Repression of the Internal Ribosome Entry Site-dependent Translation of Hepatitis C Virus by an Engineered PUF Protein
| Author | Seyed Jalal Kiani | en |
| Author | Tahereh Taheri | en |
| Author | Ahmad Nejati | en |
| Author | Monireh Maleki | en |
| Author | Sima Rafati | en |
| Author | Kayhan Azadmanesh | en |
| Author | Seyed Moayed Alavian | en |
| Author | Talat Mokhtari Azad | en |
| Author | Katayoun Samimi-Rad | en |
| Orcid | Katayoun Samimi-Rad [0000-0001-9705-2605] | en |
| Issued Date | 2017-02-01 | en |
| Abstract | Background: Pumilio/fem-3 mRNA binding factor (PUF) proteins can bind RNA in a sequence-specific manner. The deciphered RNA-recognition code of these proteins has enabled researchers to design engineered PUF proteins, capable of binding to any desired target in order to modify its ultimate fate. In this study, a modified Homo sapiens Pumilio 1-homology domain (HsPUM1-HD) was engineered to bind to the internal ribosome entry site (IRES) of hepatitis C virus (HCV) genome to potentially inhibit viral translation. Methods: Based on the RNA-recognition code, required modifications were applied to HsPUM1-HD in order to change its natural recognition sequence to a sequence in the stem-loop III of HCV IRES. RNA protein pull-down assay was performed to assess the sequence specificity of the modified HsPUM1-HD (mHsPUM1-HD). Translational inhibitory effect of mHsPUM1-HD was evaluated in a dual-luciferase reporter assay. Results: The mHsPUM1-HD was found to bind to its cognate RNA in a sequence-specific manner, as a biotinylated target RNA captured mHsPUM1-HD through binding to streptavidin magnetic beads. This protein also reduced HCV IRES-dependent firefly luciferase translation by 40% in HEK293 cells. Conclusions: The present study is the first report of an engineered HsPUM1-HD with potential anti-HCV activity. These findings suggest that PUM-HDs can be engineered to target desired RNAs of infectious agents in order to specifically interrupt protein translation, as an essential step of their life cycle. | en |
| DOI | https://doi.org/10.5812/hepatmon.45022 | en |
| Keyword | Hepatitis C Virus | en |
| Keyword | Pumilio Protein | en |
| Keyword | Internal Ribosome Entry Sites | en |
| Publisher | Brieflands | en |
| Title | Repression of the Internal Ribosome Entry Site-dependent Translation of Hepatitis C Virus by an Engineered PUF Protein | en |
| Type | Research Article | en |