scholarly journals Uncovering a Role for the Tail of the Dictyostelium discoideum SadA Protein in Cell-Substrate Adhesion

2011 ◽  
Vol 10 (5) ◽  
pp. 662-671 ◽  
Author(s):  
Anthony S. Kowal ◽  
Rex L. Chisholm
Keyword(s):  
Actin Cytoskeleton ◽  
Dictyostelium Discoideum ◽  
Cytoplasmic Tail ◽  
Tail Domain ◽  
Content Type ◽  
Substrate Adhesion ◽  
Cell Substrate ◽  
Phosphorylation Event ◽  
Carboxy Terminal ◽  
Cell Substrate Adhesion

ABSTRACTPrevious work from our laboratory showed that theDictyostelium discoideumSadA protein plays a central role in cell-substrate adhesion. SadA null cells exhibit a loss of adhesion, a disrupted actin cytoskeleton, and a cytokinesis defect. How SadA mediates these phenotypes is unknown. This work addresses the mechanism of SadA function, demonstrating an important role for the C-terminal cytoplasmic tail in SadA function. We found that a SadA tailless mutant was unable to rescue thesadAadhesion deficiency, and overexpression of the SadA tail domain reduced adhesion in wild-type cells. We also show that SadA is closely associated with the actin cytoskeleton. Mutagenesis studies suggested that four serine residues in the tail, S924/S925 and S940/S941, may regulate association of SadA with the actin cytoskeleton. GlutathioneS-transferase pull-down assays identified at least one likely interaction partner of the SadA tail, cortexillin I, a known actin bundling protein. Thus, our data demonstrate an important role for the carboxy-terminal cytoplasmic tail in SadA function and strongly suggest that a phosphorylation event in this tail regulates an interaction with cortexillin I. Based on our data, we propose a model for the function of SadA.

scholarly journals Paxillin and Phospholipase D Interact To Regulate Actin-Based Processes in Dictyostelium discoideum

2011 ◽  
Vol 10 (7) ◽  
pp. 977-984 ◽  
Author(s):  
Jelena Pribic ◽  
Rebecca Garcia ◽  
May Kong ◽  
Derrick Brazill
Keyword(s):  
Dictyostelium Discoideum ◽  
Phospholipase D ◽  
Actin Polymerization ◽  
Genetic Interaction ◽  
Actin Reorganization ◽  
Content Type ◽  
Multicellular Development ◽  
Substrate Adhesion ◽  
Cell Substrate ◽  
Cell Substrate Adhesion

ABSTRACT The actin cytoskeleton forms a membrane-associated network whose proper regulation is essential for numerous processes, including cell differentiation, proliferation, adhesion, chemotaxis, endocytosis, exocytosis, and multicellular development. In this report, we show that in Dictyostelium discoideum , paxillin (PaxB) and phospholipase D (PldB) colocalize and coimmunoprecipitate, suggesting that they interact physically. Additionally, the phenotypes observed during development, cell sorting, and several actin-required processes, including cyclic AMP (cAMP) chemotaxis, cell-substrate adhesion, actin polymerization, phagocytosis, and exocytosis, reveal a genetic interaction between paxB and pldB , suggesting a functional interaction between their gene products. Taken together, our data point to PldB being a required binding partner of PaxB during processes involving actin reorganization.

Functional and cytoskeletal changes induced by sublethal injury in proximal tubular epithelial cells

1994 ◽  
Vol 266 (1) ◽  
pp. F21-F30 ◽  
Author(s):  
V. M. Kroshian ◽  
A. M. Sheridan ◽  
W. Lieberthal
Keyword(s):  
Actin Cytoskeleton ◽  
Cell Injury ◽  
Single Cells ◽  
Atp Depletion ◽  
Substrate Adhesion ◽  
Cell Substrate ◽  
Sublethal Injury ◽  
Proximal Tubular ◽  
Cell Substrate Adhesion ◽  
Chemical Anoxia

