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Tissue Barriers Oct 2023Tight junctions (TJs) are the most apical components of junctional complexes in epithelial and endothelial cells. Barrier function is one of the major functions of TJ,... (Review)
Review
Tight junctions (TJs) are the most apical components of junctional complexes in epithelial and endothelial cells. Barrier function is one of the major functions of TJ, which restricts the ions and small water-soluble molecules from passing through the paracellular pathway. Adherens junctions (AJs) play an important role in cell-cell adhesion and cell signaling. Gap junctions (GJs) are intercellular channels regulating electrical and metabolic signals between cells. It is well known that TJ integral membrane proteins, such as claudins and occludins, are the molecular building blocks responsible for TJ barrier function. However, recent studies demonstrate that proteins of other junctional complexes can influence and regulate TJ barrier function. Therefore, the crosstalk between different cell junctions represents a common means to modulate cellular activities. In this review, we will discuss the interactions among TJ, AJ, and GJ by focusing on how AJ and GJ proteins regulate TJ barrier function in different biological systems.
Topics: Tight Junctions; Epithelial Cells; Endothelial Cells; Intercellular Junctions; Adherens Junctions
PubMed: 36220768
DOI: 10.1080/21688370.2022.2133880 -
The Journal of Cell Biology Jul 2023The mechanisms that regulate the spatial sorting of nonmuscle myosins-2 (NM2) isoforms and couple them mechanically to the plasma membrane are unclear. Here we show that...
The mechanisms that regulate the spatial sorting of nonmuscle myosins-2 (NM2) isoforms and couple them mechanically to the plasma membrane are unclear. Here we show that the cytoplasmic junctional proteins cingulin (CGN) and paracingulin (CGNL1) interact directly with NM2s through their C-terminal coiled-coil sequences. CGN binds strongly to NM2B, and CGNL1 to NM2A and NM2B. Knockout (KO), exogenous expression, and rescue experiments with WT and mutant proteins show that the NM2-binding region of CGN is required for the junctional accumulation of NM2B, ZO-1, ZO-3, and phalloidin-labeled actin filaments, and for the maintenance of tight junction membrane tortuosity and apical membrane stiffness. CGNL1 expression promotes the junctional accumulation of both NM2A and NM2B and its KO results in myosin-dependent fragmentation of adherens junction complexes. These results reveal a mechanism for the junctional localization of NM2A and NM2B and indicate that, by binding to NM2s, CGN and CGNL1 mechanically couple the actomyosin cytoskeleton to junctional protein complexes to mechanoregulate the plasma membrane.
Topics: Adherens Junctions; Cell Membrane; Cytoskeletal Proteins; Cytoskeleton; Myosins; Tight Junctions
PubMed: 37204781
DOI: 10.1083/jcb.202208065 -
Respiratory Research Nov 2023Idiopathic pulmonary fibrosis (IPF) is a heterogeneous disease that is pathologically characterized by areas of normal-appearing lung parenchyma, active fibrosis...
BACKGROUND
Idiopathic pulmonary fibrosis (IPF) is a heterogeneous disease that is pathologically characterized by areas of normal-appearing lung parenchyma, active fibrosis (transition zones including fibroblastic foci) and dense fibrosis. Defining transcriptional differences between these pathologically heterogeneous regions of the IPF lung is critical to understanding the distribution and extent of fibrotic lung disease and identifying potential therapeutic targets. Application of a spatial transcriptomics platform would provide more detailed spatial resolution of transcriptional signals compared to previous single cell or bulk RNA-Seq studies.
METHODS
We performed spatial transcriptomics using GeoMx Nanostring Digital Spatial Profiling on formalin-fixed paraffin-embedded (FFPE) tissue from 32 IPF and 12 control subjects and identified 231 regions of interest (ROIs). We compared normal-appearing lung parenchyma and airways between IPF and controls with histologically normal lung tissue, as well as histologically distinct regions within IPF (normal-appearing lung parenchyma, transition zones containing fibroblastic foci, areas of dense fibrosis, and honeycomb epithelium metaplasia).
