Network Of Cytoskeletal Proteins Research Articles

CircRNAs as Epigenetic Regulators of Integrity in Blood-Brain Barrier Architecture: Mechanisms and Therapeutic Strategies in Multiple Sclerosis.

Multiple sclerosis (MS) is a chronic inflammatory neurodegenerative disease leading to progressive demyelination and neuronal loss, with extensive neurological symptoms. As one of the most widespread neurodegenerative disorders, with an age onset of about 30 years, it turns out to be a socio-health and economic issue, thus necessitating therapeutic interventions currently unavailable. Loss of integrity in the blood-brain barrier (BBB) is one of the distinct MS hallmarks. Brain homeostasis is ensured by an endothelial cell-based monolayer at the interface between the central nervous system (CNS) and systemic bloodstream, acting as a selective barrier. MS results in enhanced barrier permeability, mainly due to the breakdown of tight (TJs) and adherens junctions (AJs) between endothelial cells. Specifically, proinflammatory mediator release causes failure in cytoplasmic exposure of junctions, resulting in compromised BBB integrity that enables blood cells to cross the barrier, establishing iron deposition and neuronal impairment. Cells with a compromised cytoskeletal protein network, fiber reorganization, and discontinuous junction structure can occur, resulting in BBB dysfunction. Recent investigations on spatial transcriptomics have proven circularRNAs (circRNAs) to be powerful multi-functional molecules able to epigenetically regulate transcription and structurally support proteins. In the present review, we provide an overview of the recent role ascribed to circRNAs in maintaining BBB integrity/permeability via cytoskeletal stability. Increased knowledge of the mechanisms responsible for impairment and circRNA's role in driving BBB damage and dysfunction might be helpful for the recognition of novel therapeutic targets to overcome BBB damage and unrestrained neurodegeneration.

The katanin A-subunits KATNA1 and KATNAL1 act co-operatively in mammalian meiosis and spermiogenesis to achieve male fertility.

Katanins, a class of microtubule-severing enzymes, are potent M-phase regulators in oocytes and somatic cells. How the complex and evolutionarily crucial, male mammalian meiotic spindle is sculpted remains unknown. Here, using multiple single and double gene knockout mice, we reveal that the canonical katanin A-subunit KATNA1 and its close paralogue KATNAL1 together execute multiple aspects of meiosis. We show KATNA1 and KATNAL1 collectively regulate the male meiotic spindle, cytokinesis and midbody abscission, in addition to diverse spermatid remodelling events, including Golgi organisation, and acrosome and manchette formation. We also define KATNAL1-specific roles in sperm flagellum development, manchette regulation and sperm-epithelial disengagement. Finally, using proteomic approaches, we define the KATNA1, KATNAL1 and KATNB1 mammalian testis interactome, which includes a network of cytoskeletal and vesicle trafficking proteins. Collectively, we reveal that the presence of multiple katanin A-subunit paralogs in mammalian spermatogenesis allows for 'customised cutting' via neofunctionalisation and protective buffering via gene redundancy.

Forces and Flows at Cell Surfaces.

Flow is an important physiological signal and modulates a variety of cell functions. However, the molecular mechanisms that cells use to sense flow have remained surprisingly opaque. The first steps in flow sensing are likely to occur at the plasma membrane, the fluid barrier between the inside and outside of a cell. This membrane is an organized, two-dimensional molecular array that has both solid and fluid properties. The mobility of membrane proteins and lipids is constrained by complex interactions with the cytoskeletal protein network that supports the membrane. Physiologically relevant flows can only generate tiny forces on individual proteins, smaller than those from thermal noise. However, cells could overcome this problem by sensing micron-scale concentration gradients of extracellular membrane proteins. This is possible in cell plasma membranes because their particular physical state allows flow to sort membrane proteins laterally over the extracellular surface.

Microscale mechanics of actin-vimentin composite networks.

Most eukaryotic cells contain a complex network of cytoskeletal proteins comprised of filamentous actin, microtubule, and intermediate filaments. The ability of cells to resist mechanical deformation, transport cargo, and change shape during movement depends on these cytoskeletal proteins. Several previous studies have mainly focused on exploring the mechanics of in vitro reconstituted networks consisting of only one of these components, which may not be able to predict the properties of composite networks. Thus, investigating the interplay between the components as a composite is central to understanding the underlying biophysical principles. Here, we couple optical tweezers microrheology with confocal imaging to measure the viscoelastic response and structural dynamics of in vitro reconstituted filamentous actin (F-actin) and vimentin intermediate filaments (VIFs) composite networks by varying the relative concentration of the proteins. Our measurements shed new light on how VIFs and F-actin work synergistically during the various cellular processes.

