Inhibition of Platelet-derived Growth Factor-induced Cell Growth Signaling by a Short Interfering RNA for EWS-Fli1 via Down-regulation of Phospholipase D2 in Ewing Sarcoma Cells

Abstract

EWS-Ewing sarcoma and other peripheral primitive neuroectodermal tumors are pediatric malignant solid tumors, Ͼ95% of which show the chromosomal translocations t(11;22, q24;q12) or t(21;22, q22;q12), which produce the fusion genes EWS-Fli1 and EWS-erg, respectively (1-4). Cytogenetic and molecular analyses of these translocation products have revealed that the 5Ј-region of the EWS gene (from band 22q12) is fused to the 3Ј-region of either the Fli-1 gene (from band 11q24) or the erg gene (from band 21q22), both of which are members of the ETS family of transcription factors. Less frequent chromosomal translocations involve the same region of the EWS gene and various other ETS genes, such as ETV-1, E1AF, or FEV (1, 2). We have previously shown that EWS preferentially binds to poly(G) and poly(U) RNA. The binding activity of EWS is located in the RGG box, which is in the carboxyl-terminal region.

Autoregulation of phospholipase D activity is coupled to selective induction of phospholipase D1 expression to promote invasion of breast cancer cells

“…Endogenous levels of PA are low in resting cells, and PLD activity is tightly regulated by mechanisms that control secretion, migration, survival and proliferation of cells. 1,2 PLD is activated by a variety of mitogens, including epidermal growth factor (EGF), platelet-derived growth factor (PDGF) and interleukin 1b (IL-1b), [3][4][5][6][7] and its involvement in many cellular biological processes, including cell proliferation, survival, vesicular trafficking and secretion, has been demonstrated. 1,8 Two distinct isoforms of mammalian PLD, PLD1 and PLD2, have been identified.…”

Ewing's sarcoma fusion protein, EWS/Fli-1 and Fli-1 protein induce PLD2 but not PLD1 gene expression by binding to an ETS domain of 5′ promoter

“…A recent study showed that the reduced EWS/Fli-1 expression by using siRNA induces downregulation of PLD2 protein expression (Nozawa et al, 2005). However, the mechanism of PLD2 gene expression by EWS/ Fli-1 was not studied.…”

“…siRNA of PLD2 and of EWS were obtained from Sigma Genosys (Hokkaido, Japan). The sequence of siRNAs of EWS and of PLD2 was according to Nozawa et al (2005).…”

“…A recent report has demonstrated that the inhibition of EWS/Fli-1 by using short interfering RNA (siRNA) downregulates PLD2 protein expression, resulting in the repression of platelet-derived growth factor-mediated cell growth signaling in EWS cells (Nozawa et al, 2005). We have extended this work by analysing the relationship between PLD gene expression and EWS/Fli-1 or Fli-1.…”

Mutation of Y179 on phospholipase D2 (PLD2) upregulates DNA synthesis in a PI3K-and Akt-dependent manner

“…PLD plays a role in growth factor-induced cellular proliferation [3][4][5][6] and facilitates oncogenic transformation [7][8][9][10]. Endogenous PLD activity is significantly increased in several types of cancers [11][12][13][14][15], as well as in cell lines transformed with oncogenes [4,5,8,10,[16][17][18][19]. PLD may participate in the activation of p42/44 ERK via Raf-1 translocation to the plasma membrane [20][21][22][23], as well as in the survival signaling mediated by PI3K/AKT activation [24].…”