scholarly journals A heterologous in-cell assay for investigating intermicrovillar adhesion complex interactions and function

2020 ◽  
Author(s):  
Meredith L. Weck ◽  
Scott W. Crawley ◽  
Matthew J. Tyska
Keyword(s):  
Epithelial Cells ◽  
Apical Surface ◽  
Tail Domain ◽  
Complex Interactions ◽  
Brush Borders ◽  
Specific Complex ◽  
Dynamic Length ◽  
And Function ◽  
Adhesion Complex ◽  
Dependent Mechanism

ABSTRACTSolute transporting epithelial cells build arrays of microvilli on their apical surface to increase membrane scaffolding capacity and enhance function potential. In epithelial tissues such as the kidney and gut, microvilli are length-matched and assembled into tightly packed ‘brush borders’, which are organized by ∼50 nm thread-like links that form between the distal tips of adjacent protrusions. Composed of protocadherins CDHR2 and CDHR5, adhesion links are stabilized at the tips by a cytoplasmic tripartite module containing the scaffolds USH1C and ANKS4B, and the actin-based motor, MYO7B. As several questions about the formation and function of this ‘intermicrovillar adhesion complex’ remain open, we devised a system that allows one to study individual binary interactions between specific complex components and MYO7B. Our approach employs a chimeric myosin consisting of the motor domain of MYO10 fused to the cargo-binding tail domain of MYO7B. When expressed in HeLa cells, which do not normally produce adhesion complex proteins, this motor exhibited robust trafficking to the tips of filopodia and was also able to transport individual components to these sites. Unexpectedly, the MYO10/MYO7B chimera was able to deliver CDHR2 and CDHR5 to distal tips in the absence USH1C or ANKS4B. Cells engineered to localize high levels of CDHR2 at filopodial tips acquired inter-filopodial adhesion and exhibited a striking dynamic length matching activity that aligned distal tips over time. These observations reveal a robust adhesion-dependent mechanism for matching the lengths of adjacent surface protrusions, and may offer insight on how epithelial cells minimize microvillar length variability.

scholarly journals A heterologous in-cell assay for investigating intermicrovillar adhesion complex interactions reveals a novel protrusion length-matching mechanism

2020 ◽  
Vol 295 (48) ◽  
pp. 16191-16206
Author(s):  
Meredith L. Weck ◽  
Scott W. Crawley ◽  
Matthew J. Tyska
Keyword(s):  
Epithelial Cells ◽  
Apical Surface ◽  
Tail Domain ◽  
Complex Interactions ◽  
Specific Complex ◽  
Matching Mechanism ◽  
Dynamic Length ◽  
And Function ◽  
Adhesion Complex ◽  
Complex Components

Solute transporting epithelial cells build arrays of microvilli on their apical surface to increase membrane scaffolding capacity and enhance function potential. In epithelial tissues such as the kidney and gut, microvilli are length-matched and assembled into tightly packed “brush borders,” which are organized by ∼50-nm thread-like links that form between the distal tips of adjacent protrusions. Composed of protocadherins CDHR2 and CDHR5, adhesion links are stabilized at the tips by a cytoplasmic tripartite module containing the scaffolds USH1C and ANKS4B and the actin-based motor MYO7B. Because several questions about the formation and function of this “intermicrovillar adhesion complex” remain open, we devised a system that allows one to study individual binary interactions between specific complex components and MYO7B. Our approach employs a chimeric myosin consisting of the MYO10 motor domain fused to the MYO7B cargo-binding tail domain. When expressed in HeLa cells, which do not normally produce adhesion complex proteins, this chimera trafficked to the tips of filopodia and was also able to transport individual complex components to these sites. Unexpectedly, the MYO10–MYO7B chimera was able to deliver CDHR2 and CDHR5 to distal tips in the absence of USH1C or ANKS4B. Cells engineered to localize high levels of CDHR2 at filopodial tips acquired interfilopodial adhesion and exhibited a striking dynamic length-matching activity that aligned distal tips over time. These findings deepen our understanding of mechanisms that promote the distal tip accumulation of intermicrovillar adhesion complex components and also offer insight on how epithelial cells minimize microvillar length variability.

scholarly journals The WD40 protein Morg1 facilitates Par6–aPKC binding to Crb3 for apical identity in epithelial cells

2013 ◽  
Vol 200 (5) ◽  
pp. 635-650 ◽  
Author(s):  
Junya Hayase ◽  
Sachiko Kamakura ◽  
Yuko Iwakiri ◽  
Yoshihiro Yamaguchi ◽  
Tomoko Izaki ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Epithelial Cells ◽  
Tight Junction ◽  
Apical Membrane ◽  
Transmembrane Protein ◽  
Cyst Formation ◽  
Cell Polarization ◽  
Small Gtpase ◽  
Apical Surface ◽  
And Function

