Improving cooperativity of transcription activators by oligomerization domains in mammalian cells

Xinmao Chen; Ziming Liu; Chunbo Lou; Ying Guan; Qi Ouyang; Yanhui Xiang

doi:10.1016/j.synbio.2022.12.003

Improving cooperativity of transcription activators by oligomerization domains in mammalian cells

Synth Syst Biotechnol. 2022 Dec 17;8(1):114-120. doi: 10.1016/j.synbio.2022.12.003. eCollection 2023 Mar.

Authors

Xinmao Chen¹, Ziming Liu², Chunbo Lou², Ying Guan^{1

3

2}, Qi Ouyang¹, Yanhui Xiang²

Affiliations

¹ School of Physics, Peking University, Beijing, 100871, China.
² Center for Cell and Gene Circuit Design, CAS Key Laboratory of Quantitative Engineering Biology, Guangdong Provincial Key Laboratory of Synthetic Genomics, Shenzhen Key Laboratory of Synthetic Genomics, Shenzhen Institute of Synthetic Biology, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen, 518055, China.
³ Department of Chemical Engineering, Tsinghua University, Beijing, 100871, China.

Abstract

Cooperative activation is critical for the applications of synthetic biology in mammalian cells. In this study, we have developed cooperative transcription factor by fusing oligomerization domain in mammalian cells. Firstly, we demonstrated that two oligomerized domains (CI434 and CI) successfully improved transcription factor cooperativity in bacterial cells but failed to increase cooperativity in mammalian cells, possibly because the additional mammalian activation domain disrupted their oligomerization capability. Therefore, we chose a different type of oligomerized domain (CarH_C), whose ability to oligomerize is not dependent on its C-terminal domains, to fuse with a transcription factor (RpaR) and activation domain (VTR3), forming a potential cooperative transcription activator RpaR-CarH-VTR3 for mammalian regulatory systems. Compared with RpaR-VTR3, the cooperativity of RpaR-CarH-VTR3 was significantly improved with higher Hill coefficient and a narrower input range in the inducible switch system in mammalian cells. Moreover, a mathematical model based on statistical mechanics model was developed and the simulation results supported the hypothesis that the tetramer of the CarH domain in mammalian cells was the reason for the cooperative capacity of RpaR-CarH-VTR3.