Keratin 8 Phosphorylation by Protein Kinase C δ Regulates Shear Stress-mediated Disassembly of Keratin Intermediate Filaments in Alveolar Epithelial Cells

Ridge, Karen M.; Linz, Laura A.; Flitney, Frederick W.; Kuczmarski, Edward R.; Chou, Ying Hao; Omary, M. Bishr; Sznajder, Jacob I.; Goldman, Robert D.

doi:10.1074/jbc.m504239200

Cited by 120 publications

(133 citation statements)

References 43 publications

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“…On mechanical stretch, keratinocytes expressing the disease mutation KRT10 Arg156His show stronger activation of p38 MAPK compared with control transfectants, which ultimately might drive hyperproliferation typical of the corresponding keratinopathy (Obarzanek-Fojt et al 2011). In lung epithelial cells, shear stress, but not stretch, caused hyperphosphorylation and disassembly of KRT8 and KRT18-containing IF, regulated by PKC-d (Ridge et al 2005). Blocking the shear stress -mediated keratin hyperphosphorylation and reorganization decreased cell viability and increased apoptosis.…”

Section: Impact Of Mechanical Stretch On the Desmosome -Keratin Complexmentioning

confidence: 99%

Desmosomes and Intermediate Filaments: Their Consequences for Tissue Mechanics

Hatzfeld

Keil

Magin

2017

Cold Spring Harb Perspect Biol

116

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Adherens junctions (AJs) and desmosomes connect the actin and keratin filament networks of adjacent cells into a mechanical unit. Whereas AJs function in mechanosensing and in transducing mechanical forces between the plasma membrane and the actomyosin cytoskeleton, desmosomes and intermediate filaments (IFs) provide mechanical stability required to maintain tissue architecture and integrity when the tissues are exposed to mechanical stress. Desmosomes are essential for stable intercellular cohesion, whereas keratins determine cell mechanics but are not involved in generating tension. Here, we summarize the current knowledge of the role of IFs and desmosomes in tissue mechanics and discuss whether the desmosome-keratin scaffold might be actively involved in mechanosensing and in the conversion of chemical signals into mechanical strength.

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Section: Impact Of Mechanical Stretch On the Desmosome -Keratin Complexmentioning

confidence: 99%

Desmosomes and Intermediate Filaments: Their Consequences for Tissue Mechanics

Hatzfeld

Keil

Magin

2017

Cold Spring Harb Perspect Biol

116

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“…Rat alveolar epithelial type II cells (AECs) were isolated from pathogen-free male Sprague-Dawley rats (200 -225 g), as described in ref. 2. In brief, the lungs were perfused by the pulmonary artery and lavaged and digested with elastase (30 units/ml, Worthington Biochemical).…”

Section: Methodsmentioning

confidence: 99%

“…They express cell-type-specific keratin intermediate filament (KIF) proteins, which assemble into complex networks of 10-nm-diameter fibers made up of heterodimers of acidic type I and neutral-basic type II isoforms (1). The primary rat alveolar epithelial cells (AECs) used in this study contain polymers of keratin 8 and 18 (K8/K18) (2). In general, intermediate filaments maintain the mechanical integrity of cells (3)(4)(5).…”

mentioning

confidence: 99%

Micromechanical properties of keratin intermediate filament networks

Sivaramakrishnan

DeGiulio²,

Lóránd³

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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106

