Protein phosphorylation in a tetradecanoyl phorbol acetate-nonproliferative variant of 3T3 cells.

Bishop, R; Martinez, R; Weber, Michael J.; Blackshear, Perry J.; Beatty, S; Lim, Robert W.; Herschman, Harvey R.

doi:10.1128/mcb.5.9.2231

Cited by 33 publications

(14 citation statements)

References 25 publications

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“…3 had the appropriate position by mixing it with purified MAP kinase, which (as shown previously [37]) migrated at the same Mr as the pp42 spot (data not shown). We conclude that the TPA-induced tyrosine phosphorylation of pp42 is defective in the 3T3-TNR9 cell variant but that tyrosine phosphorylation of at least some 45-kDa proteins occurs in these cells, a result which explains the apparent discrepancy with our earlier report (7). However, as discussed in the legend to Fig.…”

Section: Resultscontrasting

confidence: 57%

“…TNR9 cells possess normal levels of PK-C, and the activity and translocation of this enzyme in response to TPA appear to occur normally (7,18). A more recent report from Biemann and Erikson (5) indicates that the TPA-induced down-modulation of PK-C does not persist in TNR9 cells which are allowed to overgrow and form multilayers.…”

Section: Discussionmentioning

confidence: 99%

“…These cells have normal levels of PK-C and display many of the early phosphorylation and metabolic responses to TPA treatment (7,12,18) which occur in the parental 3T3 cells. However, unlike the parental 3T3 cells, they do not show an increase in ornithine decarboxylase activity or DNA synthesis in response to TPA (11,12).…”

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confidence: 99%

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Defective regulation of mitogen-activated protein kinase activity in a 3T3 cell variant mitogenically nonresponsive to tetradecanoyl phorbol acetate.

1991

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Mitogen-activated protein (MAP) kinase is a serine/threonine-specific protein kinase which is activated in response to various mitogenic agonists (e.g., epidermal growth factor, insulin, and the tumor promoter tetradecanoyl phorbol acetate [TPA]) and requires both threonine and tyrosine phosphorylation for activity. This enzyme has recently been shown to be identical or closely related to pp42, a protein which becomes tyrosine phosphorylated in response to mitogenic stimulation. Neither the kinases which regulate MAP kinase/pp42 nor the in vivo substrates for this enzyme are known. Because MAP kinase is activated and phosphorylated in response both to agents which stimulate tyrosine kinase receptors and to agents which stimulate protein kinase C, a serine/threonine kinase, we have examined the regulation and phosphorylation of this enzyme in 3T3-TNR9 cells, a variant cell line partially defective in protein kinase C-mediated signalling.In this communication, we show that in the 3T3-TNR9 variant cell line, TPA does not cause the characteristically rapid phosphorylation of pp42 or the activation and phosphorylation of MAP kinase. This defective response is not due to the absence of the MAP kinase/pp42 protein itself because both tyrosine phosphorylation of MAP kinase/pp42 and its enzymatic activation could be induced by platelet-derived growth factor in the 3T3-TNR9 cells. Thus, the defect in these variant cells apparently resides in some aspect of the regulation of MAP kinase phosphorylation. Since the 3T3-TNR9 cells are also defective with respect to the TPA-induced increase in ribosomal protein S6 kinase, these in vivo results reinforce the earlier in vitro finding that MAP kinase can regulate S6 kinase activity. These findings suggest a key role for MAP kinase in a kinase cascade involved in the control of cell proliferation.Activation of tyrosine kinase transmembrane receptors results in stimulation of a diverse array of cellular activities, such as glycolysis, protein synthesis, and transcription. However, in most cases in which the regulation of these cellular activities has been adequately investigated, critical phosphorylations of regulatory proteins occur not only on tyrosine but on serine and threonine as well. Thus, it seems likely that "switch kinases", which are regulated by tyrosine phosphorylation but function as serine/threonine kinases, are pivotal elements in the hormonal control of growth and metabolism. Elucidation of the pathways by which cell surface receptors control growth and metabolism will require analysis of the regulation of these switch kinases and determination of their downstream substrates.Mitogen-activated protein (MAP) kinase was the first switch kinase to be so identified (35). This enzyme was originally described by Ray and Sturgill (34) as a serine/ threonine-specific protein kinase which was activated by treatment of 3T3-L1 cells with insulin and which could be partially purified by sequential DEAE and hydrophobicinteraction chromatography (36). Subsequently, it was found th...