Formation of apico-basal polarity in epithelial cells is crucial for both morphogenesis (e.g., cyst formation) and function (e.g., tight junction development). Atypical protein kinase C (aPKC), complexed with Par6, is considered to translocate to the apical membrane and function in epithelial cell polarization. However, the mechanism for translocation of the Par6–aPKC complex has remained largely unknown. Here, we show that the WD40 protein Morg1 (mitogen-activated protein kinase organizer 1) directly binds to Par6 and thus facilitates apical targeting of Par6–aPKC in Madin-Darby canine kidney epithelial cells. Morg1 also interacts with the apical transmembrane protein Crumbs3 to promote Par6–aPKC binding to Crumbs3, which is reinforced with the apically localized small GTPase Cdc42. Depletion of Morg1 disrupted both tight junction development in monolayer culture and cyst formation in three-dimensional culture; apico-basal polarity was notably restored by forced targeting of aPKC to the apical surface. Thus, Par6–aPKC recruitment to the premature apical membrane appears to be required for definition of apical identity of epithelial cells.

scholarly journals Bacterial Exposure Induces and Activates Matrilysin in Mucosal Epithelial Cells

2000 ◽  
Vol 148 (6) ◽  
pp. 1305-1315 ◽  
Author(s):  
Yolanda S. López-Boado ◽  
Carole L. Wilson ◽  
Lora V. Hooper ◽  
Jeffrey I. Gordon ◽  
Scott J. Hultgren ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Acute Infection ◽  
Human Colon ◽  
Cell Types ◽  
The Body ◽  
Apical Surface ◽  
Tracheal Epithelial Cells ◽  
Lung Carcinoma Cells ◽  
Tracheal Epithelial ◽  
And Function

Matrilysin, a matrix metalloproteinase, is expressed and secreted lumenally by intact mucosal and glandular epithelia throughout the body, suggesting that its regulation and function are shared among tissues. Because matrilysin is produced in Paneth cells of the murine small intestine, where it participates in innate host defense by activation of prodefensins, we speculated that its expression would be influenced by bacterial exposure. Indeed, acute infection (10–90 min) of human colon, bladder, and lung carcinoma cells, primary human tracheal epithelial cells, and human tracheal explants with type 1–piliated Escherichia coli mediated a marked (25–50-fold) and sustained (>24 h) induction of matrilysin production. In addition, bacterial infection resulted in activation of the zymogen form of the enzyme, which was selectively released at the apical surface. Induction of matrilysin was mediated by a soluble, non-LPS bacterial factor and correlated with the release of defensin-like bacteriocidal activity. Bacteria did not induce matrilysin in other cell types, and expression of other metalloproteinases by epithelial cells was not affected by bacteria. Matrilysin was not detected in germ-free mice, but the enzyme was induced after colonization with Bacteroides thetaiotaomicron. These findings indicate that bacterial exposure is a potent and physiologically relevant signal regulating matrilysin expression in epithelial cells.

scholarly journals An alternative mode of epithelial polarity in the Drosophila midgut

2018 ◽  
Author(s):  
Jia Chen ◽  
Aram-Christopher Sayadian ◽  
Nick Lowe ◽  
Holly E. Lovegrove ◽  
Daniel St Johnston
Keyword(s):  
Epithelial Cells ◽  
Adherens Junctions ◽  
Epithelial Polarity ◽  
Alternative Mode ◽  
Epithelial Tissues ◽  
Occluding Junctions ◽  
Discs Large ◽  
And Function ◽  
Adhesion Complex

AbstractApical-basal polarity is essential for the formation and function of epithelial tissues, whereas loss of polarity is a hallmark of tumours. Studies in Drosophila have identified conserved polarity factors that define the apical (Crumbs, Stardust, Par-6, aPKC), junctional (Baz/Par-3) and basolateral (Scribbled, Discs large, Lgl) domains of epithelial cells (1, 2). Because these conserved factors mark equivalent domains in diverse vertebrate and invertebrate epithelial types, it is generally assumed that this system organises polarity in all epithelia. Here we show that this is not the case, as none of these canonical factors are required for the polarisation of the endodermal epithelium of the Drosophila adult midgut. Furthermore, unlike other Drosophila epithelia, the midgut forms occluding junctions above adherens junctions, as in vertebrates, and requires the integrin adhesion complex for polarity (3, 4). Thus, Drosophila contains two types of epithelia that polarise by different mechanisms. Since knock-outs of canonical polarity factors often have little effect on the polarity of vertebrate epithelia, this diversity of polarity mechanisms is likely to be conserved in other animals (5-8).

