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Specific interference between two unrelated internal ribosome entry site elements impairs translation efficiency
Sandrine Reigadas1, Almudena Pacheco, Jorge Ramajo
1Centro de Biologia Molecular Severo Ochoa, CSIC-UAM, Cantoblanco, 28049, Madrid, Spain.
FEBS Letters
|December 7, 2005
Summary
Two internal ribosome entry site (IRES) elements, foot-and-mouth disease virus (FMDV) and immunoglobulin heavy chain binding protein (BiP), showed competition in tricistronic vectors, reducing protein synthesis efficiency. Hepatitis C virus (HCV) IRES did not interfere.
Area of Science:
- Molecular Biology
- Virology
- Cell Biology
Background:
- Internal ribosome entry site (IRES) elements enable cap-independent translation initiation, allowing for the synthesis of multiple proteins from a single messenger RNA (mRNA) transcript in eukaryotic cells.
- IRES elements are crucial in viral replication and cellular processes, offering a mechanism for complex gene expression regulation.
Purpose of the Study:
- To investigate the potential interference between different IRES elements when co-expressed in a tricistronic construct.
- To determine if the efficiency of IRES-mediated translation is affected by the presence of other IRES elements on the same mRNA.
- To explore the mechanism of interference, specifically focusing on competition for shared trans-acting factors.
Main Methods:
- Construction of tricistronic vectors incorporating two distinct IRES elements: foot-and-mouth disease virus (FMDV) IRES and immunoglobulin heavy chain binding protein (BiP) IRES.
- Functional analysis of protein synthesis efficiency from each cistron in the tricistronic vector using in vitro and in vivo assays.
- Comparative analysis of IRES efficiency in bicistronic and tricistronic contexts.
- Assessment of interference using constructs with FMDV IRES and hepatitis C virus (HCV) IRES.
Main Results:
- The BiP IRES showed reduced protein synthesis efficiency in the tricistronic vector compared to its efficiency in bicistronic constructs.
- A specific competition was observed between the FMDV IRES and the BiP IRES, impacting their relative translation efficiencies.
- The FMDV IRES demonstrated interference effects both in cis and in trans.
- No interference was detected when the FMDV IRES was co-expressed with the HCV IRES.
Conclusions:
- The FMDV IRES and BiP IRES compete for common trans-acting factors, leading to reduced translation efficiency in a tricistronic setting.
- This competition is specific and does not involve factors required for cap-dependent or HCV IRES-dependent translation initiation.
- The findings highlight the complex interplay between different IRES elements and have implications for designing synthetic gene expression systems.