scholarly journals Physical association of the small GTPase Rho with a 68-kDa phosphatidylinositol 4-phosphate 5-kinase in Swiss 3T3 cells.

1996 ◽  
Vol 7 (3) ◽  
pp. 435-442 ◽  
Author(s):  
X D Ren ◽  
G M Bokoch ◽  
A Traynor-Kaplan ◽  
G H Jenkins ◽  
R A Anderson ◽  
...  
Keyword(s):  
Bound State ◽  
Mouse Fibroblast ◽  
Small Gtpase ◽  
Dominant Negative ◽  
Type I ◽  
3T3 Cells ◽  
Dominant Negative Inhibitor ◽  
Swiss 3T3 ◽  
Swiss 3T3 Cell ◽  
Phosphatidylinositol 4

Our previous work showed that post-translationally modified Rho in its GTP-bound state stimulated phosphatidylinositol 4-phosphate 5-kinase (PIP5K) activity in mouse fibroblast lysates. To investigate whether Rho physically interacts with PIP5K, we incubated immobilized Rho-GST with Swiss 3T3 cell lysates and tested for retained PIP5K activity. Rho-GST, but not Ras-GST or GST alone, bound significant PIP5K activity. The binding of PIP5K was independent of whether Rho was in a GTP- or GDP-bound state. An antibody against a 68-kDa human erythrocyte type I PIP5K recognized a single 68-kDa protein eluted from Rho-GST column. The Rho-associated PIP5K responded to phosphatidic acid differentially from the erythrocyte type I PIP5K, suggesting that it could be a distinct isoform not reported previously. Rho co-immunoprecipitated with the 68-kDa PIP5K from Swiss 3T3 lysates, demonstrating that endogenous Rho also interacts with PIP5K. ADP-ribosylation of Rho with C3 exoenzyme enhanced PIP5K binding by approximately eightfold, consistent with the ADP-ribosylated Rho functioning as a dominant negative inhibitor. These results demonstrate that Rho physically interacts with a 68-kDa PIP5K, although whether the association is direct or indirect is unknown.

scholarly journals ROCK and mDia1 antagonize in Rho-dependent Rac activation in Swiss 3T3 fibroblasts

2002 ◽  
Vol 157 (5) ◽  
pp. 819-830 ◽  
Author(s):  
Takahiro Tsuji ◽  
Toshimasa Ishizaki ◽  
Muneo Okamoto ◽  
Chiharu Higashida ◽  
Kazuhiro Kimura ◽  
...  
Keyword(s):  
Focal Adhesions ◽  
Small Gtpase ◽  
Dominant Negative ◽  
Stress Fibers ◽  
Induced Membrane ◽  
3T3 Fibroblasts ◽  
C3 Exoenzyme ◽  
Swiss 3T3 ◽  
Swiss 3T3 Fibroblasts ◽  
Membrane Ruffle

The small GTPase Rho acts on two effectors, ROCK and mDia1, and induces stress fibers and focal adhesions. However, how ROCK and mDia1 individually regulate signals and dynamics of these structures remains unknown. We stimulated serum-starved Swiss 3T3 fibroblasts with LPA and compared the effects of C3 exoenzyme, a Rho inhibitor, with those of Y-27632, a ROCK inhibitor. Y-27632 treatment suppressed LPA-induced formation of stress fibers and focal adhesions as did C3 exoenzyme but induced membrane ruffles and focal complexes, which were absent in the C3 exoenzyme-treated cells. This phenotype was suppressed by expression of N17Rac. Consistently, the amount of GTP-Rac increased significantly by Y-27632 in LPA-stimulated cells. Biochemically, Y-27632 suppressed tyrosine phosphorylation of paxillin and focal adhesion kinase and not that of Cas. Inhibition of Cas phosphorylation with PP1 or expression of a dominant negative Cas mutant inhibited Y-27632–induced membrane ruffle formation. Moreover, Crk-II mutants lacking in binding to either phosphorylated Cas or DOCK180 suppressed the Y-27632–induced membrane ruffle formation. Finally, expression of a dominant negative mDia1 mutant also inhibited the membrane ruffle formation by Y-27632. Thus, these results have revealed the Rho-dependent Rac activation signaling that is mediated by mDia1 through Cas phosphorylation and antagonized by the action of ROCK.

scholarly journals The Crk signaling pathway contributes to the bombesin-induced activation of the small GTPase Rap1 in Swiss 3T3 cells

Oncogene ◽  
2000 ◽  
Vol 19 (54) ◽  
pp. 6361-6368 ◽  
Author(s):  
Guido Posern ◽  
Ulf R Rapp ◽  
Stephan M Feller
Keyword(s):  
Signaling Pathway ◽  
Small Gtpase ◽  
3T3 Cells ◽  
Swiss 3T3

scholarly journals Production of plasminogen activator by established cell lines of mouse origin.