Amplification loop of the inflammatory process is induced by P2X7R activation in intestinal epithelial cells in response to neutrophil transepithelial migration

2010 ◽  
Vol 299 (1) ◽  
pp. G32-G42 ◽  
Author(s):  
Annabelle Cesaro ◽  
Patrick Brest ◽  
Véronique Hofman ◽  
Xavier Hébuterne ◽  
Scott Wildman ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Intestinal Epithelial Cells ◽  
Active Phase ◽  
Apical Surface ◽  
Intestinal Epithelial ◽  
T84 Cells ◽  
Caspase 1 ◽  
Bowel Diseases ◽  
And Function

Inflammatory bowel diseases (IBD) are characterized during their active phase by polymorphonuclear leukocyte (PMNL) transepithelial migration. The efflux of PMNL into the mucosa is associated with the production of proinflammatory cytokines and the release of ATP from damaged and necrotic cells. The expression and function of purinergic P2X7receptor (P2X7R) in intestinal epithelial cells (IEC) and its potential role in the “cross talk” between IEC and PMNL have not been explored. The aims of the present study were 1) to examine P2X7R expression in IEC (T84 cells) and in human intestinal biopsies; 2) to detect any changes in P2X7R expression in T84 cells during PMNL transepithelial migration, and during the active and quiescent phases of IBD; and 3) to test whether P2X7R stimulation in T84 monolayers can induce caspase-1 activation and IL-1β release by IEC. We found that a functional ATP-sensitive P2X7R is constitutively expressed at the apical surface of IEC T84 cells. PMNL transmigration regulates dynamically P2X7R expression and alters its distribution from the apical to basolateral surface of IEC during the early phase of PMNL transepithelial migration in vitro. P2X7R expression was weak in intestinal biopsies obtained during the active phase of IBD. We show that activation of epithelial P2X7R is mandatory for PMNL-induced caspase-1 activation and IL-1β release by IEC. Overall, these changes in P2X7R function may serve to tailor the intensity of the inflammatory response and to prevent IL-1β overproduction and inflammatory disease.

scholarly journals Actin dynamics drive microvillar motility and clustering during brush border assembly

2018 ◽  
Author(s):  
Leslie M Meenderink ◽  
Matthew J. Tyska
Keyword(s):  
Epithelial Cells ◽  
Brush Border ◽  
Driving Force ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Actin Dynamics ◽  
Apical Surface ◽  
Actin Assembly ◽  
Brush Borders ◽  
Surface Remodeling

SUMMARYDuring differentiation, transporting epithelial cells generate large arrays of microvilli known as a brush borders to enhance functional capacity. To develop our understanding of brush border formation, we used live cell imaging to visualize apical surface remodeling during early stages of this process. Strikingly, we found that individual microvilli exhibit persistent active motility, translocating across the cell surface at ~0.2 μm/min. Perturbation studies with inhibitors and photokinetic experiments revealed that microvillar motility is driven by actin assembly at the barbed-ends of core bundles, which in turn is linked to robust treadmilling of these structures. Because the apical surface of differentiating epithelial cells is crowded with nascent microvilli, persistent motility promotes collisions between protrusions and ultimately leads to their clustering and consolidation into higher order arrays. Thus, microvillar motility represents a previously unrecognized driving force for apical surface remodeling and maturation during epithelial differentiation.

Adenomatoid Tumor of the Testis, A Mesonephric Hamartoma

1971 ◽  
Vol 29 ◽  
pp. 318-319
Author(s):  
Raoul Fresco ◽  
Mary Chang-Lo
Keyword(s):  
Electron Microscopy ◽  
Epithelial Cells ◽  
Fibrous Tissue ◽  
Salient Feature ◽  
Luminal Surface ◽  
Adenomatoid Tumor ◽  
Ultrastructural Studies ◽  
Epithelial Origin ◽  
Brush Borders ◽  
Cell Processes

Confusion surrounds the nature of the “adenomatoid tumor” of the testis, as evidenced by the large number of synonyms which have been ascribed to it. Various authors have considered the tumor to be of endothelial, mesothelial or epithelial origin. There appears to be no controversy as to the stromal elements of the tumor, which consists mainly of smooth muscle and fibrous tissue. It is the irregular gland-like spaces which have given rise to the numerous theories as to its histogenesis, and even recent ultrastructural studies fail to agree on the origin of these structures.Electron microscopy of a typical intrascrotal adenomatoid tumor showed the gland-like spaces to be lined by epithelial cells (Fig. 1), rich in cytoplasmic tonofibrils and united to each other by numerous desmosomes (Fig. 2). The most salient feature of these epithelial cells was the presence on their luminal surface of numerous long and repeatedly branching microvillous structures of the type known as stereocilia (Fig. 3). These are extremely long slender cell processes which are as much as three to four times the length of those in brush borders.

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