α-catenin phosphorylation is actomyosin-sensitive and required for epithelial barrier functions through Afadin.

Zonula adherens junctions (zAJ) are spatially proximal to tight junctions (TJ), in a superstructure known as the apical junctional complex (AJC). A key component of the AJC is a circumferential ring of filamentous (F)-actin, but how actomyosin contractility drives AJC structure and epithelial barrier function is incompletely understood. Here, we show that a central mechanosensitive component of zAJ, α-catenin (α-cat), undergoes force-dependent phosphorylation in an unstructured linker region.

Profibrotic monocyte-derived alveolar macrophages as a biomarker and therapeutic target in systemic sclerosis-associated interstitial lung disease.

Interstitial lung disease (ILD) is present in over 60% of patients with systemic sclerosis (SSc) and is the leading cause of SSc-related deaths. Profibrotic monocyte-derived alveolar macrophages (MoAM) play a causal role in the pathogenesis of pulmonary fibrosis in animal models where their persistence in the niche requires signaling through Colony Stimulating Factor 1 Receptor (CSF1R). We hypothesized that the presence and proportion of MoAM in bronchoalveolar lavage (BAL) fluid from patients with SSc-ILD may be a biomarker of ILD severity.

α-Catenin force-sensitive binding and sequestration of LZTS2 leads to cytokinesis failure.

Epithelial cells can become polyploid upon tissue injury, but mechanosensitive cues that trigger this state are poorly understood. Using an Madin Darby Canine Kidney (MDCK) cell knock-out/reconstitution system, we show that α-catenin mutants that alter force-sensitive binding to F-actin or middle (M)-domain promote cytokinesis failure and binucleation, particularly near epithelial wound-fronts. We identified Leucine Zipper Tumor Suppressor 2 (LZTS2), a factor previously implicated in abscission, as a conformation sensitive proximity partner of α-catenin.

α-catenin phosphorylation is elevated during mitosis to resist apical rounding and epithelial barrier leak.

Epithelial cell cohesion and barrier function critically depend on α-catenin, an actin-binding protein and essential constituent of cadherin-catenin-based adherens junctions. α-catenin undergoes actomyosin force-dependent unfolding of both actin-binding and middle domains to strongly engage actin filaments and its various effectors; this mechanosensitivity is critical for adherens junction function. We previously showed that α-catenin is highly phosphorylated in an unstructured region that links the mechanosensitive middle and actin-binding domains (known as the P-linker region), but the cellular processes that promote α-catenin phosphorylation have remained elusive.

α-catenin middle- and actin-binding domain unfolding mutants differentially impact epithelial strength and sheet migration.

α-catenin (α-cat) displays force-dependent unfolding and binding to actin filaments through direct and indirect means, but features of adherens junction structure and function most vulnerable to loss of these allosteric mechanisms have not been directly compared. By reconstituting an α-cat F-actin-binding domain unfolding mutant known to exhibit enhanced binding to actin (α-cat-H0-FABD) into α-cat knockout Madin Darby Canine Kidney (MDCK) cells, we show that partial loss of the α-cat catch bond mechanism (via an altered H0 α-helix) leads to stronger epithelial sheet integrity with greater colocalization between the α-cat-H0-FABD mutant and actin. α-cat-H0-FABD -expressing cells are less efficient at closing scratch-wounds, suggesting reduced capacity for more dynamic cell-cell coordination.

α-catenin mechanosensitivity as a route to cytokinesis failure through sequestration of LZTS2.

Epithelial cells can become polyploid upon tissue injury, but mechanosensitive cues that trigger this state are poorly understood. Using α-catenin (α-cat) knock-out Madin Darby Canine Kidney (MDCK) cells reconstituted with wild-type and mutant forms of α-cat as a model system, we find that an established α-cat actin-binding domain unfolding mutant designed to reduce force-sensitive binding to F-actin (α-cat-H0-FABD) can promote cytokinesis failure, particularly along epithelial wound-fronts. Enhanced α-cat coupling to cortical actin is neither sufficient nor mitotic cell-autonomous for cytokinesis failure, but critically requires the mechanosensitive Middle-domain (M1-M2-M3) and neighboring cells.

Mitochondrial integrated stress response controls lung epithelial cell fate.

Alveolar epithelial type 1 (AT1) cells are necessary to transfer oxygen and carbon dioxide between the blood and air. Alveolar epithelial type 2 (AT2) cells serve as a partially committed stem cell population, producing AT1 cells during postnatal alveolar development and repair after influenza A and SARS-CoV-2 pneumonia. Little is known about the metabolic regulation of the fate of lung epithelial cells.

