scholarly journals Cell transformation as aberrant differentiation: Environmentslly dependent spontaneous transformation of NIH 3T3 cells

Cell Research ◽  
1990 ◽  
Vol 1 (2) ◽  
pp. 197-206 ◽  
Author(s):  
Kang Xu ◽  
Harry Rubin
Keyword(s):  
Cell Transformation ◽  
3T3 Cells ◽  
Spontaneous Transformation ◽  
Nih 3T3 ◽  
Nih 3T3 Cells

Adaptation and selection as factors in the spontaneous transformation of NIH-3T3 cells

1992 ◽  
Vol 13 (10) ◽  
pp. 1873-1877 ◽  
Author(s):  
Ralph Grundel ◽  
Harry Rubin
Keyword(s):  
3T3 Cells ◽  
Spontaneous Transformation ◽  
Nih 3T3 ◽  
Nih 3T3 Cells

KSHV-GPCR and CXCR2 Transforming Capacity and Angiogenic Responses Are Mediated through a JAK2-STAT3 Dependent Pathway.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4347-4347
Author(s):  
Meike Burger ◽  
Tanja Hartmann ◽  
Jan A. Burger ◽  
Ingrid U. Schraufstatter
Keyword(s):  
Kaposi's Sarcoma ◽  
Cell Transformation ◽  
Kaposi’S Sarcoma ◽  
Fiber Formation ◽  
Actin Stress Fiber ◽  
3T3 Cells ◽  
Focus Formation ◽  
Nih 3T3 ◽  
Dependent Pathway ◽  
Nih 3T3 Cells

Abstract The Kaposi’s sarcoma herpesvirus encodes a G-protein-coupled chemokine receptor termed KSHV-GPCR. Expression of this constitutively active GPCR leads to cell transformation and vascular overgrowth characteristic of Kaposi’s sarcoma. Previously, we have shown that CXCR2, the closest human homologue, is similarly able to transform cells if continously stimulated or constitutively activated by amino acid exchange D138V of the DRY sequence. Here, we demonstrate that STAT3 activation is a prerequisite for transformation in KSHV-GPCR and CXCR2 transfected NIH 3T3 cells. In KSHV-GPCR and D138V transfected cells, STAT-3 is constitutively phosphorylated on Tyr705. In CXCR2 transfected NIH 3T3 cells and human microvascular endothelial cells (HMEC), which express the CXCR2 constitutively, STAT3 is phosphorylated upon stimulation with IL-8 (CXCL8). Focus formation in NIH 3T3 cells transfected with the KSHV-GPCR, CXCR2, or the D138V mutant, was blocked by the specific JAK2 inhibitor AG490. Typical functions of the CXCR2 including actin stress fiber formation, haptotaxis, and the angiogenic response in HMEC shown by tube formation in Matrigel were blocked by AG490. These data suggest that the transforming capacity and migratory responses that are involved in tumor development, metastasis and angiogenesis in KSHV or CXCR2-expressing cells is at least partially mediated through a JAK2-STAT3 dependent pathway.

Interactions of phosphatidylinositol kinase, GTPase-activating protein (GAP), and GAP-associated proteins with the colony-stimulating factor 1 receptor

1990 ◽  
Vol 10 (11) ◽  
pp. 5601-5608
Author(s):  
M Reedijk ◽  
X Q Liu ◽  
T Pawson
Keyword(s):  
Cell Transformation ◽  
Cellular Transformation ◽  
Colony Stimulating Factor ◽  
Gtpase Activating Protein ◽  
3T3 Cells ◽  
Nih 3T3 ◽  
Fibroblast Transformation ◽  
Insert Region ◽  
Stimulating Factor ◽  
Nih 3T3 Cells

The interactions of the macrophage colony-stimulating factor 1 (CSF-1) receptor with potential targets were investigated after ligand stimulation either of mouse macrophages or of fibroblasts that ectopically express mouse CSF-1 receptors. In Rat-2 cells expressing the mouse CSF-1 receptor, full activation of the receptor and cellular transformation require exogenous CSF-1, whereas NIH 3T3 cells expressing mouse c-fms are transformed by autocrine stimulation. Activated CSF-1 receptors physically associate with a phosphatidylinositol (PI) 3'-kinase. A mutant CSF-1 receptor with a deletion of the kinase insert region was deficient in its ability to bind functional PI 3'-kinase and to induce PI 3'-kinase activity precipitable with antiphosphotyrosine antibodies. In fibroblasts, CSF-1 stimulation also induced the phosphorylation of the GTPase-activating protein (GAP)-associated protein p62 on tyrosine, although GAP itself was a relatively poor substrate. In contrast to PI 3'-kinase association, phosphorylation of p62 and GAP was not markedly affected by deletion of the kinase insert region. These results indicate that the kinase insert region selectively enhances the CSF-1-dependent association of the CSF-1 receptor with active PI 3'-kinase. The insert deletion mutant retains considerable transforming activity in NIH 3T3 cells (G. Taylor, M. Reedijk, V. Rothwell, L. Rohrschneider, and T. Pawson, EMBO J. 8:2029-2037, 1989). This mutant was more seriously impaired in Rat-2 cell transformation, although mutant-expressing Rat-2 cells still formed small colonies in soft agar in the presence of CSF-1. Therefore, phosphorylation of GAP and p62 through activation of the CSF-1 receptor does not result in full fibroblast transformation. The interaction between the CSF-1 receptor and PI 3'-kinase may contribute to c-fms fibroblast transformation and play a role in CSF-1-stimulated macrophages.

