An increase in intracellular free Ca2+ associated with serum-free growth stimulation of Swiss 3T3 fibroblasts by epidermal growth factor in the presence of bradykinin.

ABSTRACT The concentration of epidermal growth factor (EGF) in human mammary secretions was about 50 nmol/l for several weeks prepartum. It then fell to about 13 nmol/l within 4 days after parturition, in parallel with the decrease in protein concentration which is associated with the onset of lactation. In contrast, the concentration of EGF in urine samples from the same donors remained constant throughout this period. All the immunoreactive EGF in mammary secretions competed at the EGF receptors on Swiss mouse 3T3 fibroblasts. The stimulation of these cells by samples of mammary secretions was, however, much greater than that induced by EGF alone, indicating the presence of other factors which synergize with EGF. Gel filtration of mammary secretions revealed two major peaks of mitogenic activity, corresponding to EGF and a factor of higher molecular weight. The latter could synergize with added EGF, insulin or bombesin, and thus falls into a different functional class from any of these factors. J. Endocr. (1987) 112, 151–159

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Inhibition of mitogen-induced DNA synthesis by bafilomycin A1 in Swiss 3T3 fibroblasts

Biochemical Journal ◽

10.1042/bj3130065 ◽

1996 ◽

Vol 313 (1) ◽

pp. 65-70 ◽

Cited By ~ 22

Author(s):

Andrew J. SAURIN ◽

Jane HAMLETT ◽

Michael J. CLAGUE ◽

Stephen R. PENNINGTON

Keyword(s):

Growth Factor ◽

Dna Synthesis ◽

Kinase Activity ◽

Bafilomycin A1 ◽

Tyrosine Kinase Activity ◽

3T3 Fibroblasts ◽

Mitogenic Signalling ◽

Epidermal Growth ◽

Swiss 3T3 ◽

Swiss 3T3 Fibroblasts

Quiescent cells (in G0) can be stimulated to enter the cell cycle and proceed to DNA synthesis in S-phase by a wide range of growth factors and mitogens. Activation of cell-surface growth factor receptors with intrinsic protein tyrosine kinase activity initiates autophosphorylation of the receptors and subsequent activation of signal transduction cascades. After activation the receptors undergo ligand-induced internalization to endosomes, which become acidified by the action of a vacuolar H+-ATPase (V-ATPase). The extent to which vesicular acidification plays a role in mitogenic signalling by receptors with intrinsic tyrosine kinase activity remains unknown. Here we have shown that bafilomycin A1, a specific inhibitor of V-ATPase, inhibits endosome acidification and mitogen-induced DNA synthesis in Swiss 3T3 fibroblasts. Addition of bafilomycin A1 at successively later times during G1 progressively decreased the inhibition of DNA synthesis such that no inhibition was observed when bafilomycin A1 was added at the onset of S-phase. Bafilomycin A1 also induced a dramatic but reversible change in the morphology of Swiss 3T3 cells. However, the rapid activation of c-fos mRNA accumulation by epidermal growth factor and insulin was unaffected by bafilomycin A1. Together, the results suggest that activation of the V-ATPase plays an important role in the mitogenic signalling pathways that occur during the G1 phase of the cell cycle but is not required for the initial epidermal growth factor and insulin-evoked signalling events that lead to c-fos mRNA expression.

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Activation of Endogenous Thrombin Receptors Causes Clustering and Sensitization of Epidermal Growth Factor Receptors of Swiss 3t3 Cells without Transactivation

The Journal of Cell Biology ◽

10.1083/jcb.152.2.263 ◽

2001 ◽

Vol 152 (2) ◽

pp. 263-274 ◽

Cited By ~ 11

Author(s):

Michael F. Crouch ◽

Deborah A. Davy ◽

Francis S. Willard ◽

Leise A. Berven

Keyword(s):

