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At least 811 records · Page 45Linked to original sources
PubMed1998

Integrin signalling and tyrosine phosphorylation: just the FAKs?

The integrin family of transmembrane receptors have long been recognized for their structural roles in linking extracellular matrix proteins with the cellular actin cytoskeleton to regulate cell shape, cell migration and tissue architecture. Lately, it has become clear that integrin receptors can initiate intracellular signals that synergize with those from growth-factor receptor protein-tyrosine kinases in modulating cell growth. This review describes our current knowledge of integrin-stimulated tyrosine phosphorylation events and downstream signalling pathways, with emphasis on the recent progress made in understanding the molecular pathways linking the integrin receptors with mitogen-activated protein (MAP) kinase cascades.

Animals↗
PubMed1999

Tumour-associated E-cadherin mutations alter cellular morphology, decrease cellular adhesion and increase cellular motility.

A major function of the cell-to-cell adhesion molecule E-cadherin is the maintenance of cell adhesion and tissue integrity. E-cadherin deficiency in tumours leads to changes in cell morphology and motility, so that E-cadherin is considered to be a suppressor of invasion. In this study we investigated the functional consequences of three tumour-associated gene mutations that affect the extracellular portion of E-cadherin: in-frame deletions of exons 8 or 9 and a point mutation in exon 8, as they were found in human gastric carcinomas. Human MDA-MB-435S breast carcinoma cells and mouse L fibroblasts were stably transfected with the wild-type and mutant cDNAs, and the resulting changes in localization of E-cadherin, cell morphology, strength of calcium-dependent aggregation as well as cell motility and actin cytoskeleton organization were studied. We found that cells transfected with wild-type E-cadherin showed an epitheloid morphology, while all cell lines expressing mutant E-cadherin exhibited more irregular cell shapes. Cells expressing E-cadherin mutated in exon 8 showed the most scattered appearance, whereas cells with deletion of exon 9 had an intermediate state. Mutant E-cadherins were localized to the lateral regions of cell-to-cell contact sites. Additionally, both exon 8-mutated E-cadherins showed apical and perinuclear localization, and actin filaments were drastically reduced. MDA-MB-435S cells with initial calcium-dependent cell aggregation exhibited decreased aggregation and, remarkably, increased cell motility, when mutant E-cadherin was expressed. Therefore, we conclude that these E-cadherin mutations may not simply affect cell adhesion but may act in a trans-dominant-active manner, i.e. lead to increased cell motility. Our study suggests that E-cadherin mutations affecting exons 8 or 9 are the cause of multiple morphological and functional disorders and could induce the scattered morphology and the invasive behaviour of diffuse type-gastric carcinomas.

Actins↗
PubMed1998

Iqg1p, a yeast homologue of the mammalian IQGAPs, mediates cdc42p effects on the actin cytoskeleton.

The Rho-type GTPase Cdc42p has been implicated in diverse cellular functions including cell shape, cell motility, and cytokinesis, all of which involve the reorganization of the actin cytoskeleton. Targets of Cdc42p that interface the actin cytoskeleton are likely candidates for mediating cellular activities. In this report, we identify and characterize a yeast homologue for the mammalian IQGAP, a cytoskeletal target for Cdc42p. The yeast IQGAP homologue, designated Iqg1p, displays a two-hybrid interaction with activated Cdc42p and coimmunoprecipitates with actin filaments. Deletion of IQG1 results in a temperature-sensitive lethality and causes aberrant morphologies including elongated and round multinucleated cells. This together with its localization at the mother-bud neck, suggest that Iqg1p promotes budding and cytokinesis. At restrictive temperatures, the vacuoles of the mutant cells enlarge and vesicles accumulate in the bud. Interestingly, Iqg1p shows two-hybrid interactions with the ankyrin repeat-containing protein, Akr1p (Kao, L.-R., J. Peterson, J. Ruiru, L. Bender, and A. Bender. 1996. Mol. Cell. Biol. 16:168-178), which inhibits pheromone signaling and appears to promote cytokinesis and/or trafficking. We also show two-hybrid interactions between Iqg1p and Afr1p, a septin-binding protein involved in projection formation (Konopka, J.B., C. DeMattei, and C. Davis. 1995. Mol. Cell. Biol. 15:723-730). We propose that Iqg1p acts as a scaffold to recruit and localize a protein complex involved in actin-based cellular functions and thus mediates the regulatory effects of Cdc42p on the actin cytoskeleton.

