Zhang Yi, Xian Yibo, Yang Heng, Yang Xuangang, Yu Tianli, Liu Sai, Liang Minting, Jiang Xianzhi, Deng Shulin
Key Laboratory of National Forestry and Grassland Administration on Plant Conservation and Utilization in Southern China & Guangdong Provincial Key Laboratory of Applied Botany, South China Botanical Garden, Chinese Academy of Sciences, Guangzhou, Guangdong, China.
Moon (Guangzhou) Biotech Co., Ltd, Guangzhou, Guangdong, China.
Plant Biotechnol J. 2025 Apr;23(4):1053-1066. doi: 10.1111/pbi.14561. Epub 2024 Dec 26.
Exploring the new elements to re-design the expression cassette is crucial in synthetic biology. Viruses are one of the most important sources for exploring gene expression elements. In this study, we found that the DNA sequence of the SBG51 deltasatellite from the Sweet potato leaf curl virus (SPLCV) greatly enhanced the gene expression when flanked downstream of the terminator. The SBG51 sequence increased transient GFP gene expression in Nicotiana benthamiana leaves by up to ~6 times and ~10 times compared to the gene expression controlled by the UBQ10 promoter and 35S promoter alone, respectively. The increased GFP gene expression level contributed to the continuous accumulation of GFP protein and GFP fluorescence until 8 days post-inoculation (dpi). The SBG51 sequence also enhanced the gene expression in the transgenic Arabidopsis plants and maintained the spatio-temporal pattern of the FLOWERING LOCUS T (FT) and TOO MANY MOUTHS (TMM) promoters. We identified a 123 bp of AT-rich sequence containing seven "ATAAA" or "TTAAA" elements from the SBG51 DNA, which had the gene expression enhancement effect. Furthermore, the artificial synthetic sequences containing tandem repeated "ATAAA" or "TTAAA" elements were sufficient to increase the gene expression but did not alter the polyadenylation of mRNA, similar to the function of matrix attachment regions (MAR). Additionally, the compact artificial synthetic sequence also had an effect on yeast when the expression cassette was integrated into the genome. We conclude that the geminivirus deltasatellite-derived sequence and the "ATAAA"/"TTAAA" elements are powerful tools for enhancing gene expression.
探索用于重新设计表达盒的新元件在合成生物学中至关重要。病毒是探索基因表达元件的最重要来源之一。在本研究中，我们发现来自甘薯卷叶病毒（SPLCV）的SBG51卫星DNA序列在终止子下游侧翼时能极大地增强基因表达。与单独由UBQ10启动子和35S启动子控制的基因表达相比，SBG51序列使本氏烟草叶片中的瞬时GFP基因表达分别提高了约6倍和10倍。GFP基因表达水平的提高导致GFP蛋白和GFP荧光持续积累直至接种后8天（dpi）。SBG51序列还增强了转基因拟南芥植株中的基因表达，并维持了开花位点T（FT）和过多嘴巴（TMM）启动子的时空模式。我们从SBG51 DNA中鉴定出一段123 bp富含AT的序列，其中包含七个“ATAAA”或“TTAAA”元件，该序列具有基因表达增强作用。此外，含有串联重复“ATAAA”或“TTAAA”元件的人工合成序列足以增加基因表达，但不会改变mRNA的聚腺苷酸化，类似于基质附着区域（MAR）的功能。此外，当表达盒整合到基因组中时，紧凑的人工合成序列对酵母也有影响。我们得出结论，双生病毒卫星衍生序列和“ATAAA”/“TTAAA”元件是增强基因表达的有力工具。
