Real-Time Temporal Dynamics of Bicistronic Expression Mediated by Internal Ribosome Entry Site and 2A Cleaving Sequence
- 주제(키워드) 2A , expression dynamics , internal ribosome entry site , multicistronic elements , real-time fluorescent imaging
- 주제(기타) Biochemistry & Molecular Biology; Cell Biology
- 설명문(일반) [Lee, Soomin; Kim, Jeong-Ah; Kim, Kyungjin; Choe, Han Kyoung] DGIST, Dept Brain & Cognit Sci, Daegu 42988, South Korea; [Kim, Jeong-Ah] Seoul Natl Univ, Dept Biol Sci, Seoul 08826, South Korea; [Kim, Hee-Dae] Univ Arizona, Coll Med Phoenix, Dept Basic Med Sci, Phoenix, AZ 85004 USA; [Chung, Sooyoung] Ehwa Womans Univ, Scranton Coll, Dept Brain & Cognit Sci, Seoul 03760, South Korea; [Choe, Han Kyoung] KBRI, Daegu 41062, South Korea
- 관리정보기술 faculty
- 발행기관 KOREAN SOC MOLECULAR & CELLULAR BIOLOGY
- 발행년도 2019
- URI http://www.dcollection.net/handler/ewha/000000160401
- 본문언어 영어
- Published As http://dx.doi.org/10.14348/molcells.2019.2427
- PubMed https://pubmed.ncbi.nlm.nih.gov/31085809
초록/요약
Multicistronic 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.
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