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lüll Melanoma differentiation-associated gene-7 protein physically associates with the double-stranded RNA-activated protein kinase PKR Pataer A; Vorburger SA; Chada S; Balachandran S; Barber GN; Roth JA; Hunt KK; Swisher SGMol Ther 2005[May]; 11 (5): 717-23We previously reported that adenoviral-mediated overexpression of the melanoma differentiation-associated gene-7 (Ad-mda7; approved gene symbol IL24) leads to the rapid induction of PKR and activation of its downstream targets, resulting in apoptosis induction in human lung cancer cells. To evaluate the mechanism by which Ad-mda7 activates PKR, we studied the interaction between MDA-7 and PKR proteins. Following Ad-mda7 transduction of lung cancer cells, intracellular and extracellular MDA-7 protein was generated, leading to dose- and time-dependent PKR induction. Purified MDA-7 protein administered extracellularly did not induce PKR or apoptosis, suggesting that Ad-mda7-mediated PKR activation and apoptosis were not dependent on extracellular MDA-7 protein. Following Ad-mda7 transduction, RT-PCR demonstrated no increase in PKR mRNA levels despite increased levels of PKR protein, suggesting posttranscriptional regulation of PKR by MDA-7. Immunofluorescence and coimmunoprecipitation studies demonstrated that MDA-7 protein physically interacts with PKR. Transduction of PKR+/+ and PKR-/- transformed MEFs with Ad-mda7 demonstrated phosphorylated MDA-7 and PKR proteins in the lysates of PKR+/+ but not PKR-/- cells. These findings identify the first binding partner for MDA-7 and suggest that direct interaction between PKR and MDA-7 may be important for PKR activation and apoptosis induction, possibly through MDA-7 phosphorylation or activation of other downstream targets.|Adenoviridae/genetics[MESH]|Animals[MESH]|Apoptosis[MESH]|Cell Line[MESH]|Enzyme Activation[MESH]|Genes, Tumor Suppressor[MESH]|Humans[MESH]|Interleukins/genetics/*metabolism[MESH]|Lung Neoplasms/genetics/metabolism/pathology[MESH]|Mice[MESH]|Mice, Knockout[MESH]|Phosphorylation[MESH]|Protein Binding[MESH]|RNA, Double-Stranded/*metabolism[MESH]|eIF-2 Kinase/deficiency/genetics/*metabolism[MESH] |