The perinuclear region concentrates disordered proteins with predicted phase separation distributed in a 3D network of cytoskeletal filaments and organelles

Membraneless organelles have emerged during the evolution of eukaryotic cells as intracellular domains in which multiple proteins organize into complex structures to perform specialized functions without the need of a lipid bilayer compartment. Here we describe the perinuclear space of eukaryotic cells as a highly organized network of cytoskeletal filaments that facilitates assembly of biomolecular condensates. Using bioinformatic analyses, we show that the perinuclear proteome is enriched in intrinsic disorder with several proteins predicted to undergo liquid-liquid phase separation. We also analyze immunofluorescence and transmission electron microscopy images showing the association between the nucleus and other organelles, such as mitochondria and lysosomes, or the labeling of specific proteins within the perinuclear region of cells. Altogether our data support the existence of a perinuclear dense sub-micron region formed by a well-organized three-dimensional network of structural and signaling proteins, including several proteins containing intrinsically disordered regions with phase behavior. This network of filamentous cytoskeletal proteins extends a few micrometers from the nucleus, contributes to local crowding, and organizes the movement of molecular complexes within the perinuclear space. Our findings take a key step towards understanding how membraneless regions within eukaryotic cells can serve as hubs for biomolecular condensates assembly, in particular the perinuclear space. Finally, evaluation of the disease context of the perinuclear proteins revealed that alterations in their expression can lead to several pathological conditions, and neurological disorders and cancer are among the most frequent.

A NanoFE simulation-based surrogate machine learning model to predict mechanical functionality of protein networks from live confocal imaging

Sub-cellular mechanics plays a crucial role in a variety of biological functions and dysfunctions. Due to the strong structure-function relationship in cytoskeletal protein networks, light can be shed on their mechanical functionality by investigating their structures. Here, we present a data-driven approach employing a combination of confocal live imaging of fluorescent tagged protein networks, in silico mechanical experiments and machine learning to investigate this relationship. Our designed image processing and nanoFE mechanical simulation framework resolves the structure and mechanical behaviour of cytoskeletal networks and the developed gradient boosting surrogate models linking network structure to its functionality. In this study, for the first time, we perform mechanical simulations of Filamentous Temperature Sensitive Z (FtsZ) complex protein networks with realistic network geometry depicting its skeletal functionality inside organelles (here, chloroplasts) of the moss Physcomitrella patens. Training on synthetically produced simulation data enables predicting the mechanical characteristics of FtsZ network purely based on its structural features (R2⩾0.93), therefore allowing to extract structural principles enabling specific mechanical traits of FtsZ, such as load bearing and resistance to buckling failure in case of large network deformation.

TiO2 nanoparticles disrupt cell adhesion and the architecture of cytoskeletal networks of human osteoblast-like cells in a size dependent manner.

Human exposure to titanium dioxide nanoparticles (nano-TiO2 ) is increasing. An internal source of nano-TiO2 is represented by titanium-based orthopedic and dental implants can release nanoparticles (NPs) upon abrasion. Little is known about how the size of NPs influences their interaction with cytoskeletal protein networks and the functional/homeostatic consequences that might follow at the implant-bone interface with regard to osteoblasts. We investigated the effects of size of anatase nano-TiO2 on SaOS-2 human osteoblast-like cells exposed to clinically relevant concentrations (0.05, 0.5, 5 mg/L) of 5 and 40 nm spherical nano-TiO2 . Cell viability and proliferation, adhesion, spread and migration were assessed, as well as the orientation of actin and microtubule cytoskeletal networks. The phosphorylation of focal adhesion kinase (p-FAKY397 ) and the expression of vinculin in response to nano-TiO2 were also assessed. Treatment with nano-TiO2 disrupted the actin and microtubule cytoskeletal networks leading to morphological modifications of SaOS-2 cells. The phosphorylation of p-FAKY397 and the expression of vinculin were also modified depending on the particle size, which affected cell adhesion. Consequently, the cell migration was significantly impaired in the 5 nm-exposed cells compared to unexposed cells. The present work shows that the orientation of cytoskeletal networks and the focal adhesion proteins and subsequently the adhesion, spread and migration of SaOS-2 cells were affected by the selected nano-TiO2 in a size dependent manner. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 2582-2593, 2018.

MicrobeJ, a tool for high throughput bacterial cell detection and quantitative analysis.

