Journal
CELLULAR AND MOLECULAR LIFE SCIENCES
Volume 74, Issue 8, Pages 1431-1455Publisher
SPRINGER BASEL AG
DOI: 10.1007/s00018-016-2409-5
Keywords
CITE; Cellular IRES; RNA transfection; Cap-independent translation; RRL; Bicistronic vector
Categories
Funding
- Russian Science Foundation [16-14-10065]
- Russian Foundation for Basic Research [13-04-00903a, 16-04-0162816]
- Russian Science Foundation [16-14-10065] Funding Source: Russian Science Foundation
Ask authors/readers for more resources
The idea of internal initiation is frequently exploited to explain the peculiar translation properties or unusual features of some eukaryotic mRNAs. In this review, we summarize the methods and arguments most commonly used to address cases of translation governed by internal ribosome entry sites (IRESs). Frequent mistakes are revealed. We explain why cap-independent does not readily mean IRES-dependent and why the presence of a long and highly structured 5' untranslated region (5'UTR) or translation under stress conditions cannot be regarded as an argument for appealing to internal initiation. We carefully describe the known pitfalls and limitations of the bicistronic assay and artefacts of some commercially available in vitro translation systems. We explain why plasmid DNA transfection should not be used in IRES studies and which control experiments are unavoidable if someone decides to use it anyway. Finally, we propose a workflow for the validation of IRES activity, including fast and simple experiments based on a single genetic construct with a sequence of interest.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available