Actins↗
PubMed2002

The Edg family G protein-coupled receptors for lysophospholipids: their signaling properties and biological activities.

Sphingosine-1-phosphate (S1P) and lysophosphatidic acid (LPA) are blood-borne lysophospholipids with a wide spectrum of biological activities, which include stimulation of cell growth, prevention of apoptosis, regulation of actin cytoskeleton, and modulation of cell shape, cell migration, and invasion. Activated platelets appear to be a major source of both S1P and LPA in blood. Despite the diversity of their biosynthetic origins, they are considered to share substantial structural similarity. Indeed, recent investigation has revealed that S1P and LPA act via a single family of G protein-coupled receptors designated as Edg. Thus, the Edg isoforms, Edg1 (also called S1P(1)), Edg5 (S1P(2)), Edg3 (S1P(3)), Edg6 (S1P(4)), and Edg8 (S1P(5)), are specific receptors for S1P (and SPC with a lower affinity), whereas Edg2 (LPA(1)), Edg4 (LPA(2)), and Edg7 (LPA(3)) serve as receptors specific for LPA. Each receptor isoform displays a unique tissue expression pattern and coupling to a distinct set of heterotrimeric G proteins, leading to the activation of an isoform-specific panel of multiple intracellular signaling pathways. Recent studies on knockout mice have unveiled non-redundant Edg receptor functions that are essential for normal development and vascular maturation. In addition, the Edg lysophospholipid signaling system may play a role in modulating cell motility under such pathological conditions as inflammation, tumor cell dissemination and vascular remodeling.

Animals↗
PubMed1976

Quantitative determination of N-acetylglucosamine residues at the non-reducing ends of peptidoglycan chains by enzymic attachment of [14C]-D-galactose.

The ability of human milk galactosyltransferase to attach D-galactose residues quantitatively to the C-4 of N-acetylglucosamine moieties at the ends of oligosaccharides has been utilized for the specific labeling and quantitative determination of the chain length of the glycan moiety of the bacterial cell wall. The average polysaccharide chain length of the soluble, uncrosslinked peptidoglycan secreted by Micrococcus luteus cells on incubation with penicillin G was studied with this technique and found to be approximately 70 hexosamines long. Furthermore, the peptidoglycan chain length of Escherichia coli sacculi of different cell shapes and dimensions was determined both in rod-shaped cells and in filaments induced by temperature shift of a division mutant or by addition of cephalexin or nalidixic acid. The average chain length found in most of these sacculi was between 70 and 100 hexosamines long. Small spherical 'mini' cells had chain lengths similar to those of the isogenic rod-like cells.

Acetylglucosamine↗
PubMed1989

Developmental expression of the glial fibrillary acidic protein mRNA in the central nervous system and in cultured astrocytes.

The expression of glial fibrillary acidic protein (GFAP)-mRNA during mouse brain development and in astroglial primary cultures has been investigated by using two approaches: Northern-blot evaluation using a specific cDNA probe, and cell-free translation associated with immunoprecipitation. During brain maturation (4-56 days postnatal), the GFAP-mRNA underwent a biphasic evolution. An increase was observed between birth and day 15 (i.e., during the period of astroglial proliferation), which was followed by a decrease until day 56 (i.e., during astroglial cell differentiation). At older stages (300 days), an increase was observed, which might reflect gliosis. During astroglial in vitro development (7-32 days in culture), the GFAP-mRNA showed similar variations. An increase, observed during the period of astroglial proliferation (7-18 days), was followed by a decrease which occurred in parallel to marked changes in cell shape, cell process outgrowth, and the organization and accumulation of gliofilaments. During the same culture period (7-32 days), alpha-tubulin mRNA, which was used as an internal standard, did not vary significantly. These results show that the increase of the GFAP protein and of gliofilaments observed both in vivo and in vitro during astroglial differentiation cannot be ascribed to an accumulation of the GFAP-mRNA. It might be that more than one mechanism regulates the levels of free and polymerized GFAP and of its encoding mRNA.

Animals↗
PubMed2006

Effects of myasthenia gravis patient sera on human myoblast cultures.

OBJECTIVES: To investigate the role of acetylcholine receptor (AChR) and other anti-muscle autoantibodies in myasthenia gravis (MG). Since many of these autoantibodies target proteins with structural or signalling functions, we examined the effect of MG sera on muscle cell morphology. MATERIALS AND METHODS: Primary human myoblast cultures were exposed to MG sera and morphological changes observed by light and fluorescence microscopy. RESULTS: MG patient sera caused changes in cell shape (cell retraction) and led to the formation of inclusion bodies and intracellular vesicles. A disordered arrangement of actin microfilaments was also observed. The effects were not complement-mediated, were both dose- and time-dependent, and appeared to correlate with disease severity of the MG donor. CONCLUSION: The factors responsible for these effects in vitro may also play a role in the pathogenesis of MG in vivo. Further study of these factors may improve our understanding of MG pathogenesis.

