Shapiro et al. , 2015) prompted an investigation into possible sources of infection. One hypothesis was that raw meat diets containing reservoir species may provide a source of C. burnetii transmission. The aim of this pilot study was to determine whether C. burnetii DNA was present in raw meat sold exclusively for companion animal consumption. The sample population consisted of raw meat packages (n = 58) of primarily kangaroo origin, with three to four aliquots (50–120 mg) randomly selected from each package. Genomic DNA was extracted from whole tissue in each of these aliquots using a modified protocol. Three quantitative PCR assays were used for the detection of C. burnetii targeting the IS1111 gene, the heat shock operon htpAB and the C. burnetii outer membrane protein‐coding gene, com1 . Coxiella burnetii DNA was detected in 25/58 samples (43%) using the IS1111 , htpAB and/or com1 PCR assays and confirmed by DNA sequencing. All samples amplifying a product in the com1 assay also amplified a product in the htpAB and IS1111 assays. A total of 17/58 (29%) packets were positive with all three genes, 4/58 (7%) were positive with two genes (IS1111 and htpAB ) and 4/58 (7%) were positive with the IS1111 gene only. Coxiella burnetii DNA was five times more likely to be found in offal than skeletal muscle meat samples. All meat samples in which C. burnetii DNA was found were from kangaroo tissues, while samples labelled as non‐kangaroo meat (n = 4) were negative. Multi‐locus variable number of tandem repeat analysis (MLVA) identified three different genotypes of C. burnetii that have all been identified previously from Australian human clinical Q fever cases. Further investigations are required to determine the potential role of certain raw meats in the transmission of C. burnetii to cats and humans.

Patients with metastatic melanoma have a poorer prognosis. Prion protein (PrP) in melanoma is known to play an important role in cancer cell migration and invasion by interacting with filamin A (FLNa), a cytolinker protein. To investigate if PrP may contribute to cancer cell mobility independent of its binding to FLNa, we knocked out PRNP in M2 melanoma cell, which lacked FLNa expression. We found that deletion of PRNP in M2 significantly reduced its motility. When PRNP was deleted, the level of Akt was decreased. As a consequence, phosphorylation of small heat shock protein (hsp27) was also reduced, which resulted in polymerization of F-actin rendering the cells less migratory. Accordingly, when PrP was re-expressed in PRNP null M2 cells, the mobility of the recurred cells was rescued, so were the expression levels of Akt and phosphorylated hsp27, resulting in a decrease in the polymerization of F-actin. These results revealed that PrP can play a FLNa independent role in cytoskeletal organization and tumor cell migration by modulating Akt-hsp27-F-actin axis.

Filamin A is the most widely expressed isoform of filamin in mammalian tissues. It can be hydrolyzed by Calpain, producing a 90-kDa carboxyl-terminal fragment (ABP90). Calpeptin is a chemical inhibitor of Calpain, which can inhibit this effect. It has been shown that ABP90 acts as a transcription factor which is involved in mediating cell signaling. However, the significance of ABP90 and its clinical signature with underlying mechanisms have not been well studied in glioblastoma multiforme (GBM). ABP90 protein was measured in 36 glioma patients by Western blot. Human GBM cell lines U87 and A172 were used to clarify the precise role of ABP90. CCK-8 assay was used to analyze the cell viability. Transwell invasion assay and wound healing assay were used to analyze the migration and invasion. Expression of matrix metalloproteinase 2/tissue inhibitors of metalloproteinase 2 (MMP2/TIMP2) protein was analyzed by Western blot. ABP90 protein expression was lower in GBM tissues. The patients with low ABP90 protein expression had a shorter OS time (p = 0.046). After being treated with Calpain, the expression of ABP90 was upregulated, which led to a decline of cell viability, enhanced the efficacy of temozolomide and restrained the cell invasion. Calpeptin could inhibit the effect. The mechanism might be involved in the balance of MMP2/TIMP2. Our present data suggest that ABP90 expression is a significant prognostic factor and may play an important role in cell viability, chemotherapeutic sensitivity and invasion of GBM.

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a serious, debilitating disorder with a wide spectrum of symptoms, including pain, depression, and neurocognitive deterioration. Over 17 million people around the world have ME/CFS, predominantly women with peak onset at 30–50 years. Given the wide spectrum of symptoms and unclear etiology, specific biomarkers for diagnosis and stratification of ME/CFS are lacking. Here we show that actin network proteins in circulating extracellular vesicles (EVs) offer specific non-invasive biomarkers for ME/CFS. We found that circulating EVs were significantly increased in ME/CFS patients correlating to C-reactive protein, as well as biological antioxidant potential. Area under the receiver operating characteristic curve for circulating EVs was 0.80, allowing correct diagnosis in 90–94% of ME/CFS cases. From two independent proteomic analyses using circulating EVs from ME/CFS, healthy controls, idiopathic chronic fatigue, and depression, proteins identified from ME/CFS patients are involved in focal adhesion, actin skeletal regulation, PI3K-Akt signaling pathway, and Epstein-Barr virus infection. In particular, talin-1, filamin-A, and 14-3-3 family proteins were the most abundant proteins, representing highly specific ME/CFS biomarkers. Our results identified circulating EV number and EV-specific proteins as novel biomarkers for diagnosing ME/CFS, providing important information on the pathogenic mechanisms of ME/CFS.