Mouse proximal tubular (MPT) cells in culture were subjected to ATP depletion by incubating them with cyanide in the absence of dextrose for 1 h. This insult resulted in marked alterations in the actin cytoskeleton. These changes were not associated with a decrease in cell viability and thus reflected sublethal injury. The effect of sublethal injury on the functional integrity of the intercellular tight junction (TJ) was then examined in MPT cell monolayers grown on permeable supports. During chemical anoxia, monolayer permeability to the paracellular marker mannitol progressively increased to 297 +/- 62% of baseline after 1 h. Chemical anoxia also caused a reversible loss in cell-substrate adhesion when MPT cells were studied as confluent monolayers or as single cells. Thus disruption of the actin cytoskeleton in nonlethally injured cells results in important reversible alterations in renal epithelial function characterized by impairment of the “gate” function of the TJ as well as impaired cell-substrate adhesion. We hypothesize that sublethal epithelial cell injury without accompanying necrosis may contribute to the decrement in renal function characteristic of ischemic renal injury.

scholarly journals Glycoprotein gp130 ofDictyostelium discoideumInfluences Macropinocytosis and Adhesion

2005 ◽  
Vol 16 (6) ◽  
pp. 2681-2693 ◽  
Author(s):  
Catherine P. Chia ◽  
Sujatha Gomathinayagam ◽  
Robert J. Schmaltz ◽  
Laura K. Smoyer
Keyword(s):  
Plasma Membrane ◽  
Amino Acid ◽  
Recombinant Protein ◽  
Dictyostelium Discoideum ◽  
Substrate Adhesion ◽  
Cell Substrate ◽  
Protein Gels ◽  
The Difference ◽  
Cell Substrate Adhesion ◽  
Axenic Growth

Glycoprotein gp130, found on the plasma membrane of Dictyostelium discoideum amoebae, was postulated previously to play a role in phagocytosis. The gene for gp130 was cloned and when translated, yielded a 768 amino acid preproprotein of 85.3 kDa. It had nearly 40% similarity to the 138 kDa family of glycoproteins implicated in sexual cell fusion during macrocyst formation in D. discoideum. The difference between the calculated size and observed Mrof 130 kDa on protein gels likely was due to N-glycosylation that was confirmed by lectin blots. Consistent with its surface-exposure, an antibody raised against recombinant protein stained the plasma membrane of D. discoideum amoebae. Gp130 and its transcripts were high during axenic growth of cells, but relatively low during growth on bacteria. The gene for gp130 was disrupted and cell lines lacking the glycoprotein were efficient phagocytes, indicating that gp130 was dispensable for phagocytosis. Gp130-null cells were similar in size to parent DH1 cells, had enhanced macropinocytosis and grew faster to higher densities. They also exhibited weaker cell-substrate adhesion but displayed greater cell-cell cohesion. Collectively, the data indicated that gp130 influenced macropinocytosis and played a role in adhesion during vegetative growth.

scholarly journals A New Member of the Rho Family, Rnd1, Promotes Disassembly of Actin Filament Structures and Loss of Cell Adhesion

1998 ◽  
Vol 141 (1) ◽  
pp. 187-197 ◽  
Author(s):  
Catherine D. Nobes ◽  
Inger Lauritzen ◽  
Marie-Geneviève Mattei ◽  
Sonia Paris ◽  
Alan Hall ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Actin Cytoskeleton ◽  
Rho Gtpase ◽  
Focal Adhesions ◽  
Substrate Adhesion ◽  
Cell Substrate ◽  
Rho Family ◽  
Cell Substrate Adhesion ◽  
Distinct Branch

Members of the Rho GTPase family regulate the organization of the actin cytoskeleton in response to extracellular growth factors. We have identified three proteins that form a distinct branch of the Rho family: Rnd1, expressed mostly in brain and liver; Rnd2, highly expressed in testis; and Rnd3/RhoE, showing a ubiquitous low expression. At the subcellular level, Rnd1 is concentrated at adherens junctions both in confluent fibroblasts and in epithelial cells. Rnd1 has a low affinity for GDP and spontaneously exchanges nucleotide rapidly in a physiological buffer. Furthermore, Rnd1 lacks intrinsic GTPase activity suggesting that in vivo, it might be constitutively in a GTP-bound form. Expression of Rnd1 or Rnd3/RhoE in fibroblasts inhibits the formation of actin stress fibers, membrane ruffles, and integrin-based focal adhesions and induces loss of cell–substrate adhesion leading to cell rounding (hence Rnd for “round”). We suggest that these proteins control rearrangements of the actin cytoskeleton and changes in cell adhesion.

scholarly journals Pharmacological Activation of Rap1 Antagonizes the Endothelial Barrier Disruption Induced by Exotoxins ExoS and ExoT of Pseudomonas aeruginosa

