Real-Time Temporal Dynamics of Bicistronic Expression Mediated by Internal Ribosome Entry Site and 2A Cleaving Sequence
AffiliationUniv Arizona, Coll Med Phoenix, Dept Basic Med Sci
internal ribosome entry site
real-time fluorescent imaging
MetadataShow full item record
PublisherKOREAN SOC MOLECULAR & CELLULAR BIOLOGY
CitationLee, S., Kim, J. A., Kim, H. D., Chung, S., Kim, K., & Choe, H. K. (2019). Real-Time Temporal Dynamics of Bicistronic Expression Mediated by Internal Ribosome Entry Site and 2A Cleaving Sequence. Molecules and cells, 42(5), 418.
JournalMOLECULES AND CELLS
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AbstractMulticistronic elements, such as the internal ribosome entry site (IRES) and 2A-like cleavage sequence, serve crucial roles in the eukaryotic ectopic expression of exogenous genes. For utilization of multicistronic elements, the cleavage efficiency and order of elements in multicistronic vectors have been investigated; however, the dynamics of multicistronic element-mediated expression remains unclear. Here, we investigated the dynamics of encephalomyocarditis virus (EMCV) IRES- and porcine teschovirus-1 2A (p2A)-mediated expression. By utilizing real-time fluorescent imaging at a minute-level resolution, we monitored the expression of fluorescent reporters bridged by either EMCV IRES or p2A in two independent cultured cell lines, HEK293 and Neuro2a. We observed significant correlations for the two fluorescent reporters in both multicistronic elements, with a higher correlation coefficient for p2A in HEK293 but similar coefficients for IRES-mediated expression and p2A-mediated expression in Neuro2a. We further analyzed the causal relationship of multicistronic elements by convergent cross mapping (CCM). CCM revealed that in all four conditions examined, the expression of the preceding gene causally affected the dynamics of the subsequent gene. As with the cross correlation, the predictive skill of p2A was higher than that of IRES in HEK293, while the predictive skills of the two multicistronic elements were indistinguishable in Neuro2a. To summarize, we report a significant temporal correlation in both EMCV IRES- and p2A-mediated expression based on the simple bicistronic vector and real-time fluorescent monitoring. The current system also provides a valuable platform to examine the dynamic aspects of expression mediated by diverse multicistronic elements under various physiological conditions.
NoteOpen access journal
VersionFinal published version
SponsorsNational Research Foundation of Korea (NRF) of the Ministry of Science and ICT [NRF-2017R1C1B2008775, NRF-2017R1A4A1015534, NRF-2018M3C7A1022310]; KBRI basic research program through Korea Brain Research Institute - Ministry of Science and ICT [17-BR-04]
- [Quantitative comparison of expression for genes linked in bicistronic vectors via ires or 2A-peptide of porcine teschovirus-1 sequence].
- Authors: Kuzmich AI, Vvedenskiĭ AV, Kopantsev EP, Vinogradova TV
- Issue date: 2013 Jul-Aug
- High cleavage efficiency of a 2A peptide derived from porcine teschovirus-1 in human cell lines, zebrafish and mice.
- Authors: Kim JH, Lee SR, Li LH, Park HJ, Park JH, Lee KY, Kim MK, Shin BA, Choi SY
- Issue date: 2011
- In vitro and in vivo comparison of viral and cellular internal ribosome entry sites for bicistronic vector expression.
- Authors: Licursi M, Christian SL, Pongnopparat T, Hirasawa K
- Issue date: 2011 Jun
- Long term expression of bicistronic vector driven by the FGF-1 IRES in mouse muscle.
- Authors: Allera-Moreau C, Delluc-Clavières A, Castano C, Van den Berghe L, Golzio M, Moreau M, Teissié J, Arnal JF, Prats AC
- Issue date: 2007 Oct 28
- Multicistronic lentiviral vectors containing the FMDV 2A cleavage factor demonstrate robust expression of encoded genes at limiting MOI.
- Authors: Chinnasamy D, Milsom MD, Shaffer J, Neuenfeldt J, Shaaban AF, Margison GP, Fairbairn LJ, Chinnasamy N
- Issue date: 2006 Mar 15