1977 ◽  
Vol 73 (1) ◽  
pp. 47-55 ◽  
Author(s):  
D B Rifkin ◽  
R Pollack
Keyword(s):  
Plasminogen Activator ◽  
Cell Lines ◽  
3T3 Cells ◽  
Established Cell Lines ◽  
Embryo Cultures ◽  
Established Cell ◽  
Swiss 3T3 ◽  
Swiss 3T3 Cell ◽  
Transformed Cell Lines ◽  
3T3 Cell Lines

The correlation between malignant transformation and increased plasminogen activator synthesis has been studied in a variety of established cell lines. In contrast to the behavior of secondary mouse embryo cultures, which always show increased fibrinolytic activity when transformed, no such correlation was found within the BALB/c 3T3 line and its transformed derivatives. Cell lines were established from tumors initiated in BALB/c mice by several transformed cell lines. These lines were generally found to contain no more plasminogen activator than the cells used for inoculation. A correlation was found between transformation and plasminogen activator synthesis within Swiss 3T3 cell lines. However, the correlation was not maintained by serum revertants of transformed Swiss 3T3 cells.

scholarly journals Analysis of the fibroblast transformation potential of GTPase-deficient gip2 oncogenes.

1992 ◽  
Vol 12 (1) ◽  
pp. 190-197 ◽  
Author(s):  
S K Gupta ◽  
C Gallego ◽  
J M Lowndes ◽  
C M Pleiman ◽  
C Sable ◽  
...  
Keyword(s):  
Growth Regulation ◽  
Normal Growth ◽  
Soft Agar ◽  
Dominant Negative ◽  
3T3 Cells ◽  
3T3 Fibroblasts ◽  
Signal Transduction Protein ◽  
Swiss 3T3 ◽  
Nih 3T3 ◽  
Mutant Polypeptides

Expression of GTPase-deficient Gi2 alpha subunit (alpha i2) mutant polypeptides and overexpression of the wild-type alpha i2 polypeptide in Rat 1a, Swiss 3T3, and NIH 3T3 fibroblasts altered normal growth regulation and induced a loss of contact inhibition. In Rat 1a cells (but not in NIH 3T3 or Swiss 3T3 cells), expression of the GTPase-deficient alpha i2 mutant polypeptides allowed colony formation in soft agar, which correlated with a loss in anchorage dependence and a decreased serum requirement. The altered growth regulatory properties of Rat 1a cells induced by expression of alpha i2 mutant polypeptides was not significantly inhibited by cotransfection with a dominant negative Ha-ras mutant polypeptide (Asn-17rasH), indicating that the activated Gi2 membrane signal transduction protein is uniquely capable of altering the regulation of Rat 1a cell growth by a predominantly c-ras-independent mechanism. The results show that GTPase-deficient alpha i2 mutant polypeptides have the properties of an oncogene that can induce the phenotypic characteristics of transformation in Rat 1a cells but that only a subset of these changes is observed with NIH 3T3 and Swiss 3T3 cells.

The promotion of hepatic maturation of human pluripotent stem cells in 3D co-culture using type I collagen and Swiss 3T3 cell sheets

Biomaterials ◽  
2012 ◽  
Vol 33 (18) ◽  
pp. 4526-4534 ◽  
Author(s):  
Yasuhito Nagamoto ◽  
Katsuhisa Tashiro ◽  
Kazuo Takayama ◽  
Kazuo Ohashi ◽  
Kenji Kawabata ◽  
...  
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Type I Collagen ◽  
Human Pluripotent Stem Cells ◽  
Type I ◽  
Cell Sheets ◽  
Swiss 3T3 ◽  
Swiss 3T3 Cell

Analysis of the fibroblast transformation potential of GTPase-deficient gip2 oncogenes

1992 ◽  
Vol 12 (1) ◽  
pp. 190-197
Author(s):  
S K Gupta ◽  
C Gallego ◽  
J M Lowndes ◽  
C M Pleiman ◽  
C Sable ◽  
...  
Keyword(s):  
Growth Regulation ◽  
Normal Growth ◽  
Soft Agar ◽  
Dominant Negative ◽  
3T3 Cells ◽  
3T3 Fibroblasts ◽  
Signal Transduction Protein ◽  
Swiss 3T3 ◽  
Nih 3T3 ◽  
Mutant Polypeptides