2015 ◽  
Vol 83 (5) ◽  
pp. 1820-1829 ◽  
Author(s):  
Stéphanie Bouillot ◽  
Ina Attrée ◽  
Philippe Huber
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Vascular Wall ◽  
Content Type ◽  
Clinical Strains ◽  
Cell Monolayers ◽  
Substrate Adhesion ◽  
Cell Substrate ◽  
Type 3 Secretion System ◽  
Cell Substrate Adhesion ◽  
Type 3

Most clinical strains ofPseudomonas aeruginosa, a leading agent of nosocomial infections, are multiresistant to antibiotherapy. Because of the paucity of new available antibiotics, the investigation of strategies aimed at limiting the action of its major virulence factors has gained much interest. The type 3 secretion system ofP. aeruginosaand its effectors are known to be major determinants of toxicity and are required for bacterial dissemination in the host. Bacterial transmigration across the vascular wall is considered to be an important step in the infectious process. Using human endothelial primary cells, we demonstrate that forskolin (FSK), a drug inducing cyclic AMP (cAMP) elevation in eukaryotic cells, strikingly reduced the cell retraction provoked by two type 3 toxins, ExoS and ExoT, found in the majority of clinical strains. Conversely, cytotoxicity of a strain carrying the type 3 effector ExoU was unaffected by FSK. In addition, FSK altered the capacity of two ExoS/ExoT strains to transmigrate across cell monolayers. In agreement with these findings, other drugs and a cytokine inducing the increase of cAMP intracellular levels have also protected cells from retraction. cAMP is an activator of both protein kinase A and EPAC, a GTPase exchange factor of Rap1. Using activators or inhibitors of either pathway, we show that the beneficial effect of FSK is exerted by the activation of the EPAC/Rap1 axis, suggesting that its protective effect is mediated by reinforcing cell-cell and cell-substrate adhesion.

scholarly journals Regulation of cell-substrate adhesion by proteoglycans immobilized on extracellular substrates

1989 ◽  
Vol 264 (14) ◽  
pp. 8012-8018 ◽  
Author(s):  
M Yamagata ◽  
S Suzuki ◽  
S K Akiyama ◽  
K M Yamada ◽  
K Kimata
Keyword(s):  
Substrate Adhesion ◽  
Cell Substrate ◽  
Cell Substrate Adhesion

scholarly journals A GTPase controls cell-substrate adhesion in Xenopus XTC fibroblasts.

1992 ◽  
Vol 118 (5) ◽  
pp. 1235-1244 ◽  
Author(s):  
M H Symons ◽  
T J Mitchison
Keyword(s):  
Control Cell ◽  
Response To Treatment ◽  
Nucleotide Analogs ◽  
Cell Contractility ◽  
Cell Rounding ◽  
Substrate Adhesion ◽  
Cell Substrate ◽  
Adhesive Interactions ◽  
Cell Substrate Adhesion ◽  
Gamma S

Cell-substrate adhesion is crucial at various stages of development and for the maintenance of normal tissues. Little is known about the regulation of these adhesive interactions. To investigate the role of GTPases in the control of cell morphology and cell-substrate adhesion we have injected guanine nucleotide analogs into Xenopus XTC fibroblasts. Injection of GTP gamma S inhibited ruffling and increased spreading, suggesting an increase in adhesion. To further investigate this, we made use of GRGDSP, a peptide which inhibits binding of integrins to vitronectin and fibronectin. XTC fibroblasts injected with non-hydrolyzable analogs of GTP took much more time to round up than mock-injected cells in response to treatment with GRGDSP, while GDP beta S-injected cells rounded up in less time than controls. Injection with GTP gamma S did not inhibit cell rounding induced by trypsin however, showing that cell contractility is not significantly affected by the activation of GTPases. These data provide evidence for the existence of a GTPase which can control cell-substrate adhesion from the cytoplasm. Treatment of XTC fibroblasts with the phorbol ester 12-o-tetradecanoylphorbol-13-acetate reduced cell spreading and accelerated cell rounding in response to GRGDSP, which is essentially opposite to the effect exerted by non-hydrolyzable GTP analogs. These results suggest the existence of at least two distinct pathways controlling cell-substrate adhesion in XTC fibroblasts, one depending on a GTPase and another one involving protein kinase C.

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