A Foamy Virus Vector System for Stable and Efficient RNAi Expression in Mammalian Cells
| Autor: | George Vassilopoulos, Elena K. Siapati, Magdalini Papadaki |
|---|---|
| Rok vydání: | 2011 |
| Předmět: |
Genetic Vectors
Green Fluorescent Proteins Cell Fusion Proteins bcr-abl Mice Transgenic Biology Real-Time Polymerase Chain Reaction RNA polymerase III Viral vector Small hairpin RNA Mice Transduction (genetics) Transduction Genetic RNA interference Genetics medicine Animals Humans Gene silencing Gene Silencing RNA Small Interfering Promoter Regions Genetic Molecular Biology Bone Marrow Transplantation DNA Primers RNA Polymerase III Flow Cytometry Hematopoietic Stem Cells Molecular biology medicine.anatomical_structure Microscopy Fluorescence Spumavirus Molecular Medicine RNA Interference Stem cell K562 Cells |
| Zdroj: | Human Gene Therapy. 22:1293-1303 |
| ISSN: | 1557-7422 1043-0342 |
| DOI: | 10.1089/hum.2010.223 |
| Popis: | The promise of the RNA interference (RNAi) technology is equally dependent on the efficiency and stability of gene silencing. The aim of the present study was the development of foamy virus (FV) vectors for stable RNAi, utilizing two potent RNA polymerase III (Pol III) promoters. Using green fluorescent protein as a target gene, we examined the efficiency of mouse U6 (mU6) and human H1 Pol III promoters in different human cell lines and mouse hematopoietic stem cells (HSCs) ex vivo and in vivo, following bone marrow transplantation. Both our mU6 and H1 FV vectors mediated very efficient gene silencing with as low as one vector copy per cell. However, transduction of human cell lines with FV vectors expressing short hairpin RNA from mU6 led to the gradual elimination of cells in culture, as opposed to H1-harboring cells, underscoring the importance of the expression system or cellular context in the evaluation of the overall RNAi effects. The efficiency and stability of the H1 vectors were further shown by the successful silencing of BCR-ABL in K562 cells. Accordingly, mU6 vectors induced efficient and stable gene silencing in mouse HSCs following bone marrow transplantation. Our work is the first in vivo study on the efficiency and stability of RNAi gene silencing in HSCs with FV vectors, currently a safe alternative for viral gene transfer. |
| Databáze: | OpenAIRE |
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