Hypercapnia alters stroma-derived Wnt production to limit β-catenin signaling and proliferation in AT2 cells.

Persistent symptoms and radiographic abnormalities suggestive of failed lung repair are among the most common symptoms in patients with COVID-19 after hospital discharge. In mechanically ventilated patients with acute respiratory distress syndrome (ARDS) secondary to SARS-CoV-2 pneumonia, low tidal volumes to reduce ventilator-induced lung injury necessarily elevate blood CO2 levels, often leading to hypercapnia. The role of hypercapnia on lung repair after injury is not completely understood.

Alpha-T-catenin is expressed in peripheral nerves as a constituent of Schwann cell adherens junctions.

The adherens junction component, alpha-T-catenin (αTcat) is an established contributor to cardiomyocyte junction structure and function, but recent genomic studies link CTNNA3 polymorphisms to diseases with no clear cardiac underpinning, including asthma, autism and multiple sclerosis, suggesting causal contributions from a different cell-type. We show Ctnna3 mRNA is highly expressed in peripheral nerves (e.g.

A Novel MIP-1-Expressing Macrophage Subtype in BAL Fluid from Healthy Volunteers.

Tissue availability remains an important limitation of single-cell genomic technologies for investigating cellular heterogeneity in human health and disease. BAL represents a minimally invasive approach to assessing an individual's lung cellular environment for diagnosis and research. However, the lack of high-quality, healthy lung reference data is a major obstacle to using single-cell approaches to study a plethora of lung diseases.

Lung Injury Induces Alveolar Type 2 Cell Hypertrophy and Polyploidy with Implications for Repair and Regeneration.

Epithelial polyploidization after injury is a conserved phenomenon recently shown to improve barrier restoration during wound healing. Whether lung injury can induce alveolar epithelial polyploidy is not known. We show that bleomycin injury induces alveolar type 2 cell (AT2) hypertrophy and polyploidy.

A spatially restricted fibrotic niche in pulmonary fibrosis is sustained by M-CSF/M-CSFR signalling in monocyte-derived alveolar macrophages.

Ontologically distinct populations of macrophages differentially contribute to organ fibrosis through unknown mechanisms.We applied lineage tracing, single-cell RNA sequencing and single-molecule fluorescence hybridisation to a spatially restricted model of asbestos-induced pulmonary fibrosis.We demonstrate that tissue-resident alveolar macrophages, tissue-resident peribronchial and perivascular interstitial macrophages, and monocyte-derived alveolar macrophages are present in the fibrotic niche.

Single-Cell Transcriptomic Analysis of Human Lung Provides Insights into the Pathobiology of Pulmonary Fibrosis.

The contributions of diverse cell populations in the human lung to pulmonary fibrosis pathogenesis are poorly understood. Single-cell RNA sequencing can reveal changes within individual cell populations during pulmonary fibrosis that are important for disease pathogenesis. To determine whether single-cell RNA sequencing can reveal disease-related heterogeneity within alveolar macrophages, epithelial cells, or other cell types in lung tissue from subjects with pulmonary fibrosis compared with control subjects.

Force-dependent allostery of the α-catenin actin-binding domain controls adherens junction dynamics and functions.

α-catenin is a key mechanosensor that forms force-dependent interactions with F-actin, thereby coupling the cadherin-catenin complex to the actin cytoskeleton at adherens junctions (AJs). However, the molecular mechanisms by which α-catenin engages F-actin under tension remained elusive. Here we show that the α1-helix of the α-catenin actin-binding domain (αcat-ABD) is a mechanosensing motif that regulates tension-dependent F-actin binding and bundling.

αT-catenin: A developmentally dispensable, disease-linked member of the α-catenin family.

α-Catenins are actin-filament binding proteins and critical subunits of the cadherin-catenin cell-cell adhesive complex. They are found in nominally-defined epithelial (E), neural (N), and testis (T) forms transcribed from three distinct genes. While most of α-catenin research has focused on the developmentally essential founding member, αE-catenin, this review discusses recent studies on αT-catenin (CTNNA3), a developmentally dispensable isoform that is emerging as relevant to cardiac, allergic and neurological diseases.

α-Catenin homodimers are recruited to phosphoinositide-activated membranes to promote adhesion.