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E pithelial cells form an indispensable physical barrier to withstand mechanical forces in the surrounding environment. They express cell-type-specific keratin intermediate filament (KIF) proteins, which assemble into complex networks of 10-nm-diameter fibers made up of heterodimers of acidic type I and neutral-basic type II isoforms (1). The primary rat alveolar epithelial cells (AECs) used in this study contain polymers of keratin 8 and 18 (K8/K18) (2). In general, intermediate filaments maintain the mechanical integrity of cells (3-5). For example, K8 knockout mutation is embryonic lethal in the mouse because of hemorrhaging in the liver (6), and mutations in epidermal keratins cause skin-blistering diseases (7).The eukaryotic cytoskeleton comprises intermediate filaments (IFs), microfilaments (MFs), and microtubules (MTs). All are dynamic structures, undergoing continuous assembly and disassembly in the intracellular environment (8-10). However, KIF assembly differs significantly from MT and MF (8,9), in that it is nucleotide-independent and that the critical concentration of subunits required for assembly is lower than required for MT and MF. Intermediate filaments resist conditions such as detergent treatment with or without high-salt extraction (11), which disassemble and solubilize the constituent proteins of MT and MF. KIF can be efficiently disassembled only by employing high concentrations of denaturants such as urea (4). We took advantage of the robust nature of the KIF network to measure its mechanical properties, without perturbation by MTs and MFs, in AECs by using particle-tracking microrheology (PTM) (12-14).We show that there is a correlation between the architecture of the KIF network and the local mechanical properties in AECs, and that shear stress applied across the cell surface causes a structural remodeling of KIF and a substantial increase in the elastic modulus of the network. Results and DiscussionShear Stress Alters the Mesh Size of KIF Networks. To understand the specific structural role of the KIF network, we quantified its mechanical properties in the absence of MT and MF. The intact network can be prepared in AECs by detergent treatment and high-salt extraction (4, 10), conditions that remove membranes, soluble cytoplasmic components, and MT and MF (15). To ensure that the extraction procedure did not alter cell shape and size or significantly affect the three-dimensional organization of the KIF network, AECs were transiently transfected with a construct expressing GFP-K18 (10). Cells were imaged pre-and postextraction by confocal microscopy; neither cell height (total depth of the Z-stack) nor the organization of the KIF was altered [supporting information (SI) Fig. 5].The ultrastructure of the KIF network, now devoid of MT and MF (SI Fig. 6), was examined by electron microscopy (11, 15). Cell images were divided into three zones that did not vary significantly across the cell population (SI Fig. 7A). As shown in Fig. 1A, the mesh size within the KIF network, defined as the mean ...

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“…For example, the major cytoskeletal components of alveolar epithelial cells can be disassembled by shear stresses of the order of 30 dynes/ cm 2 . 45 It is also well known that …”

Section: Application To Pulmonary Airway Closurementioning

confidence: 99%

The effect of viscoelasticity on the stability of a pulmonary airway liquid layer

2010

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The lungs consist of a network of bifurcating airways that are lined with a thin liquid film. This film is a bilayer consisting of a mucus layer on top of a periciliary fluid layer. Mucus is a non-Newtonian fluid possessing viscoelastic characteristics. Surface tension induces flows within the layer, which may cause the lung's airways to close due to liquid plug formation if the liquid film is sufficiently thick. The stability of the liquid layer is also influenced by the viscoelastic nature of the liquid, which is modeled using the Oldroyd-B constitutive equation or as a Jeffreys fluid. To examine the role of mucus alone, a single layer of a viscoelastic fluid is considered. A system of nonlinear evolution equations is derived using lubrication theory for the film thickness and the film flow rate. A uniform film is initially perturbed and a normal mode analysis is carried out that shows that the growth rate g for a viscoelastic layer is larger than for a Newtonian fluid with the same viscosity. Closure occurs if the minimum core radius, R min ͑t͒, reaches zero within one breath. Solutions of the nonlinear evolution equations reveal that R min normally decreases to zero faster with increasing relaxation time parameter, the Weissenberg number We. For small values of the dimensionless film thickness parameter , the closure time, t c , increases slightly with We, while for moderate values of , ranging from 14% to 18% of the tube radius, t c decreases rapidly with We provided the solvent viscosity is sufficiently small. Viscoelasticity was found to have little effect for Ͼ0.18, indicating the strong influence of surface tension. The film thickness parameter and the Weissenberg number We also have a significant effect on the maximum shear stress on tube wall, max͑ w ͒, and thus, potentially, an impact on cell damage. Max͑ w ͒ increases with for fixed We, and it decreases with increasing We for small We provided the solvent viscosity parameter is sufficiently small. For large Ϸ 0.2, there is no significant difference between the Newtonian flow case and the large We cases.

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Keratin 8 Phosphorylation by Protein Kinase C δ Regulates Shear Stress-mediated Disassembly of Keratin Intermediate Filaments in Alveolar Epithelial Cells

Cited by 120 publications

References 43 publications

Desmosomes and Intermediate Filaments: Their Consequences for Tissue Mechanics

Desmosomes and Intermediate Filaments: Their Consequences for Tissue Mechanics

Micromechanical properties of keratin intermediate filament networks

The effect of viscoelasticity on the stability of a pulmonary airway liquid layer

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