Apoptosis↗
PubMed1983

Straight mutants of Spirillum volutans can swim.

Nonhelical mutant cells of Spirillum volutans ATCC 19554 can swim as fast as the helical cells. Consequently, a helical cell shape is not required for motility of this species, and the function of the polar flagellar fascicles is not merely to cause rotation, and therefore translocation, of the corkscrew-shaped cell.

Movement↗
PubMed2006

Dynamic filaments of the bacterial cytoskeleton.

Bacterial cells contain a variety of structural filamentous proteins necessary for the spatial regulation of cell shape, cell division, and chromosome segregation, analogous to the eukaryotic cytoskeletal proteins. The molecular mechanisms by which these proteins function are beginning to be revealed, and these proteins show numerous three-dimensional structural features and biochemical properties similar to those of eukaryotic actin and tubulin, revealing their evolutionary relationship. Recent technological advances have illuminated links between cell division and chromosome segregation, suggesting a higher complexity and organization of the bacterial cell than was previously thought.

Actins↗
PubMed2000

Substrate flexibility regulates growth and apoptosis of normal but not transformed cells.

One of the hallmarks of oncogenic transformation is anchorage-independent growth (27). Here we demonstrate that responses to substrate rigidity play a major role in distinguishing the growth behavior of normal cells from that of transformed cells. We cultured normal or H-ras-transformed NIH 3T3 cells on flexible collagen-coated polyacrylamide substrates with similar chemical properties but different rigidity. Compared with cells cultured on stiff substrates, nontransformed cells on flexible substrates showed a decrease in the rate of DNA synthesis and an increase in the rate of apoptosis. These responses on flexible substrates are coupled to decreases in cell spreading area and traction forces. In contrast, transformed cells maintained their growth and apoptotic characteristics regardless of substrate flexibility. The responses in cell spreading area and traction forces to substrate flexibility were similarly diminished. Our results suggest that normal cells are capable of probing substrate rigidity and that proper mechanical feedback is required for regulating cell shape, cell growth, and survival. The loss of this response can explain the unregulated growth of transformed cells.

3T3 Cells↗
PubMed2004

Overexpression of hyperactive integrin-linked kinase leads to increased cellular radiosensitivity.

Integrin-linked kinase (ILK), bound to the cytoplasmic tails of integrin beta1, beta2, and beta3, is thought to signal through AKT and glycogen synthase kinase-3beta (GSK-3beta) for survival and proliferation regulation. To determine the role of ILK in the cellular radiation response, stably transfected A549 lung cancer cells overexpressing either wild-type (ILK-wk) or hyperactive ILK (ILK-hk) were studied for survival, signaling, proliferation, and examined in immunofluorescence and adhesion assays. Strong radiosensitization was observed in ILK-hk in contrast to ILK-wk mutants and empty vector controls. ILK small interfering RNA transfections showed radioresistance similar to irradiation on fibronectin. AKT, GSK-3beta-cyclin D1, mitogen-activated protein kinase kinase 1/2-mitogen-activated protein kinase, and c-Jun NH2-terminal kinase signaling was dysregulated in irradiated ILK-hk mutants. Immunofluorescence stainings of ILK-hk cells indicated disturbed ILK and paxillin membrane localization with concomitant decrease in focal adhesions. Profound ILK-hk-dependent changes in morphology were characterized by spindle-like cell shape, cell size reduction, increased cell protrusions, strong formation of membranous f-actin rings, and significantly reduced adhesion to matrix proteins. Additionally, ILK-wk and ILK-hk overexpression impaired beta1-integrin clustering and protein Tyr-phosphorylation. Taken together, the data provide evidence that ILK signaling modulates the cellular radiation response involving diverse signaling pathways and through changes in f-actin-based processes such as focal adhesion formation, cell adhesion, and spreading. Identification of ILK and its signaling partners as potential targets for tumor radiosensitization might promote innovative anticancer strategies by providing insight into the mechanism of cell adhesion-mediated radioresistance, oncogenic transformation, and tumor growth and spread.

