The Regulation of Agonist-Stimulated Phospholipase D Activity in Swiss 3T3 Fibroblasts

1993 ◽  
Vol 21 (4) ◽  
pp. 492S-492S
Author(s):  
CELIA. P. BRISCOE ◽  
MICHAEL. J. O. WAKELAM
Keyword(s):  
Phospholipase D ◽  
3T3 Fibroblasts ◽  
Swiss 3T3 ◽  
Swiss 3T3 Fibroblasts

scholarly journals Multiple sources of sn-1,2-diacylglycerol in platelet-derived-growth-factor-stimulated Swiss 3T3 fibroblasts. Evidence for activation of phosphoinositidase C and phosphatidylcholine-specific phospholipase D

1991 ◽  
Vol 279 (2) ◽  
pp. 559-565 ◽  
Author(s):  
R Plevin ◽  
S J Cook ◽  
S Palmer ◽  
M J O Wakelam
Keyword(s):  
Growth Factor ◽  
Phospholipase D ◽  
Lag Time ◽  
Water Soluble ◽  
Platelet Derived Growth Factor ◽  
Second Phase ◽  
Multiple Sources ◽  
3T3 Fibroblasts ◽  
Swiss 3T3 ◽  
Swiss 3T3 Fibroblasts

Platelet-derived growth factor (PDGF) stimulated sn-1,2-diacylglycerol (DAG) mass formation in Swiss 3T3 fibroblasts with a lag time of some 30 s. The response was biphasic, with the second phase being sustained over time. PDGF also stimulated the formation of Ins(1,4,5)P3 with a similar lag time to the DAG response, suggesting that DAG is derived from PtdIns(4,5)P2 hydrolysis at this time point. PDGF-stimulated phosphatidylcholine (PtdCho) hydrolysis in Swiss 3T3 fibroblasts, as measured by the formation of water-soluble choline metabolites and phosphatidylbutanol (PtdBut) accumulation, was by a phospholipase D (PLD)-catalysed pathway which was kinetically downstream of initial PtdIns(4,5)P2 hydrolysis. Accumulation of PtdBut increased up to 15 min, suggesting that PLD activity is not rapidly densitized in response to PDGF. The kinetics of PtdCho hydrolysis closely paralleled the second phase of DAG formation, strongly suggesting that during prolonged stimulation periods PtdCho is a major source of DAG in these cells. However, since PtdIns(4,5)P2 breakdown was also prolonged, PDGF-stimulated DAG may be derived from both phospholipids. Down-regulation of protein kinase C (PKC), by pre-treatment with phorbol 12-myristate 13-acetate, abolished both [3H]choline and [3H]PtdBut formation, suggesting that PLD-catalysed PtdCho hydrolysis may be dependent on PKC activation, supporting its dependence on prior PtdIns(4,5)P2 hydrolysis.

scholarly journals The roles of multiple pathways in regulating bombesin-stimulated phospholipase D activity in Swiss 3T3 fibroblasts

1995 ◽  
Vol 306 (1) ◽  
pp. 115-122 ◽  
Author(s):  
C P Briscoe ◽  
A Martin ◽  
M Cross ◽  
M J O Wakelam
Keyword(s):  
Tyrosine Phosphorylation ◽  
Phospholipase D ◽  
Tyrosine Kinases ◽  
Kinase Inhibitor ◽  
Protein Tyrosine ◽  
Permeabilized Cells ◽  
3T3 Fibroblasts ◽  
Swiss 3T3 ◽  
Swiss 3T3 Fibroblasts ◽  
Stimulation Of

The regulation of bombesin-stimulated phospholipase D (PLD) activity in Swiss 3T3 fibroblasts was examined. Increasing protein-tyrosine phosphorylation by using pervanadate to inhibit tyrosine phosphatases was found to stimulate protein kinase C (PKC)-independent [3H]phosphatidylbutanol ([3H]PtdBut) accumulation within 5 min, which continued to increase up to 30 min. The stimulation of PLD activity in response to submaximal [bombesin] could be decreased by approx. 50% by the tyrosine kinase inhibitor genistein, whereas pretreatment with genistein and the PKC inhibitor Ro-31-8220 completely abolished the generation of [3H]PtdBut in response to a maximal concentration of bombesin. The addition of guanosine 5′-[gamma-thio]triphosphate (GTP[S]) into permeabilized cells resulted in an increase in [3H]PtdBut, which was abolished by depletion of cellular ATP. The additional presence of 30 microM GTP[S] did not increase the stimulation of PLD activity by any [bombesin] tested, whereas it was synergistic with that stimulated in response to phorbol 12-myristate 13-acetate. These findings suggest that bombesin-stimulated PLD activity is indirectly regulated by G-proteins, possibly through a kinase intermediate. Furthermore, activation of protein tyrosine kinases is proposed to account for the PKC-independent arm of bombesin-stimulated PLD activity. No evidence was obtained for a form of PLD directly regulated by tyrosine phosphorylation.

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.

cDNA-cloning, sequencing and expression in glucocorticoid-stimulated quiescent Swiss 3T3 fibroblasts of mouse lipocortin I

1989 ◽  
Vol 159 (1) ◽  
pp. 155-162 ◽  
Author(s):  
Christine Philipps ◽  
Stefan Rose-John ◽  
Gabriele Rincke ◽  
Gerhard Fürstenberger ◽  
Friedrich Marks
Keyword(s):  
Cdna Cloning ◽  
3T3 Fibroblasts ◽  
Swiss 3T3 ◽  
Sequencing And Expression ◽  
Swiss 3T3 Fibroblasts
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