Extracellular matrices (ECMs) in the intervertebral disc (IVD), lung and artery are thought to undergo age-dependant accumulation of damage by chronic exposure to mechanisms such as reactive oxygen species, proteases and glycation. It is unknown whether this damage accumulation is species-dependant (via differing lifespans and hence cumulative exposures) or whether it can influence the progression of age-related diseases such as atherosclerosis. Peptide location fingerprinting (PLF) is a new proteomic analysis method, capable of the non-targeted identification of structure-associated changes within proteins. Here we applied PLF to publicly available ageing human IVD (outer annulus fibrosus), ageing mouse lung and human arterial atherosclerosis datasets and bioinformatically identified novel target proteins alongside common age-associated differences within protein structures which were conserved between three ECM-rich organs, two species, three IVD tissue regions, sexes and in an age-related disease. We identify peptide yield differences across protein structures which coincide with biological regions, potentially reflecting the functional consequences of ageing or atherosclerosis for macromolecular assemblies (collagen VI), enzyme/inhibitor activity (alpha-2 macroglobulin), activation states (complement C3) and interaction states (laminins, perlecan, fibronectin, filamin-A, collagen XIV and apolipoprotein-B). Furthermore, we show that alpha-2 macroglobulin and collagen XIV exhibit possible shared structural consequences in IVD ageing and arterial atherosclerosis, providing novel links between an age-related disease and intrinsic ageing. Crucially, we also demonstrate that fibronectin, laminin beta chains and filamin-A all exhibit conserved age-associated structural differences between mouse lung and human IVD, providing evidence that ECM, and their associating proteins, may be subjected to potentially similar mechanisms or consequences of ageing across both species, irrespective of differences in lifespan and tissue function. © 2022 The Author(s) Funding agencies: Manchester Institute for Collaborative Research on Ageing (MICRA); Wal-greens Boots Alliance (WBA) (MJS), UK; the British Heart Foundation Centre ofResearch Excellence (BHF CRE); WellcomeTrust grant (108413/A/15/D); RGC European Union - Hong Kong Research and Innovation Cooperation Co-funding Mechanism (E-HKU703/18); European Union’s Horizon 2020research and innovation program (iPSpine; grant agreement number 825925); Shenz-hen “Key Medical Discipline Construction Fund”(SZXK077); UK Engineering and PhysicalSciences Research Council (EPSRC; EP/V011065/1); Swedish Heart Lung Foundation; Stroke Founda-tion; Olle Engkvist Foundation; Swedish GamlaTjänarinnor Foundation; Linköping University Hospital Research Fund.

With the number of cancer cases projected to significantly increase over time, researchers are currently exploring "nontraditional" research fields in the pursuit of novel therapeutics. One emerging area that is steadily gathering interest revolves around cellular mechanical machinery. When looking broadly at the physical properties of cancer, it has been debated whether a cancer could be defined as either stiffer or softer across cancer types. With numerous articles supporting both sides, the evidence instead suggests that cancer is not particularly regimented. Instead, cancer is highly adaptable, allowing it to endure the constantly changing microenvironments cancer cells encounter, such as tumor compression and the shear forces in the vascular system and body. What allows cancer cells to achieve this adaptability are the particular proteins that make up the mechanical network, leading to a particular mechanical program of the cancer cell. Coincidentally, some of these proteins, such as myosin II, α-actinins, filamins, and actin, have either altered expression in cancer and/or some type of direct involvement in cancer progression. For this reason, targeting the mechanical system as a therapeutic strategy may lead to more efficacious treatments in the future. However, targeting the mechanical program is far from trivial. As involved as the mechanical program is in cancer development and metastasis, it also helps drive many other key cellular processes, such as cell division, cell adhesion, metabolism, and motility. Therefore, anti-cancer treatments targeting the mechanical program must take great care to avoid potential side effects. Here, we introduce the potential of targeting the mechanical program while also providing its challenges and shortcomings as a strategy for cancer treatment.

Filamin C (FLN c) is a novel allergen in shellfish. In this study, FLN c from

In epithelia, normal cells recognize and extrude out newly emerged transformed cells by competition. This process is the most fundamental epithelial defence against cancer, whose occasional failure promotes oncogenesis. However, little is known about what factors determine the success or failure of this defence. Here we report that mechanical stiffening of extracellular matrix attenuates the epithelial defence against HRasV12-transformed cells. Using photoconversion labelling, protein tracking, and loss-of-function mutations, we attribute this attenuation to stiffening-induced perinuclear sequestration of a cytoskeletal protein, filamin. On soft matrix mimicking healthy epithelium, filamin exists as a dynamically single population, which moves to the normal cell-transformed cell interface to initiate the extrusion of transformed cells. However, on stiff matrix mimicking fibrotic epithelium, filamin redistributes into two dynamically distinct populations, including a new perinuclear pool that cannot move to the cell-cell interface. A matrix stiffness-dependent differential between filamin-Cdc42 and filamin-perinuclear cytoskeleton interaction controls this distinctive filamin localization and hence, determines the success or failure of epithelial defence on soft versus stiff matrix. Together, our study reveals how pathological matrix stiffening leads to a failed epithelial defence at the initial stage of oncogenesis., Epithelial cells have the ability to competitively remove potentially cancerous cells from the tissue. Here the authors discover that pathological stiffening of extracellular matrix leads to the loss of this basic epithelial defence against cancer.