Single cell analysis of bacteria and subcellular protein localization dynamics has shown that bacteria have elaborate life cycles, cytoskeletal protein networks, and complex signal transduction pathways driven by localized proteins. The volume of multi-dimensional images generated in such experiments and the computation time required to detect, associate, and track cells and subcellular features pose considerable challenges, especially for high-throughput experiments. Therefore, there is a need for a versatile, computationally efficient image analysis tool capable of extracting the desired relationships from images in a meaningful and unbiased way. Here we present MicrobeJ, a plug-in for the open-source platform ImageJ. MicrobeJ provides a comprehensive framework to process images derived from a wide variety of microscopy experiments with special emphasis on large image sets. It performs the most common intensity and morphology measurements as well as customized detection of poles, septa, fluorescent foci, and organelles, determines their sub-cellular localization with sub-pixel resolution, and tracks them over time. Because a dynamic link is maintained between the images, measurements, and all data representations derived from them, the editor and suite of advanced data presentation tools facilitates the image analysis process and provides a robust way to verify the accuracy and veracity of the data.

Molecular Control of Actin Dynamics In Vivo: Insights from Drosophila.

The actin cytoskeleton provides mechanical support for cells and generates forces to drive cell shape changes and cell migration in morphogenesis. Molecular understanding of actin dynamics requires a genetically traceable model system that allows interdisciplinary experimental approaches to elucidate the regulatory network of cytoskeletal proteins in vivo. Here, we will discuss some examples of how advances in Drosophila genetics and high-resolution imaging techniques contribute to the discovery of new actin functions, signaling pathways, and mechanisms of actin regulation in vivo.

Prolonged high fat diet ingestion, obesity, and type 2 diabetes symptoms correlate with phenotypic plasticity in myenteric neurons and nerve damage in the mouse duodenum.

Symptoms of diabetic gastrointestinal dysmotility indicate neuropathy of the enteric nervous system. Long-standing diabetic enteric neuropathy has not been fully characterized, however. We used prolonged high fat diet ingestion (20 weeks) in a mouse model to mimic human obese and type 2 diabetic conditions, and analyzed changes seen in neurons of the duodenal myenteric plexus. Ganglionic and neuronal size, number of neurons per ganglionic area, density indices of neuronal phenotypes (immunoreactive nerve cell bodies and varicosities per ganglion or tissue area) and nerve injury were measured. Findings were compared with results previously seen in mice fed the same diet for 8 weeks. Compared to mice fed standard chow, those on a prolonged high fat diet had smaller ganglionic and cell soma areas. Myenteric VIP- and ChAT-immunoreactive density indices were also reduced. Myenteric nerve fibers were markedly swollen and cytoskeletal protein networks were disrupted. The number of nNOS nerve cell bodies per ganglia was increased, contrary to the reduction previously seen after 8 weeks, but the density index of nNOS varicosities was reduced. Mice fed high fat and standard chow diets experienced an age-related reduction in total neurons, with bias towards neurons of sensory phenotype. Meanwhile, ageing was associated with an increase in excitatory neuronal markers. Collectively, these results support a notion that nerve damage underlies diabetic symptoms of dysmotility, and reveals adaptive ENS responses to the prolonged ingestion of a high fat diet. This highlights a need to mechanistically study long-term diet-induced nerve damage and age-related impacts on the ENS.

Alteration Young’s moduli by protein 4.1 phosphorylation play a potential role in the deformability development of vertebrate erythrocytes

The mechanical properties of vertebrate erythrocytes depend on their cytoskeletal protein networks. Membrane skeleton proteins spectrin and protein 4.1 (4.1R) cross-link with actin to maintain membrane stability under mechanical stress. Phosphorylation of 4.1R alters the affinity of 4.1R for spectrin–actin binding and this modulates the mechanical properties of human erythrocytes. In this study, phorbol 12-myristate-13-acetate (PMA)-induced phosphorylation of 4.1R was tested, erythrocyte deformability was determined and the erythrocyte elastic modulus was detected in human, chick, frog and fish. Furthermore, amino acid sequences of the functionally important domains of 4.1R were analyzed. Results showed that PMA-induced phosphorylation of 4.1R decreased erythrocyte deformability and this property was stable after 1h. The values of Young’s modulus alteration gradually decreased from human to fish (0.388±0.035kPa, 0.219±0.022kPa, 0.191±0.036kPa and 0.141±0.007kPa). Ser-312 and Ser-331 are located within the consensus sequence recognized by protein kinase C (PKC); however, Ser-331 in zebrafish was replaced by Ala-331. The sequence of the 8 aa motif from vertebrate 4.1R showed only one amino acid mutation in frog and numerous substitutions in fish. Analyses of Young’s modulus suggested that the interaction between 4.1R with the spectrin–actin binding domain may have a special relationship with the development of erythrocyte deformability. In addition, amino acid mutations in 4.1R further supported this relationship. Thus, we hypothesize that alteration of membrane skeleton protein binding affinity may play a potential role in the development of erythrocyte deformability, and alteration of Young’s modulus values may provide a method for determining the deformability development of vertebrate erythrocytes.