A unique feature of α-catenin localized outside the cadherin-catenin complex is its capacity to form homodimers, but the subcellular localization and functions of this form of α-catenin remain incompletely understood. We identified a cadherin-free form of α-catenin that is recruited to the leading edge of migrating cells in a phosphatidylinositol 3-kinase-dependent manner. Surface plasmon resonance analysis shows that α-catenin homodimers, but not monomers, selectively bind phosphatidylinositol-3,4,5-trisphosphate-containing lipid vesicles with high affinity, where three basic residues, K488, K493, and R496, contribute to binding.

Lrp5/β-Catenin Signaling Controls Lung Macrophage Differentiation and Inhibits Resolution of Fibrosis.

Previous studies established that attenuating Wnt/β-catenin signaling limits lung fibrosis in the bleomycin mouse model of this disease, but the contribution of this pathway to distinct lung cell phenotypes relevant to tissue repair and fibrosis remains incompletely understood. Using microarray analysis, we found that bleomycin-injured lungs from mice that lack the Wnt coreceptor low density lipoprotein receptor-related protein 5 (Lrp5) and exhibit reduced fibrosis showed enrichment for pathways related to extracellular matrix processing, immunity, and lymphocyte proliferation, suggesting the contribution of an immune-matrix remodeling axis relevant to fibrosis. Activation of β-catenin signaling was seen in lung macrophages using the β-catenin reporter mouse, Axin2.

A Simple Method to Assess Abundance of the β-Catenin Signaling Pool in Cells.

β-catenin (CTNNB1) is a dual-function cell-cell adhesion/transcriptional co-activator protein and an essential transducer of canonical Wnt signals. Although a number of established techniques and reagents are available to quantify the nuclear signaling activity of β-catenin (e.g.

Adiponectin inhibits Wnt co-receptor, Lrp6, phosphorylation and β-catenin signaling.

Adiponectin is a pleiotropic adipokine implicated in obesity, metabolic syndrome and cardiovascular disease. Recent studies have identified adiponectin as a negative regulator of tissue fibrosis. Wnt/β-catenin signaling has also been implicated in metabolic syndrome and can promote tissue fibrosis, but the extent to which adiponectin cross-regulates Wnt/β-catenin signaling is unknown.

α-Catenin phosphorylation promotes intercellular adhesion through a dual-kinase mechanism.

The cadherin-catenin adhesion complex is a key contributor to epithelial tissue stability and dynamic cell movements during development and tissue renewal. How this complex is regulated to accomplish these functions is not fully understood. We identified several phosphorylation sites in mammalian αE-catenin (also known as catenin α-1) and Drosophila α-Catenin within a flexible linker located between the middle (M)-region and the carboxy-terminal actin-binding domain.

Wnt coreceptor Lrp5 is a driver of idiopathic pulmonary fibrosis.

Rationale: Wnt/β-catenin signaling has been implicated in lung fibrosis, but how this occurs and whether expression changes in Wnt pathway components predict disease progression is unknown.

Objectives: To determine whether the Wnt coreceptor Lrp5 drives pulmonary fibrosis in mice and is predictive of disease severity in humans.

Methods: We examined mice with impaired Wnt signaling caused by loss of the Wnt coreceptor Lrp5 in models of lung fibrosis induced by bleomycin or an adenovirus encoding an active form of transforming growth factor (TGF)-β.

α-Catenin is an inhibitor of transcription.

α-Catenin (α-cat) is an actin-binding protein required for cell-cell cohesion. Although this adhesive function for α-cat is well appreciated, cells contain a substantial amount of nonjunctional α-cat that may be used for other functions. We show that α-cat is a nuclear protein that can interact with β-catenin (β-cat) and T-cell factor (TCF) and that the nuclear accumulation of α-cat depends on β-cat.

Nuclear β-catenin is increased in systemic sclerosis pulmonary fibrosis and promotes lung fibroblast migration and proliferation.

Pulmonary fibrosis is a disease that results in loss of normal lung architecture, but the signaling events that drive tissue destruction are incompletely understood. Wnt/β-catenin signaling is important in normal lung development, but whether abnormal signaling occurs in lung fibrosis due to systemic sclerosis and the consequences of β-catenin signaling toward the fibrogenic phenotype remain poorly defined. In this study, we show nuclear β-catenin accumulation in fibroblastic foci from lungs of patients with systemic sclerosis-associated advanced pulmonary fibrosis.

Beta-catenin phosphorylated at serine 45 is spatially uncoupled from beta-catenin phosphorylated in the GSK3 domain: implications for signaling.

C. elegans and Drosophila generate distinct signaling and adhesive forms of beta-catenin at the level of gene expression. Whether vertebrates, which rely on a single beta-catenin gene, generate unique adhesive and signaling forms at the level of protein modification remains unresolved.