A Dominant Negative RAF-1 Mutant Prevents V-SRC-Induced Transformation

Qureshi, Sajjad A.; Joseph, Cecil K.; Hendrickson, Michael L.; Song, Jianguo; Gupta, Ruchika; Bruder, JT; Rapp, Ulf R.; Foster, David A.

doi:10.1006/bbrc.1993.1510

Cited by 20 publications

(19 citation statements)

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“…Previous studies using cells in culture have suggested a role for RalA in mitogenic signaling and transformation (Urano et al, 1996). However, dominant negative RalA mutants were less e cient in blocking the transformed phenotype than dominant negative mutants of Ras and Raf (Qureshi et al, 1993;Jiang et al, 1995a,b;Urano et al, 1996). The studies provided here demonstrate conclusively that RalA inhibits completely the ability to form tumors.…”

Section: The Dominant Negative S28n Rala Mutant Prevents Vsrc and V-rsupporting

confidence: 55%

“…However, the inhibitory e ect of the dominant negative RalA on focus formation was not as strong as that observed for dominant negative mutants of Ras and Raf on v-Src-induced transformation where colony formation in soft agar was inhibited almost completely (Qureshi et al, 1993;Jiang et al, 1995a,b). We therefore investigated the e ect of the dominant negative RalA on the ability of the v-Srcand v-Ras-transformed cells to form colonies in soft agar.…”

Section: The Dominant Negative S28n Rala Mutant Prevents Vsrc and V-rmentioning

confidence: 97%

“…Increased expression of the RalA (S28N) protein levels was veri®ed by Western blot analysis (data not shown). For growth of cells in soft agar, 1610 3 cells were suspended in top agar (DMEM, 20% calf serum, 0.38% agar) and overlaid onto hardened bottom agar (DMEM, 20% calf serum, 0.7% agar) as described previously (Qureshi et al, 1993). In all experiments as a control NIH3T3 cells were vector (pZipneo)-transfected as described previously (Jiang et al, 1995a,b).…”

Section: Cells and Cell Culture Conditionsmentioning

confidence: 99%

“…PLD converts phosphatidylcholine to phosphatidic acid, which has been implicated in vesicle budding and transport, as well as the transduction of intracellular signals (Bi et al, 1997;Exton, 1998;Roth and Sternweis, 1997). Phosphatidic acid may also be metabolized further to the biologically active diacylglycerol or lysophosphatidic acid (Foster, 1993;Exton, 1994). PLD activity is increased in response to most, if not all, mitogenic signals (Foster, 1993;Exton, 1998).…”

Section: Introductionmentioning

confidence: 99%

“…Phosphatidic acid may also be metabolized further to the biologically active diacylglycerol or lysophosphatidic acid (Foster, 1993;Exton, 1994). PLD activity is increased in response to most, if not all, mitogenic signals (Foster, 1993;Exton, 1998). A PLD activity was found to be associated with the Ras-family GTPase RalA, which is required for the activation of PLD by both v-Src and vRas (Jiang et al, 1995b) and the active complex includes PLD1, RalA and Arf (Luo et al, 1997(Luo et al, , 1998.…”

Section: Introductionmentioning

confidence: 99%

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RalA requirement for v-Src- and v-Ras-induced tumorigenicity and overproduction of urokinase-type plasminogen activator: involvement of metalloproteases

et al. 1999

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Overproduction of urokinase-type plasminogen activator (uPA) and metalloproteases (MMPs) is strongly correlated with tumorigenicity and with invasive and metastatic phenotypes of human and experimental tumors. We demonstrated previously that overproduction of uPA in tumor cells is mediated by a phospholipase D (PLD)-and protein kinase C-dependent mechanism. The oncogenic stimulus of v-Src and v-Ras results in the activation of PLD, which is dependent upon the monomeric GTPase RalA. We have therefore investigated whether RalA plays a role in uPA and MMP overproduction that is observed in response to oncogenic signals. We report here that NIH3T3 cells transformed by both v-Src and v-Ras, constitutively overproduce uPA and that expression of a dominant negative RalA mutant (S28N) blocks overproduction of uPA in both the v-Srcand v-Ras-transformed cells. v-Src and v-Ras also induced an upregulation of the activity of MMP-2 and MMP-9 as detected by zymograms, however only the vSrc induction correlated with MMP protein levels detected by Western blot analysis. The dominant negative RalA mutant blocked increased MMP-2 and 9 overproduction induced by v-Src, but not the increased activity of MMP-2 and 9 induced by v-Ras. And, consistent with a role for the RalA/PLD pathway in mitogenesis and tumor development, the dominant negative RalA mutant completely blocked tumor formation by v-Src-and v-Ras-transformed NIH3T3 cells injected subcutaneously in syngeneic mice. The data presented here implicate RalA and PLD as signaling mediators for tumor formation and protease production by transformed cells.

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Section: The Dominant Negative S28n Rala Mutant Prevents Vsrc and V-rsupporting

confidence: 55%

Section: The Dominant Negative S28n Rala Mutant Prevents Vsrc and V-rmentioning

confidence: 97%

Section: Cells and Cell Culture Conditionsmentioning

confidence: 99%