The degree of resistance of erythrocyte membrane cytoskeletal proteins to supra-physiologic concentrations of calcium: an in vitro study.

Calcium is a key regulator of cell dynamics. Dysregulation of its cytosolic concentration is implicated in the pathophysiology of several diseases. This study aimed to assess the effects of calcium on the network of membrane cytoskeletal proteins. Erythrocyte membranes were obtained from eight healthy donors and incubated with 250 µM and 1.25 mM calcium solutions. Membrane cytoskeletal proteins were quantified using SDS-PAGE at baseline and after 3 and 5 days of incubation. Supra-physiologic concentrations of calcium (1.25 mM) induced a significant proteolysis in membrane cytoskeletal proteins, compared with magnesium (p < 0.001). Actin exhibited the highest sensitivity to calcium-induced proteolysis (6.8 ± 0.3 vs. 5.3 ± 0.6, p < 0.001), while spectrin (39.9 ± 1.0 vs. 40.3 ± 2.0, p = 0.393) and band-6 (6.3 ± 0.3 vs. 6.8 ± 0.8, p = 0.191) were more resistant to proteolysis after incubation with calcium in the range of endoplasmic reticulum concentrations (250 µM). Aggregation of membrane cytoskeletal proteins was determined after centrifugation and was significantly higher after incubation with calcium ions compared with control, EDTA and magnesium solutions (p < 0.001). In a supra-physiologic range of 1.25-10 mM of calcium ions, there was a nearly perfect linear relationship between calcium concentration and aggregation of erythrocyte membrane cytoskeletal proteins (R(2) = 0.971, p < 0.001). Our observation suggests a strong interaction between calcium ions and membrane cytoskeletal network. Cumulative effects of disrupted calcium homeostasis on cytoskeletal proteins need to be further investigated at extended periods of time in disease states.

Protein coat keeps axons buttoned up

![Figure][1] After 4 days in vitro (left), the AIS of a hippocampal neuron contains a microtubule bundle. 17 days later (right), the microtubules are coated by a dense, fibrillar–globular coat. Jones et al. reveal how a submembranous network of cytoskeletal proteins helps maintain

Technological advances for deciphering the complexity of psychiatric disorders: merging proteomics with cell biology

Proteomic studies have increased our understanding of the molecular pathways affected in psychiatric disorders. Mass spectrometry and two-dimensional gel electrophoresis analyses of post-mortem brain samples from psychiatric patients have revealed effects on synaptic, cytoskeletal, antioxidant and mitochondrial protein networks. Multiplex immunoassay profiling studies have found alterations in hormones, growth factors, transport and inflammation-related proteins in serum and plasma from living first-onset patients. Despite these advances, there are still difficulties in translating these findings into platforms for improved treatment of patients and for discovery of new drugs with better efficacy and side effect profiles. This review describes how the next phase of proteomic investigations in psychiatry should include stringent replication studies for validation of biomarker candidates and functional follow-up studies which can be used to test the impact on physiological function. All biomarker candidates should now be tested in series with traditional and emerging cell biological approaches. This should include investigations of the effects of post-translational modifications, protein dynamics and network analyses using targeted proteomic approaches. Most importantly, there is still an urgent need for development of disease-relevant cellular models for improved translation of proteomic findings into a means of developing novel drug treatments for patients with these life-altering disorders.

Abstract 4918: A Rho-directed ROCK:mDia2 signaling axis drives morphological plasticity and invasive transitions in EOC spheroids.

