Repository of Research and Investigative Information

Repository of Research and Investigative Information

Baqiyatallah University of Medical Sciences

Synergistic Effect of Expressed miR-128 and Puma protein on Targeted Induction of Tumor Cell Apoptosis

(2016) Synergistic Effect of Expressed miR-128 and Puma protein on Targeted Induction of Tumor Cell Apoptosis. Iranian Journal of Biotechnology. pp. 125-131. ISSN 1728-3043

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Abstract

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.

Item Type: Article
Keywords: AAV Adeno-Associated Virus Gene therapy Puma Suicide gene recombinant adenoassociated virus cancer-cells gene-therapy p53 tumorigenesis activation chromatin htert dna Biotechnology & Applied Microbiology
Divisions:
Page Range: pp. 125-131
Journal or Publication Title: Iranian Journal of Biotechnology
Journal Index: ISI
Volume: 14
Number: 3
Identification Number: https://doi.org/10.15171/ijb.1429
ISSN: 1728-3043
Depositing User: مهندس مهدی شریفی
URI: http://eprints.bmsu.ac.ir/id/eprint/4978

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