RESULTS
We identified 254 differentially expressed genes (DEGs) between IPF and controls in histologically normal-appearing regions of lung parenchyma; pathway analysis identified disease processes such as EIF2 signaling (important for cap-dependent mRNA translation), epithelial adherens junction signaling, HIF1α signaling, and integrin signaling. Within IPF, we identified 173 DEGs between transition and normal-appearing lung parenchyma and 198 DEGs between dense fibrosis and normal lung parenchyma; pathways dysregulated in both transition and dense fibrotic areas include EIF2 signaling pathway activation (upstream of endoplasmic reticulum (ER) stress proteins ATF4 and CHOP) and wound healing signaling pathway deactivation. Through cell deconvolution of transcriptome data and immunofluorescence staining, we confirmed loss of alveolar parenchymal signals (AGER, SFTPB, SFTPC), gain of secretory cell markers (SCGB3A2, MUC5B) as well as dysregulation of the upstream regulator ATF4, in histologically normal-appearing tissue in IPF.
CONCLUSIONS
Our findings demonstrate that histologically normal-appearing regions from the IPF lung are transcriptionally distinct when compared to similar lung tissue from controls with histologically normal lung tissue, and that transition zones and areas of dense fibrosis within the IPF lung demonstrate activation of ER stress and deactivation of wound healing pathways.
Topics: Humans; Eukaryotic Initiation Factor-2; Idiopathic Pulmonary Fibrosis; Lung; Transcriptome; Fibrosis
PubMed: 37978501
DOI: 10.1186/s12931-023-02572-6 -
Cardiovascular Research Jun 2024SCUBE2 (Signal peptide-CUB-epidermal growth factor-like domain-containing protein 2) is a secreted or membrane-bound protein originally identified from endothelial cells...
AIMS
SCUBE2 (Signal peptide-CUB-epidermal growth factor-like domain-containing protein 2) is a secreted or membrane-bound protein originally identified from endothelial cells (ECs). Our previous work showed that SCUBE2 forms a complex with E-cadherin and stabilizes epithelial adherens junctions (AJs) to promote epithelial phenotypes. However, it remains unclear whether SCUBE2 also interacts with vascular endothelial (VE)-cadherin and modulates EC barrier function. In this study, we investigated whether and how SCUBE2 in ECs regulates vascular barrier maintenance.
METHODS AND RESULTS
We showed that SCUBE2 colocalized and interacted with VE-cadherin and VE-protein tyrosine phosphatase (VE-PTP) within EC AJs. Furthermore, SCUBE2 knockdown disrupted EC AJs and increased EC permeability. Expression of EC SCUBE2 was suppressed at both mRNA and protein levels via the nuclear factor-κB (NF-κB) signaling pathway in response to pro-inflammatory cytokines or permeability-inducing agents. In line with these findings, EC-specific deletion of Scube2 (EC-KO) in mice impaired baseline barrier function and worsened vascular leakiness of peripheral capillaries after local injection of histamine or vascular endothelial growth factor. EC-KO mice were also sensitive to pulmonary vascular hyperpermeability and leukocyte infiltration in response to acute endotoxin- or influenza virus-induced systemic inflammation. Meanwhile, EC-specific SCUBE2-overexpressing mice were protected from these effects. Molecular studies suggested that SCUBE2 acts as a scaffold molecule enabling VE-PTP to dephosphorylate VE-cadherin, which prevents VE-cadherin internalization and stabilizes EC AJs. As such, loss of SCUBE2 resulted in hyperphosphorylation of VE-cadherin at tyrosine 685, which led to its endocytosis, thus destabilizing EC AJs and reducing barrier function. All of these effects were exacerbated by inflammatory insults.
CONCLUSIONS
We found that SCUBE2 contributes to vascular integrity by recruiting VE-PTP to dephosphorylate VE-cadherin and stabilize AJs, thereby promoting EC barrier function. Moreover, our data suggest that genetic overexpression or pharmacological upregulation of SCUBE2 may help to prevent vascular leakage and edema in inflammatory diseases.
PubMed: 38870316
DOI: 10.1093/cvr/cvae132 -
Seminars in Immunopathology Jan 2024The lungs serve as the primary organ for respiration, facilitating the vital exchange of gases with the bloodstream. Given their perpetual exposure to external... (Review)
Review
The lungs serve as the primary organ for respiration, facilitating the vital exchange of gases with the bloodstream. Given their perpetual exposure to external particulates and pathogens, they possess intricate protective barriers. Cellular adhesion in the lungs is robustly maintained through tight junctions, adherens junctions, and desmosomes. Furthermore, the pulmonary system features a mucociliary clearance mechanism that synthesizes mucus and transports it to the outside. This mucus is enriched with chemical barriers like antimicrobial proteins and immunoglobulin A (IgA). Additionally, a complex immunological network comprising epithelial cells, neural cells, and immune cells plays a pivotal role in pulmonary defense. A comprehensive understanding of these protective systems offers valuable insights into potential pathologies and their therapeutic interventions.