Actins↗
PubMed1997

Time scale dependent viscoelastic and contractile regimes in fibroblasts probed by microplate manipulation.

Many essential phenomena in biology involve changes in cell shape. Cell deformation occurs in response to physical forces either coming from the external environment or intracellularly generated. In most tests of cell rheology, an external constraint is usually superimposed on an already mechanically active cell, thus the measurements may reflect both active motion and passive viscoelastic deformation. To show that active and passive processes could be distinguished on a time scale basis, we designed a novel piezo-controlled micromanipulation system to impose dynamic mechanical deformations on individual cells. Chick fibroblasts were seized between two glass microplates; one of the plates, more flexible, served as a sensor of the applied force. Controlled amounts of unidirectional compression and traction in the range of 10(-8)-10(-7) N were applied, using either step functions or sinusoidal signals at chosen frequencies. These tests allowed identification of three time scale dependent regimes. (1) A dominant elastic response, characterized by a linear stress-strain relationship, was especially apparent at short times (seconds); (2) A viscous behavior, characterized by force relaxation and irreversible cell deformation, was noticeable at intermediate times (minutes). Data from traction and oscillatory excitation tests were well fitted by a three-element Kelvin viscoelastic model, allowing the calculation of two elastic moduli in the range of 600-1,000 N/m2 and an apparent viscosity of about 10(4) Pa.s. (3) A contractile regime, in which actin-dependent traction forces were developed in response to uniaxial load was apparent at longer times (several tens of minutes). These forces were in the order of 4 x 10(-8) N above viscous relaxation. Thus we could distinguish, on a time scale basis, the specific contributions of passive viscoelasticity and active traction, and evaluate their mechanical characteristics within one experiment on a single cell.

Animals↗
PubMed1994

[Structure and function of mammalian brain microtubule-associated proteins].

Cytoplasmic microtubules are fibrous intracellular organelles found in almost all eukaryotic cells and play an important role in maintenance of cell shape, cell division, axonal transport, secretion and receptor activity. Besides tubulin dimers, microtubule proteins consist of several other components called MAPs which promote microtubule assembly and form long filamentous projection on the surface of the polymer. In mammalian brain, two classes of MAPs have been characterized; one is structural MAPs including MAP1 (1A and 1B), MAP2 (2A, 2B and 2C) and tau which function in the morphogenesis and maintenance of neural tissues and cells, and the other contains motor MAPs (kinesin and MAP1C) which are related to translocation of vesicles along microtubules in axon and to mitosis. The primary sequences of MAPs have been determined from their cDNAs. The functions of structural MAPs are modulated by their binding to other intracellular components, different expressions of isoforms during brain development and phosphorylation-dephosphorylation by various protein kinases and phosphatases. Biochemical characterization of MAP2 and tau have been well investigated. However, little is known about the function of MAP1 under the biochemical level, because MAP1 is unstable and high sensitive to proteases. We have developed a simple and rapid purification procedure for MAP1 using poly (L-aspartic acid) and taxol, and observed MAP1-F-actin interaction as well as MAP1-microtubules interaction. Recently, we have found that three specific kinases which can phosphorylate MAP1A and 1B are associated with MAP1 preparation and called it MAP1 kinase. Some evidence suggest that one of them is an unknown kinase and others are casein kinase I- and II-like kinases. Further studies to examine MAP1 kinase and phosphorylation of MAP1 provide a valuable insight for understanding thoroughly the microtubule-mediated functions.

Animals↗
PubMed2001

[Growth and metabolism of human skin fibroblasts cultured on microcarriers].

Human foreskin fibroblasts were cultured in vitro on microcarriers in spinners and traditional static flasks. The cultured cells obtained with these approaches were compared in cell shape, cell growth, cell production, and the metabolisms of glucose, lactate, and ammonium. The cells from microcarrier cultures through medium exchange were 8 times more than those from traditional static flasks. It was found that the specific growth rate and specific glucose consumption rate in microcarrier spinner cultures were 0.64/d-1 and 5.56 mmol/10(9) cell/d, respectively, higher than those in static flask cultures. However, the average lactate yield on glucose consumption in spinner cultures was only 0.955 mmol/mmol, lower than that in static flask cultures, 1. 125 mmol/mmol. This indicated that the energy metabolism in spinner cultures was significantly more efficient than that in static flasks. The experimental results from this work suggest that the microcarrier culture system is a suitable way to expand the seeding cells for tissue engineering, due to its ideal cultivation environment provided.

Cell Division↗
PubMed2004

[Impact of radiofrequency/microwave radiation on cell and cytoskeleton structure].