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Section: Resultscontrasting

confidence: 57%

Section: Discussionmentioning

confidence: 99%

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Defective regulation of mitogen-activated protein kinase activity in a 3T3 cell variant mitogenically nonresponsive to tetradecanoyl phorbol acetate.

1991

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“…on May 11, 2018 by guest http://mcb.asm.org/ Downloaded from 3T3-TNR9 cells, in contrast to our earlier results (5). They also reported that 3T3-TNR9 cells, in contrast to parental 3T3 cells, were defective in the ability to down regulate PKC in response to chronic TPA treatment.…”

Section: Addendumcontrasting

confidence: 56%

A Swiss 3T3 variant cell line resistant to the effects of tumor promoters cannot be transformed by src.

Nori¹,

Shawver²,

Weber³

1990

Mol. Cell. Biol.

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To study the relationship between oncogenesis by v-src and normal cellular signalling pathways, we determined the effects of v-src on 3T3-TNR9 cells, a Swiss 3T3 variant which does not respond mitogenically to tumor promoters such as 12-O-tetradecanoyl-phorbol-13-acetate (TPA). We found that src was unable to transform these variant cells, whether the oncogene was introduced by infection with a murine retrovirus vector or by transfection with plasmid DNA. 3T3-TNR9 cells were not inherently resistant to transformation, since infection with similar recombinant retroviruses containing either v-ras or v-abl did induce transformation. Further analysis of Swiss 3T3 and 3T3-TNR9 cell populations infected with the v-src-containing retrovirus revealed that although the amount of v-src DNA in each was approximately the same, the level of the v-src message and protein and the overall level of phosphotyrosine expressed in the infected variants was much less than in infected parental cells. Cotransfection experiments using separate v-src and neo plasmids revealed a decrease in the number of G418-resistant colonies when transfections of TNR9 cells occurred in the presence of the src-containing plasmid, suggesting a growth inhibitory effect of v-src on 3T3-TNR9 cells, as has also been found for TPA itself. Since v-src cannot transform this variant cell line, which does not respond mitogenically to the protein kinase C agonist TPA, we suggest that src makes use of the protein kinase C pathway as part of its signalling activities.

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“…This was surprising, since we generally find that TPA-induced phosphorylations are extremely rapid, requiring only a few minutes (5,6). Thus, the steady, prolonged increase in transporter phosphorylation in response to TPA may reflect not only the activation of protein kinase C but also the biosynthesis of new transporters or the translocation of transporters to a site where they are susceptible to phosphorylation by protein kinase C (22).…”

Section: Discussionmentioning

confidence: 96%

Degradation and biosynthesis of the glucose transporter protein in chicken embryo fibroblasts transformed by the src oncogene.

Shawver¹,

Olson²,

White³

et al. 1987

Mol. Cell. Biol.

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The rate of glucose transport in cultured fibroblasts is regulated in response to a number of physiological variables, including malignant transformation by src, glucose starvation, and stimulation with mitogens. Much of this transport regulation can be accounted for by variations in the amount of transporter protein in the cells. To determine the mechanisms by which levels of the transporter are regulated, we measured the rates of synthesis and degradation of the transporter by pulse-chase experiments and immunoprecipitation of the transporter. We found that transformation by the src oncogene results in a large decrease in the rate at which the transporter protein is degraded but that it does not appreciably increase the rate of transporter biosynthesis. On the other hand, glucose starvation and mitogen stimulation increase the rate of transporter biosynthesis, although a role for control of degradation is possible in these circumstances also. Variations in the rate of glucose transport or the amount of the transporter are not associated with phosphorylation of the transporter protein.D-Glucose is transported into fibroblasts by a stereospecific carrier-mediated facilitated transport system (for a review, see reference 47). The rate at which glucose enters these cells is regulated by a number of factors, including malignant transformation

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Protein phosphorylation in a tetradecanoyl phorbol acetate-nonproliferative variant of 3T3 cells.

Cited by 33 publications

References 25 publications

Defective regulation of mitogen-activated protein kinase activity in a 3T3 cell variant mitogenically nonresponsive to tetradecanoyl phorbol acetate.

Defective regulation of mitogen-activated protein kinase activity in a 3T3 cell variant mitogenically nonresponsive to tetradecanoyl phorbol acetate.

A Swiss 3T3 variant cell line resistant to the effects of tumor promoters cannot be transformed by src.

Degradation and biosynthesis of the glucose transporter protein in chicken embryo fibroblasts transformed by the src oncogene.

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