Abstract Epithelial ovarian cancer (EOC) is often detected late after it has metastasized within the peritoneal cavity. EOC cells are typically detected in the peritoneal ascites as single cells, multi-cell clusters or highly ordered, compacted spheroids. Compacted EOC spheroids represent a more invasive and tumorigenic population of ovarian tumor cells both in vitro and in vivo and are highly chemoresistant. Yet little is known about molecular mechanisms triggering an invasive transition within single cells emanating from these clinically dangerous structures that are fundamental to EOC dissemination and disease progression. In this regard, the field of molecular players driving EOC spheroid adhesion/migration/invasion has remained relatively static over the last decade. However, spheroid formation and ensuing invasion requires tight regulation of the actin cytoskeleton; therefore, a thorough understanding of the contribution of proteins regulating the spheroid F-actin cytoskeleton in both cell-cell adhesions and adhesion to/transmigration through peritoneal mesothelium is essential for progress in therapy of metastatic EOC. The mDia formin protein family has emerged as key regulators of cytoskeletal dynamics during cancer cell division and migration. mDia2 assembles non-branched F-actin networks underlying cell-cell junctions and is critical for driving single cell mesenchymal motility. Loss of mDia2 expression and/or function drives an amoeboid morphological transition that enhanced cell invasion and metastasis in breast and prostate cancer models. Our data strongly implicate a tightly controlled network of cytoskeletal proteins including the GTPase RhoA and its effectors mDia2 formin and ROCK in driving highly adaptable single cell motility from EOC spheroids within three-dimensional (3D) matrices. Here we show that mDia2 is involved not only in EOC spheroid growth and formation, yet is enriched in the invasive cellular front of spheroids embedded in 3D collagen gels. However, mDia2 disruption significantly enhances invasive dissemination of amoeboid single cells from EOC spheroids embedded in collagen gels. ROCK, which promotes myosin-based cellular contractility and amoeboid motility programs, similarly played a role in spheroid formation, yet its disruption via Y27632 treatment significantly inhibited spheroid invasion while driving an elongated cellular morphology. Treatment of invasive mDia2-depleted EOC spheroids with Y27632 blocked invasion more effectively than Y27632 treatment alone, indicating that under conditions of mDia2 depletion, invasion is dependent upon Rho-directed ROCK activity. Collectively, these data indicate interplay between a Rho-directed ROCK and mDia2 signaling axis in driving invasive transitions and morphological plasticity in EOC spheroids. Citation Format: Krista Pettee, Andrea Nestor-Kalinoski, Kathryn M. Eisenmann. A Rho-directed ROCK:mDia2 signaling axis drives morphological plasticity and invasive transitions in EOC spheroids. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4918. doi:10.1158/1538-7445.AM2013-4918

Deoxyelephantopin impedes mammary adenocarcinoma cell motility by inhibiting calpain-mediated adhesion dynamics and inducing reactive oxygen species and aggresome formation

We previously showed that deoxyelephantopin (DET), a plant sesquiterpene lactone, exhibits more profound suppression than paclitaxel (PTX) of lung metastasis of mammary adenocarcinoma TS/A cells in mice. Proteomics studies suggest that DET affects actin cytoskeletal protein networks and downregulates calpain-mediated proteolysis of several actin-associated proteins, whereas PTX mainly interferes with microtubule proteins. Here, DET was observed to significantly deregulate adhesion formation in TS/A cells, probably through inhibition of m-calpain activity. Epithelial growth factor (EGF)-mediated activation of Rho GTPase Rac1 and formation of lamellipodia in TS/A cells were remarkably suppressed by DET treatment. Further, DET impaired vesicular trafficking of EGF and induced protein carbonylation and formation of centrosomal aggregates in TS/A cells. DET-induced reactive oxygen species were observed to be the upstream stimulus for the formation of centrosomal ubiquitinated protein aggregates that might subsequently restrict cancer cell motility. PTX, however, caused dramatic morphological changes, interfered with microtubule networking, and moderately inhibited calpain-mediated cytoskeletal and focal adhesion protein cleavage in TS/A cells. This study provides novel mechanistic insights into the pharmacological action of DET against metastatic mammary cell migration and suggests that modulation of oxidative stress might be a potential strategy for treatment of metastatic breast cancer.