scholarly journals Human c-fgr induces a monocyte-specific enzyme in NIH 3T3 cells.

1991 ◽  
Vol 11 (12) ◽  
pp. 6279-6285 ◽  
Author(s):  
K Inoue ◽  
B Wongsasant ◽  
T Akiyama ◽  
K Toyoshima
Keyword(s):  
Tyrosine Kinases ◽  
Cell Transformation ◽  
Peritoneal Macrophages ◽  
Marker Enzyme ◽  
3T3 Cells ◽  
Dihydroxyvitamin D3 ◽  
Tyrosine Residues ◽  
Nih 3T3 ◽  
Nih 3T3 Cells

The mutant c-fgr protein (p58c-fgr/F523) containing Phe-523 instead of Tyr-523 exhibited transforming activity in NIH 3T3 cells like other protein-tyrosine kinases of the src family, but normal p58c-fgr (p58c-fgr/wt) did not. The mutant protein showed tyrosine kinase activity threefold higher than that of the normal protein in vitro. Surprisingly, transfection of the normal c-fgr gene into NIH 3T3 cells resulted in induction of sodium fluoride (NaF)-sensitive alpha-naphthyl butyrate esterase (alpha-NBE), a marker enzyme of cells of monocytic origin, which was not induced in v-src-, v-fgr-, or lyn-transfected NIH 3T3 cells. The NaF-sensitive alpha-NBE induced in c-fgr transfectants was shown by isoelectric focusing to have a pI of 5.2 to 5.4, a range which was the same as those for thioglycolate-induced murine peritoneal macrophages and 1 alpha,25-dihydroxyvitamin D3-treated WEHI-3B cells. Immunoblotting studies with antiphosphotyrosine antibodies revealed that 58-, 62-, 75-, 120-, 200-, and 230-kDa proteins were commonly phosphorylated at tyrosine residues in NIH 3T3 cells transfected with normal and mutated c-fgr, while 95-kDa protein was significantly phosphorylated at tyrosine residues in cells transfected with the mutated c-fgr. These findings suggest that tyrosine phosphorylation of specific cellular substrate proteins is important in induction of NaF-sensitive alpha-NBE and cell transformation by p58c-fgr.

scholarly journals E2A-HLF-mediated cell transformation requires both the trans-activation domains of E2A and the leucine zipper dimerization domain of HLF.

1995 ◽  
Vol 15 (6) ◽  
pp. 3247-3255 ◽  
Author(s):  
T Yoshihara ◽  
T Inaba ◽  
L H Shapiro ◽  
J Y Kato ◽  
A T Look
Keyword(s):  
Leucine Zipper ◽  
Target Genes ◽  
Cell Transformation ◽  
Fusion Gene ◽  
Phase Cell ◽  
3T3 Cells ◽  
Dimerization Domain ◽  
Activation Domains ◽  
Nih 3T3 ◽  
Nih 3T3 Cells

The E2A-HLF fusion gene, formed by the t(17;19)(q22;p13) translocation in childhood acute pro-B-cell leukemia, encodes a hybrid protein that contains the paired trans-activation domains of E2A (E12/E47) linked to the basic region/leucine zipper DNA-binding and dimerization domain of hepatic leukemia factor (HLF). To assess the transforming potential of this novel gene, we introduced it into NIH 3T3 murine fibroblasts by using an expression vector that also contained the neomycin resistance gene. Cells selected for resistance to the neomycin analog G418 formed aberrant colonies in monolayer cultures, marked by increased cell density and altered morphology. Transfected cells also grew readily in soft agar, producing colonies whose sizes correlated with E2A-HLF expression levels. Subclones expanded from colonies with high levels of the protein reproducibly formed tumors in nude mice and grew to higher plateau-phase cell densities in reduced-serum conditions than did parental NIH 3T3 cells. By contrast, NIH 3T3 cells expressing mutant E2A-HLF proteins that lacked either of the bipartite E2A trans-activation domains or the HLF leucine zipper domain failed to show oncogenic properties, including anchorage-independent cell growth. Thus, both of the E2A trans-activation motifs and the HLF leucine zipper dimerization domain are essential for the transforming potential of the chimeric E2A-HLF protein, suggesting a model in which aberrant regulation of the expression pattern of downstream target genes contributes to leukemogenesis.

scholarly journals Interactions of phosphatidylinositol kinase, GTPase-activating protein (GAP), and GAP-associated proteins with the colony-stimulating factor 1 receptor.

