2019 ; Vol.29-1: 127~140
|Author||Hae Li Ko, Hyo-Jung Park, Jihye Kim, Ha Kim, Hyewon Youn, Jae-Hwan Nam|
|Place of duty||Department of Biotechnology, The Catholic University of Korea, Bucheon, Korea |
|Title||Development of an RNA Expression Platform Controlled by Viral Internal Ribosome Entry Sites|
J. Microbiol. Biotechnol.2019 ;
|Abstract||Since 1990, many nucleic acid expression platforms consisting of DNA or RNA have been
developed. However, although RNA expression platforms have been relatively neglected,
several such platforms capped at the 5’ end of RNA by an anti-reverse cap analog have now
been developed. At the same time, the capping reaction is a bottleneck in the production of
such platforms, with high cost and low efficiency. Here, we investigated several viral and
eukaryotic internal ribosome entry sites (IRESs) to develop an optimal RNA expression
platform, because IRES-dependent translation does not require a capping step. RNA
expression platforms constructed with IRESs from the 5’ untranslated regions of the
encephalomyocarditis virus (EMCV) and the intergenic region of the cricket paralysis virus
(CrPV) showed sufficient expression efficiency compared with cap-dependent RNA
expression platforms. However, eukaryotic IRESs exhibited a lower viral IRES expression
efficiency. Interestingly, the addition of a poly(A) sequence to the 5’ end of the coxsackievirus
B3 (CVB3) IRES (pMA-CVB3) increased the expression level compared with the CVB3 IRES
without poly(A) (pCVB3). Therefore, we developed two multiexpression platforms (termed
pMA-CVB3-EMCV and pCrPV-EMCV) by combining the IRESs of CVB3, CrPV, and EMCV in
a single-RNA backbone. The pMA-CVB3-EMCV-derived RNA platform showed the highest
expression level. Moreover, it clearly exhibited expression in mouse muscles in vivo. These
RNA expression platforms prepared using viral IRESs will be useful in developing potential
RNA-based prophylactic or therapeutic vaccines, because they have better expression
efficiency and do not need a capping step.|
|Key_word||Internal ribosome entry sites, RNA expression platform, coxsackievirus B3, cricket paralysis virus, encephalomyocarditis virus, poly(A)|
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