Evidence for Altered Communication Between the L-Type Ca2+ Channel and Mitochondria in a Model of Cardiomyopathy

Progression of cardiac hypertrophy to failure and development of many cardiomyopathies involves myocyte remodeling, disorganisation of cytoskeletal proteins and reduced energy metabolism. The mechanisms by which cytoskeletal disruption leads to mitochondrial dysfunction are poorly understood.Calcium influx through the L-type Ca2+ channel (LTCC) is a requirement for contraction in the heart. The LTCC can influence mitochondrial superoxide production, NADH production and metabolic activity in a calcium-dependent manner. Activation of the channel can also increase mitochondrial membrane potential (Ψm) in a calcium-independent manner. This response is dependent upon the cytoskeleton. We hypothesized that disruption of normal cytoskeletal architecture will result in altered communication between the LTCC and mitochondria. We investigated this hypothesis in the murine model of Duchenne Muscular Dystrophy(mdx). Myocytes from 8 week old mdx mice that exhibit disorganised cytoskeletal protein networks but not yet overt cardiomyopathy, demonstrated significantly slower LTCC inactivation rate (26.2±1.8ms, n=13 vs 21.1±1.3ms, n=16; p<0.05) and significantly greater calcium influx (681±40nM, n=7 vs 432±51nM, n=5). However activation of the LTCC did not increase Ψm (n=6) or metabolic activity (measured as formation of formazan from tetrazolium salt) in mdx myocytes (n=8). Application of 4,4′diisothiocyano-2,2′-stilbenedisulfonic acid, that blocks anion transport mimicked the response of BayK(-) in control myocytes (n=4) and “restored” the increase in Ψm in mdx myocytes (n=4).The activities of the mitochondrial respiratory complexes were normal in mitochondria isolated from 8 week old mdx hearts and from 40 week old mdx hearts that had developed cardiomyopathy. We conclude that the communication between the L-type Ca2+ channel and mitochondria is altered in the dystrophin-deficient cardiomyocyte. This appears to involve an alteration in the association between the channel protein, actin filaments and mitochondrial anion transport and precedes the development of cardiomyopathy.

Disruption of Spectrin-Like Cytoskeleton in Differentiating Keratinocytes by PKCδ Activation Is Associated with Phosphorylated Adducin

Spectrin is a central component of the cytoskeletal protein network in a variety of erythroid and non-erythroid cells. In keratinocytes, this protein has been shown to be pericytoplasmic and plasma membrane associated, but its characteristics and function have not been established in these cells. Here we demonstrate that spectrin increases dramatically in amount and is assembled into the cytoskeleton during differentiation in mouse and human keratinocytes. The spectrin-like cytoskeleton was predominantly organized in the granular and cornified layers of the epidermis and disrupted by actin filament inhibitors, but not by anti-mitotic drugs. When the cytoskeleton was disrupted PKCδ was activated by phosphorylation on Thr505. Specific inhibition of PKCδ(Thr505) activation with rottlerin prevented disruption of the spectrin-like cytoskeleton and the associated morphological changes that accompany differentiation. Rottlerin also inhibited specific phosphorylation of the PKCδ substrate adducin, a cytoskeletal protein. Furthermore, knock-down of endogenous adducin affected not only expression of adducin, but also spectrin and PKCδ, and severely disrupted organization of the spectrin-like cytoskeleton and cytoskeletal distribution of both adducin and PKCδ. These results demonstrate that organization of a spectrin-like cytoskeleton is associated with keratinocytes differentiation, and disruption of this cytoskeleton is mediated by either PKCδ(Thr505) phosphorylation associated with phosphorylated adducin or due to reduction of endogenous adducin, which normally connects and stabilizes the spectrin-actin complex.

Characterization of cellular elastic modulus using structure based double layer model

The mechanical characterization of cells is important for understanding cellular behavior and physiological functions. We used atomic force microscopy (AFM) to obtain a force-displacement curve and estimate the elastic modulus of hepatocellular carcinoma cells (HEP-G2) utilizing both linear Hertz-Sneddon (HS) and non-linear elastic models. In order to overcome the limitations of HS model, which assumes a linear homogeneous cell body, a cell is modeled as a double-layered body with an outer cytoplasmic layer made mostly of interconnected fibers of cytoskeleton proteins and a nucleus. By disrupting all cytoskeletal protein networks, we estimate the elastic modulus of the core nucleus using FEM for a single ellipsoid. Based on the nucleic modulus and cellular dimensions found by 3D confocal imaging, we develop a novel double-layered cellular (DLC) finite element model. The DLC model provides a more reliable estimate of the elastic modulus of the cell than conventionally used HS model and correlates closely with experimental results.

Single-cell electrical lysis of erythrocytes detects deficiencies in the cytoskeletal protein network

The network of erythrocyte cytoskeletal proteins significantly influences erythrocyte physical and biological properties. Here we show that the kinetics of erythrocyte lysis during exposure to an electric field is sensitively correlated with defects in the cytoskeletal network. Histograms compiled from single-cell electrical lysis data show characteristics of erythrocyte populations that are deficient in a specific cytoskeletal protein, revealing the presence of cell subpopulations.