1990 ◽  
Vol 10 (11) ◽  
pp. 5601-5608 ◽  
Author(s):  
M Reedijk ◽  
X Q Liu ◽  
T Pawson
Keyword(s):  
Cell Transformation ◽  
Cellular Transformation ◽  
Colony Stimulating Factor ◽  
Gtpase Activating Protein ◽  
3T3 Cells ◽  
Nih 3T3 ◽  
Fibroblast Transformation ◽  
Insert Region ◽  
Stimulating Factor ◽  
Nih 3T3 Cells

The interactions of the macrophage colony-stimulating factor 1 (CSF-1) receptor with potential targets were investigated after ligand stimulation either of mouse macrophages or of fibroblasts that ectopically express mouse CSF-1 receptors. In Rat-2 cells expressing the mouse CSF-1 receptor, full activation of the receptor and cellular transformation require exogenous CSF-1, whereas NIH 3T3 cells expressing mouse c-fms are transformed by autocrine stimulation. Activated CSF-1 receptors physically associate with a phosphatidylinositol (PI) 3'-kinase. A mutant CSF-1 receptor with a deletion of the kinase insert region was deficient in its ability to bind functional PI 3'-kinase and to induce PI 3'-kinase activity precipitable with antiphosphotyrosine antibodies. In fibroblasts, CSF-1 stimulation also induced the phosphorylation of the GTPase-activating protein (GAP)-associated protein p62 on tyrosine, although GAP itself was a relatively poor substrate. In contrast to PI 3'-kinase association, phosphorylation of p62 and GAP was not markedly affected by deletion of the kinase insert region. These results indicate that the kinase insert region selectively enhances the CSF-1-dependent association of the CSF-1 receptor with active PI 3'-kinase. The insert deletion mutant retains considerable transforming activity in NIH 3T3 cells (G. Taylor, M. Reedijk, V. Rothwell, L. Rohrschneider, and T. Pawson, EMBO J. 8:2029-2037, 1989). This mutant was more seriously impaired in Rat-2 cell transformation, although mutant-expressing Rat-2 cells still formed small colonies in soft agar in the presence of CSF-1. Therefore, phosphorylation of GAP and p62 through activation of the CSF-1 receptor does not result in full fibroblast transformation. The interaction between the CSF-1 receptor and PI 3'-kinase may contribute to c-fms fibroblast transformation and play a role in CSF-1-stimulated macrophages.

scholarly journals Dual Roles for NFAT Transcription Factor Genes as Oncogenes and Tumor Suppressors

2008 ◽  
Vol 28 (23) ◽  
pp. 7168-7181 ◽  
Author(s):  
Bruno K. Robbs ◽  
Andre L. S. Cruz ◽  
Miriam B. F. Werneck ◽  
Giuliana P. Mognol ◽  
João P. B. Viola
Keyword(s):  
Cell Cycle ◽  
Transcription Factor ◽  
Transcription Factors ◽  
Cell Transformation ◽  
Tumor Formation ◽  
3T3 Cells ◽  
Activated T Cells ◽  
Nih 3T3 ◽  
Nih 3T3 Cells

ABSTRACT Nuclear factor of activated T cells (NFAT) was first described as an activation and differentiation transcription factor in lymphocytes. Several in vitro studies suggest that NFAT family members are redundant proteins. However, analysis of mice deficient for NFAT proteins suggested different roles for the NFAT family of transcription factors in the regulation of cell proliferation and apoptosis. NFAT may also regulate several cell cycle and survival factors influencing tumor growth and survival. Here, we demonstrate that two constitutively active forms of NFAT proteins (CA-NFAT1 and CA-NFAT2 short isoform) induce distinct phenotypes in NIH 3T3 cells. Whereas CA-NFAT1 expression induces cell cycle arrest and apoptosis in NIH 3T3 fibroblasts, CA-NFAT2 short isoform leads to increased proliferation capacity and induction of cell transformation. Furthermore, NFAT1-deficient mice showed an increased propensity for chemical carcinogen-induced tumor formation, and CA-NFAT1 expression subverted the transformation of NIH 3T3 cells induced by the H-rasV12 oncogene. The differential roles for NFAT1 are at least partially due to the protein C-terminal domain. These results suggest that the NFAT1 gene acts as a tumor suppressor gene and the NFAT2 short isoform acts gene as an oncogene, supporting different roles for the two transcription factors in tumor development.