Topics: Humans; Lung; Animals; Mucociliary Clearance; Respiratory Mucosa; Tight Junctions; Cell Adhesion; Mucus
PubMed: 38451292
DOI: 10.1007/s00281-024-01003-y -
Neurobiology of Disease Sep 2023Impairment of the blood-brain barrier (BBB) is considered to be a common feature among neurodegenerative diseases, including Alzheimer's, Parkinson's and prion diseases....
BACKGROUND
Impairment of the blood-brain barrier (BBB) is considered to be a common feature among neurodegenerative diseases, including Alzheimer's, Parkinson's and prion diseases. In prion disease, increased BBB permeability was reported 40 years ago, yet the mechanisms behind the loss of BBB integrity have never been explored. Recently, we showed that reactive astrocytes associated with prion diseases are neurotoxic. The current work examines the potential link between astrocyte reactivity and BBB breakdown.
RESULTS
In prion-infected mice, the loss of BBB integrity and aberrant localization of aquaporin 4 (AQP4), a sign of retraction of astrocytic endfeet from blood vessels, were noticeable prior to disease onset. Gaps in cell-to-cell junctions along blood vessels, together with downregulation of Occludin, Claudin-5 and VE-cadherin, which constitute tight and adherens junctions, suggested that loss of BBB integrity is linked with degeneration of vascular endothelial cells. In contrast to cells isolated from non-infected adult mice, endothelial cells originating from prion-infected mice displayed disease-associated changes, including lower levels of Occludin, Claudin-5 and VE-cadherin expression, impaired tight and adherens junctions, and reduced trans-endothelial electrical resistance (TEER). Endothelial cells isolated from non-infected mice, when co-cultured with reactive astrocytes isolated from prion-infected animals or treated with media conditioned by the reactive astrocytes, developed the disease-associated phenotype observed in the endothelial cells from prion-infected mice. Reactive astrocytes were found to produce high levels of secreted IL-6, and treatment of endothelial monolayers originating from non-infected animals with recombinant IL-6 alone reduced their TEER. Remarkably, treatment with extracellular vesicles produced by normal astrocytes partially reversed the disease phenotype of endothelial cells isolated from prion-infected animals.
CONCLUSIONS
To our knowledge, the current work is the first to illustrate early BBB breakdown in prion disease and to document that reactive astrocytes associated with prion disease are detrimental to BBB integrity. Moreover, our findings suggest that the harmful effects are linked to proinflammatory factors secreted by reactive astrocytes.
Topics: Animals; Mice; Blood-Brain Barrier; Astrocytes; Endothelial Cells; Claudin-5; Interleukin-6; Occludin; Prion Diseases; Prions
PubMed: 37597815
DOI: 10.1016/j.nbd.2023.106264 -
Current Biology : CB Jul 2023Collective cell movements contribute to tissue development and repair and spread metastatic disease. In epithelia, cohesive cell movements require reorganization of...
Collective cell movements contribute to tissue development and repair and spread metastatic disease. In epithelia, cohesive cell movements require reorganization of adherens junctions and the actomyosin cytoskeleton. However, the mechanisms that coordinate cell-cell adhesion and cytoskeletal remodeling during collective cell migration in vivo are unclear. We investigated the mechanisms of collective cell migration during epidermal wound healing in Drosophila embryos. Upon wounding, the cells adjacent to the wound internalize cell-cell adhesion molecules and polarize actin and the motor protein non-muscle myosin II to form a supracellular cable around the wound that coordinates cell movements. The cable anchors at former tricellular junctions (TCJs) along the wound edge, and TCJs are reinforced during wound closure. We found that the small GTPase Rap1 was necessary and sufficient for rapid wound repair. Rap1 promoted myosin polarization to the wound edge and E-cadherin accumulation at TCJs. Using embryos expressing a mutant form of the Rap1 effector Canoe/Afadin that cannot bind Rap1, we found that Rap1 signals through Canoe for adherens junction remodeling, but not for actomyosin cable assembly. Instead, Rap1 was necessary and sufficient for RhoA/Rho1 activation at the wound edge. The RhoGEF Ephexin localized to the wound edge in a Rap1-dependent manner, and Ephexin was necessary for myosin polarization and rapid wound repair, but not for E-cadherin redistribution. Together, our data show that Rap1 coordinates the molecular rearrangements that drive embryonic wound healing, promoting actomyosin cable assembly through Ephexin-Rho1, and E-cadherin redistribution through Canoe, thus enabling rapid collective cell migration in vivo.
