%0 Journal Article %T Synergistic Effect of Expressed miR-128 and Puma protein on Targeted Induction of Tumor Cell Apoptosis %J Iranian Journal of Biotechnology %I National Institute of Genetic Engineering and Biotechnology of Iran %Z 1728-3043 %A Khoshtinat Nikkhoi, Shahryar %A Dorostkar, Ruhollah %A Ranjbar, Saeed %A Heydarzadeh, Hedieh %A Tat, Mahdi %A Ghalavand, Majdedin %A Farasat, Alireza %A Hashemzadeh, Mohammad Sadegh %D 2016 %\ 09/01/2016 %V 14 %N 3 %P 185-191 %! Synergistic Effect of Expressed miR-128 and Puma protein on Targeted Induction of Tumor Cell Apoptosis %K AAV %K Adeno-Associated Virus %K Gene Therapy %K Puma %K Suicide gene %R DOI:10.15171/ijb.1429 %X Background: Puma is a highly robust pro-apoptotic protein. The protein becomes activated by p53 ensuing beyond-repair DNA damage. Downregulation of SIRT 1, by miR-128, elevates activated p53 that foment Puma indirectly. Objectives: In the present study, we used two-expression Adeno-Associated Virus (AAV) system for co-expression of miR-128 and Puma in order to evaluate apoptotic response; both in the tumor and normal cells, respectively.  Materials and Methods: Three recombinant AAVs constructs were generated. The First rAAV bearing Puma under the control of hTERT (p-AAV), the second construct designed such that to carry miR-128 downstream of CMV (mi-AAV), and the last construct comprises of the both CMV-miR-128 and hTERT- Puma. Real-Time PCR and western blotting were used to evaluate expression levels of the transduced genes. Results: MTT assay and DAPI staining shown suicidal effect of each recombinant AAV vectors. p-AAV cytotoxicity was recorded for 62% of the tumor cells, while for normal cells it was only 20% cytotoxic. The second construct, mi-AAV, was not as potent and selective as p-AAV. This construct was shown to be 27% and 16% cytotoxic for BT-474 and HEK-293 cells, respectively. Co-expression of Puma and miR-128 (p-mi-AAV) was accomplished with a selective cytotoxicity toward BT-474. This construct was 85% toxic for tumor cells, although it was only 25% toxic for the normal cell line (HEK-293). Conclusions: In this study, we have shown that not only Puma is able to instigate apoptotic response but also its co-expression along with miR-128 could significantly enhance apoptosis in a synergistic manner. %U https://www.ijbiotech.com/article_15461_dab0558438e11cbd03b3d9af42232f1a.pdf