scholarly journals Vav3 Mediates Receptor Protein Tyrosine Kinase Signaling, Regulates GTPase Activity, Modulates Cell Morphology, and Induces Cell Transformation

2000 ◽  
Vol 20 (24) ◽  
pp. 9212-9224 ◽  
Author(s):  
Liyu Zeng ◽  
Pallavi Sachdev ◽  
Lunbiao Yan ◽  
Joseph L. Chan ◽  
Thomas Trenkle ◽  
...  
Keyword(s):  
Growth Factor ◽  
Cell Morphology ◽  
Protein Tyrosine Kinase ◽  
Cell Transformation ◽  
Receptor Protein ◽  
Gtpase Activity ◽  
3T3 Cells ◽  
Receptor Protein Tyrosine Kinase ◽  
Nih 3T3 ◽  
Nih 3T3 Cells

ABSTRACT A recently reported new member of the Vav family proteins, Vav3 has been identified as a Ros receptor protein tyrosine kinase (RPTK) interacting protein by yeast two-hybrid screening. Northern analysis shows that Vav3 has a broad tissue expression profile that is distinct from those of Vav and Vav2. Two species of Vav3 transcripts, 3.4 and 5.4 kb, were detected with a differential expression pattern in various tissues. Transient expression of Vav in 293T and NIH 3T3 cells demonstrated that ligand stimulation of several RPTKs (epidermal growth factor receptor [EGFR], Ros, insulin receptor [IR], and insulin-like growth factor I receptor [IGFR]) led to tyrosine phosphorylation of Vav3 and its association with the receptors as well as their downstream signaling molecules, including Shc, Grb2, phospholipase C (PLC-γ), and phosphatidylinositol 3 kinase. In vitro binding assays using glutathione S-transferase-fusion polypeptides containing the GTPase-binding domains of Rok-α, Pak, or Ack revealed that overexpression of Vav3 in NIH 3T3 cells resulted in the activation of Rac-1 and Cdc42 whereas a deletion mutant lacking the N-terminal calponin homology and acidic region domains activated RhoA and Rac-1 but lost the ability to activate Cdc42. Vav3 induced marked membrane ruffles and microspikes in NIH 3T3 cells, while the N-terminal truncation mutants of Vav3 significantly enhanced membrane ruffle formation but had a reduced ability to induce microspikes. Activation of IR further enhanced the ability of Vav3 to induce membrane ruffles, but IGFR activation specifically promoted Vav3-mediated microspike formation. N-terminal truncation of Vav3 activated its transforming potential, as measured by focus-formation assays. We conclude that Vav3 mediates RPTK signaling and regulates GTPase activity, its native and mutant forms are able to modulate cell morphology, and it has the potential to induce cell transformation.

Human c-fgr induces a monocyte-specific enzyme in NIH 3T3 cells

1991 ◽  
Vol 11 (12) ◽  
pp. 6279-6285
Author(s):  
K Inoue ◽  
B Wongsasant ◽  
T Akiyama ◽  
K Toyoshima
Keyword(s):  
Tyrosine Kinases ◽  
Cell Transformation ◽  
Peritoneal Macrophages ◽  
Marker Enzyme ◽  
3T3 Cells ◽  
Dihydroxyvitamin D3 ◽  
Tyrosine Residues ◽  
Nih 3T3 ◽  
Nih 3T3 Cells

The mutant c-fgr protein (p58c-fgr/F523) containing Phe-523 instead of Tyr-523 exhibited transforming activity in NIH 3T3 cells like other protein-tyrosine kinases of the src family, but normal p58c-fgr (p58c-fgr/wt) did not. The mutant protein showed tyrosine kinase activity threefold higher than that of the normal protein in vitro. Surprisingly, transfection of the normal c-fgr gene into NIH 3T3 cells resulted in induction of sodium fluoride (NaF)-sensitive alpha-naphthyl butyrate esterase (alpha-NBE), a marker enzyme of cells of monocytic origin, which was not induced in v-src-, v-fgr-, or lyn-transfected NIH 3T3 cells. The NaF-sensitive alpha-NBE induced in c-fgr transfectants was shown by isoelectric focusing to have a pI of 5.2 to 5.4, a range which was the same as those for thioglycolate-induced murine peritoneal macrophages and 1 alpha,25-dihydroxyvitamin D3-treated WEHI-3B cells. Immunoblotting studies with antiphosphotyrosine antibodies revealed that 58-, 62-, 75-, 120-, 200-, and 230-kDa proteins were commonly phosphorylated at tyrosine residues in NIH 3T3 cells transfected with normal and mutated c-fgr, while 95-kDa protein was significantly phosphorylated at tyrosine residues in cells transfected with the mutated c-fgr. These findings suggest that tyrosine phosphorylation of specific cellular substrate proteins is important in induction of NaF-sensitive alpha-NBE and cell transformation by p58c-fgr.

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