Topics: Animals; Actomyosin; Cell Adhesion; Drosophila melanogaster; Drosophila Proteins; Cell Movement; Myosins; Adherens Junctions; Cadherins
PubMed: 37244252
DOI: 10.1016/j.cub.2023.05.009 -
International Journal of Molecular... Jul 2023The intercalated disk is a cardiac specific structure composed of three main protein complexes-adherens junctions, desmosomes, and gap junctions-that work in concert to... (Review)
Review
The intercalated disk is a cardiac specific structure composed of three main protein complexes-adherens junctions, desmosomes, and gap junctions-that work in concert to provide mechanical stability and electrical synchronization to the heart. Each substructure is regulated through a variety of mechanisms including proteolysis. Calpain proteases, a class of cysteine proteases dependent on calcium for activation, have recently emerged as important regulators of individual intercalated disk components. In this review, we will examine how calcium homeostasis regulates normal calpain function. We will also explore how calpains modulate gap junctions, desmosomes, and adherens junctions activity by targeting specific proteins, and describe the molecular mechanisms of how calpain dysregulation leads to structural and signaling defects within the heart. We will then examine how changes in calpain activity affects cardiomyocytes, and how such changes underlie various heart diseases.
Topics: Calpain; Calcium; Myocardium; Myocytes, Cardiac; Adherens Junctions
PubMed: 37511485
DOI: 10.3390/ijms241411726 -
Life Sciences Mar 2024Cingulin and its paralog paracingulin are vital components of the apical junctional complex in vertebrate epithelial and endothelial cells. They are both found in tight... (Review)
Review
Cingulin and its paralog paracingulin are vital components of the apical junctional complex in vertebrate epithelial and endothelial cells. They are both found in tight junctions (TJ), and paracingulin is also detectable in adherens junctions (AJ) as TJ cytoplasmic plaque proteins. Cingulin and paracingulin interact with other proteins to perform functions. They interact with cytoskeletal proteins, modulate the activity of small GTPases, such as RhoA and Rac1, and regulate gene expression. In addition, cingulin and paracingulin regulate barrier function and many pathological processes, including inflammation and tumorigenesis. In this review, we summarize the discovery and structure, expression and subcellular distribution, and molecular interactions of cingulin family proteins and discuss their role in development, physiology, and pathological processes.
Topics: Membrane Proteins; Endothelial Cells; Clinical Relevance; Cytoskeletal Proteins; Tight Junctions; Epithelial Cells
PubMed: 38354973
DOI: 10.1016/j.lfs.2024.122504 -
Molecular Medicine Reports Aug 2023Osteoporosis increases the risk of fracture. Improving the diagnosis and treatment of osteoporosis has clinical applications. The differentially expressed genes...
Osteoporosis increases the risk of fracture. Improving the diagnosis and treatment of osteoporosis has clinical applications. The differentially expressed genes (DEcircRs, DEmRs, DEmiRs) of osteoporotic patients and controls were analyzed using the GEO database, and enrichment analysis of DEmRs was performed. circRNAs and mRNAs, which were predicted to have a target relationship with DEmRs, were obtained to compare competing endogenous RNA (ceRNA) regulatory networks by comparison with differentially expressed genes. Molecular experiments were utilized to validate the expression of genes within the network. The interactions between genes within the ceRNA network were validated by luciferase reporter assays. Following overexpression of circ_0070304 in bone marrow mesenchymal stem cells (BMSCs), the osteogenic differentiation of the cells was assessed by Alizarin Red staining. A total of 110 intersectional DEmRs between patients with osteoporosis and controls from GSE35958 and GSE56815, which were mainly enriched in estrogen, the thyroid hormone signaling pathway, and adherens junctions were identified. A ceRNA network [circ_0070304/miR‑183‑5p/ring finger and CCCH‑type domains 2 (RC3H2)] was then constructed. Circ_0070304 acted as a sponge for miR‑183‑5p and regulated RC3H2 expression. Overexpression of circ_0070304 upregulated ROCK1 and induced osteogenic differentiation. The ceRNA regulatory network that was obtained is expected to be a new target for osteoporosis treatment and to provide new insights into the diagnosis and treatment of osteoporosis in greater depth.
Topics: Humans; MicroRNAs; Osteogenesis; Osteoporosis; RNA, Circular; RNA, Messenger; rho-Associated Kinases
PubMed: 37326104
DOI: 10.3892/mmr.2023.13033