The increasing number of mobile phone users raises concern about possible hazardous effects of radiofrequency/microwave (RF/MW) electromagnetic fields. The most intriguing issue is the existence of the so called non-thermal effect which is not related to increased temperature. Possible biological effects, associated with the exposure to RF/MW fields have not been elucidated so far. A number of in vitro studies have attempted to see whether permissible levels of RF/MW affect the cell membrane, gene transcription, apoptosis, and whether they induce stress. However, the effect of RF/MW on the cytoskeleton, one of the most important cell structures, remains unknown. Cytoskeleton is a functional and structural part of the cell, composed of actin, intermedial filaments and microtubules. Microtubules direct the distribution of cellular organelles, determine cell shape, cell polarity and segregation of chromosomes during mitosis.

Animals↗
PubMed2005

Beta-actin in human colon adenocarcinoma cell lines with different metastatic potential.

Human colon adenocarcinoma LS180 parental cell line and selected variants, characterized by different metastatic capacity were used to examine, whether a correlation exists between beta-actin expression, its subcellular distribution and metastatic potential of these cells. Cytosolic fraction (supernatant 105000 x g), isolated from the tumor cells was used as a source for actin quantification. The higher level of beta-actin was observed in the cytosol of three selected sublines to compare with LS180 parental line. Statistically significant increase of beta-actin level in highly motile EB3 cells variant should be underlined to compare with the other sublines. Distinct differences in the phenotype of adenocarcinoma cell variants were found, such as the changes in cells shape, cells spreading and ability to attach to the surface of culture dish. Actin cytoskeleton was visualized with fluorescence microscopy application and microfilaments rhodamine-conjugated phalloidin staining. beta-actin subcellular localization was done by immunofluorescence staining with monoclonal anti-beta actin antibodies. In the elongated cells (LS180, 3LNLN), this isoactin is dispersed in the whole cell body and concentrates in pseudopods and at the leading edges, when in the rounded variant (EB3) beta-actin dominates mainly in cortical ring under cellular membrane and it is also seen in the subtle protrusions. Summary of our former (Nowak et al., 2002, Acta Biochim. Polon., 49: 823) and current data lead to the conclusion that there is a distinct correlation between metastatic capacity of examined human colon adenocarcinoma cells, the state of actin polymerization, actin cytoskeleton organization and beta-actin expression.

Actins↗
PubMed1985

Correlation of frequency of induced mutation and metastatic potential in tumor cell lines from a single mouse mammary tumor.

Spontaneous mutation rates were determined in mouse mammary tumor subpopulation lines that differ in metastatic phenotype. Although there was almost a 9-fold difference in spontaneous rates to ouabain resistance among the three lines tested, the difference did not correlate with ability to metastasize. Similarly a 10-fold difference in spontaneous rates to 6-thioguanine resistance did not correlate with metastatic ability. In contrast, the frequency of ethyl methanesulfonate-induced mutations was associated with metastatic potential. Thus, ethyl methanesulfonate only induced significant numbers of 6-thioguanine resistant colonies in 66 and 410.4 cells, the only 2 of 5 lines tested that spontaneously metastasize at high frequency, and of ouabain resistant colonies in 66, 410.4, and 168 cells, the only lines tested that produce experimental lung metastases after i.v. injection. Differential sensitivity to induced mutation was not correlated with differences in plating efficiency, wild type sensitivity to ethyl methanesulfonate, 6-thioguanine, or ouabain toxicity, ploidy, cell shape, cell size, or ability to engage in metabolic cooperation.

Animals↗
PubMed1984

Microtubules influence compaction in preimplantation mouse embryos.

The role of microtubules during compaction of the 8-cell-stage mouse embryo was investigated using the drugs Taxol (which leads to a non-controlled polymerization of tubulin) and Nocodazole (which causes depolymerization of microtubules). Taxol inhibits compaction in most non-compacted embryos and reverses it in already compacted embryos. These effects were observed on both cell flattening (as judged by phase-contrast microscopy) and on cell surface polarization (as judged by scanning electron microscopy and the surface binding of fluorescent concanavalin A). In contrast Nocodazole does not inhibit cell flattening, but rather accelerates its completion. Nocodazole influences the detailed organization of the surface pole and appears to reduce the incidence of surface polarization but does not reverse polarity once established to a significant extent. We conclude that microtubules exercise a constraining role during compaction, influencing cell shape, cell organization and the time at which compaction takes place.

Alkaloids↗