Expression of GTPase-deficient Gi2 alpha subunit (alpha i2) mutant polypeptides and overexpression of the wild-type alpha i2 polypeptide in Rat 1a, Swiss 3T3, and NIH 3T3 fibroblasts altered normal growth regulation and induced a loss of contact inhibition. In Rat 1a cells (but not in NIH 3T3 or Swiss 3T3 cells), expression of the GTPase-deficient alpha i2 mutant polypeptides allowed colony formation in soft agar, which correlated with a loss in anchorage dependence and a decreased serum requirement. The altered growth regulatory properties of Rat 1a cells induced by expression of alpha i2 mutant polypeptides was not significantly inhibited by cotransfection with a dominant negative Ha-ras mutant polypeptide (Asn-17rasH), indicating that the activated Gi2 membrane signal transduction protein is uniquely capable of altering the regulation of Rat 1a cell growth by a predominantly c-ras-independent mechanism. The results show that GTPase-deficient alpha i2 mutant polypeptides have the properties of an oncogene that can induce the phenotypic characteristics of transformation in Rat 1a cells but that only a subset of these changes is observed with NIH 3T3 and Swiss 3T3 cells.

Syndecan-4 and integrins: combinatorial signaling in cell adhesion

1999 ◽  
Vol 112 (20) ◽  
pp. 3415-3420 ◽  
Author(s):  
J.R. Couchman ◽  
A. Woods
Keyword(s):  
Extracellular Matrix ◽  
Heparan Sulfate ◽  
Focal Adhesion ◽  
Core Protein ◽  
Adhesion Formation ◽  
Focal Adhesions ◽  
Dominant Negative ◽  
Focal Adhesion Formation ◽  
Dominant Negative Inhibitor ◽  
Phosphatidylinositol 4

It is now becoming clear that additional transmembrane components can modify integrin-mediated adhesion. Syndecan-4 is a transmembrane heparan sulfate proteoglycan whose external glycosaminoglycan chains can bind extracellular matrix ligands and whose core protein cytoplasmic domain can signal during adhesion. Two papers in this issue of JCS demonstrate, through transfection studies, that syndecan-4 plays roles in the formation of focal adhesions and stress fibers. Overexpression of syndecan-4 increases focal adhesion formation, whereas a partially truncated core protein that lacks the binding site for protein kinase C(α) and phosphatidylinositol 4, 5-bisphosphate acts as a dominant negative inhibitor of focal adhesion formation. Focal adhesion induction does not require interaction between heparan sulfate glycosaminoglycan and ligand but can occur when non-glycanated core protein is overexpressed; this suggests that oligomerization of syndecan-4 plays a major role in signaling from the extracellular matrix in adhesion.

scholarly journals Inositol trisphosphate formation and calcium mobilization in Swiss 3T3 cells in response to platelet-derived growth factor

1984 ◽  
Vol 222 (1) ◽  
pp. 195-201 ◽  
Author(s):  
M J Berridge ◽  
J P Heslop ◽  
R F Irvine ◽  
K D Brown
Keyword(s):  
Growth Factor ◽  
Second Messengers ◽  
Platelet Derived Growth Factor ◽  
Maximal Effect ◽  
3T3 Cells ◽  
Intact Cells ◽  
Swiss 3T3 ◽  
Phosphatidylinositol 4 ◽  
Dose Dependent ◽  
Hydrolysis Of

Swiss 3T3 cells incubated for 60 h with [3H]inositol incorporated radioactivity into phosphatidylinositol (PI) and the two polyphosphoinositides phosphatidylinositol 4-phosphate (PIP) and phosphatidylinositol 4,5-bisphosphate (PIP2). On stimulation with platelet-derived growth factor (PDGF) there were significant increases in the levels of inositol 1-phosphate (IP1), inositol 1,4-bisphosphate (IP2) and inositol 1,4,5-trisphosphate (IP3). The effect of PDGF and IP3 on Ca2+ mobilization was studied in both intact cells and in ‘leaky’ cells that had been permeabilized with saponin. In intact cells, PDGF stimulated the efflux of 45Ca2+, whereas IP3 had no effect. Conversely, IP3 stimulated 45Ca2+ efflux from ‘leaky’ cells, which were insensitive to PDGF. ‘Leaky’ cells, which accumulated 45Ca2+ to a steady state within 20 min, were found to release approx. 40% of the label within 1 min after addition of 10 microM-IP3. This stimulation of 45Ca2+ release by IP3 was reversible and was also dose-dependent, with a half-maximal effect at approx. 0.3 microM. It seems likely that an important action of PDGF on Swiss 3T3 cells is to stimulate the hydrolysis of PIP2 to form IP3 and diacylglycerol, both of which may function as second messengers. Our results indicate that IP3 mobilizes intracellular Ca2+, and we propose that diacylglycerol may act through C-kinase to activate the Na+/H+ antiport. By generating two second messengers, PDGF can simultaneously elevate the intracellular level of Ca2+ and alkalinize the cytoplasm by lowering the level of H+.

Sign in / Sign up

Export Citation Format

Share Document