Growth Factor ◽

Epidermal Growth Factor ◽

Egf Receptor ◽

Thrombin Receptor ◽

3T3 Cells ◽

Downstream Effectors ◽

Epidermal Growth ◽

Swiss 3T3 ◽

G Protein Coupled ◽

Stimulation Of

The G protein–coupled thrombin receptor can induce cellular responses in some systems by transactivating the epidermal growth factor (EGF) receptor. This is in part due to the stimulation of ectoproteases that generate EGF receptor ligands. We show here that this cannot account for the stimulation of proliferation or migration by thrombin of Swiss 3T3 cells. Thrombin has no direct effect on the activation state of the EGF receptor or of its downstream effectors. However, thrombin induces the subcellular clustering of the EGF receptor at filamentous actin–containing structures at the leading edge and actin arcs of migrating cells in association with other signaling molecules, including Shc and phospholipase Cγ1. In these thrombin-primed cells, the subsequent migratory response to EGF is potentiated. Thrombin did not potentiate the EGF-stimulated EGF receptor phosphorylation. Thus, in Swiss 3T3 cells the G protein–coupled thrombin receptor can potentiate the EGF tyrosine kinase receptor response when activated by EGF, and this appears to be due to the subcellular concentration of the receptor with downstream effectors and not to the overall ability of EGF to induce receptor transphosphorylation. Thus, the EGF receptor subcellular localization which is altered by thrombin appears to be an important determinant of the efficacy of downstream EGF receptor signaling in cell migration.

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Growth stimulation of A431 cells by epidermal growth factor: identification of high-affinity receptors for epidermal growth factor by an anti-receptor monoclonal antibody.

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.80.5.1337 ◽

1983 ◽

Vol 80 (5) ◽

pp. 1337-1341 ◽

Cited By ~ 473

Author(s):

T. Kawamoto ◽

J. D. Sato ◽

A. Le ◽

J. Polikoff ◽

G. H. Sato ◽

...

Keyword(s):

Monoclonal Antibody ◽

Growth Factor ◽

Epidermal Growth Factor ◽

Growth Stimulation ◽

A431 Cells ◽

High Affinity ◽

Epidermal Growth ◽

Receptor Monoclonal Antibody ◽

Stimulation Of

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Stimulation of growth and migration of liver epithelial cells by insulin and epidermal growth factor in serum-free medium. immunohistological detection of migration tracks

Fresenius Zeitschrift für Analytische Chemie ◽

10.1007/bf00593894 ◽

1984 ◽

Vol 317 (6) ◽

pp. 741-742 ◽

Cited By ~ 2

Author(s):

E. G. Bade ◽

B. Nitzgen ◽

C. Lögl

Keyword(s):

Growth Factor ◽

Epidermal Growth Factor ◽

Epithelial Cells ◽

Serum Free Medium ◽

Free Medium ◽

Serum Free ◽

Liver Epithelial Cells ◽

Epidermal Growth ◽

And Migration ◽

Stimulation Of

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Calcineurin is essential for DNA synthesis in Swiss 3T3 fibroblasts

Biochemical Journal ◽

10.1042/bj3170675 ◽

1996 ◽

Vol 317 (3) ◽

pp. 675-680 ◽

Cited By ~ 24

Author(s):

Masahiro TOMONO ◽

Kotaro TOYOSHIMA ◽

Motohiro ITO ◽

Hisao AMANO

Keyword(s):

Growth Factors ◽

Growth Factor ◽

Dna Synthesis ◽

Early Stage ◽

Growth Stimulation ◽

3T3 Fibroblasts ◽

Protein Phosphatase 2B ◽

Dependent Protein ◽

Swiss 3T3 ◽

Swiss 3T3 Fibroblasts

DNA synthesis was measured 16 h after stimulation of Swiss 3T3 fibroblasts in the resting phase with various growth factors (platelet-derived growth factor, fibroblast growth factor, lysophosphatidic acid and thrombin). When extracellular Ca2+ was chelated by EGTA, or when the influx of Ca2+ from outside to inside the cell was blocked by cobalt, DNA synthesis was completely inhibited. As there was no effect whatsover on DNA synthesis when Ca2+ was chelated, or when the influx of Ca2+ was blocked up to the first 4 h after growth stimulation, it was concluded that, at an early stage, Ca2+ influx from outside to inside the cell is not related to the transition from the G1 to the S phase. A Ca2+/calmodulin-dependent protein kinase II inhibitor (KN-62) had no effect on DNA synthesis. However, cyclosporin A and FK-506, which are inhibitors of Ca2+/calmodulin-dependent protein phosphatase 2B (calcineurin), markedly inhibited DNA synthesis stimulated by all of the growth factors. These results indicate that calcineurin plays a role, not only in activation of T-cells of the immune system in the initial phase, but also in DNA synthesis in fibroblasts. It was concluded that Ca2+ influx from outside to inside the cell during the mid-to-late G1 phase, followed by calcineurin activation, is essential as a mechanism of growth signal transduction.

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