Li Xinyi, Wang Song, Li Qianyi, Li Xiangyu, Lin Sirao, Zhao Wenyu, Liu Yingqi, Wu Bowen, Huang Ying, Jia Bin, Hu Zhangli
Guangdong Technology Research Center for Marine Algal Bioengineering, Guangdong Provincial Key Laboratory for Plant Epigenetics, Shenzhen Engineering Laboratory for Marine Algal Biotechnology, College of Life Sciences and Oceanography, Shenzhen University, Shenzhen, China.
School of Life Science and Technology, Southeast University, Nanjing, China.
Plant Cell Environ. 2025 Jun;48(6):3913-3924. doi: 10.1111/pce.15360. Epub 2025 Jan 22.
Chlamydomonas reinhardtii, a prominent chassis in synthetic biology, faces limitations in regulating the expression of exogenous genes. A destabilization domain (DD)/Shield-1 system, originally derived from mammals, offers a ligand-dependent control of stability, making it a valuable tool. This system utilises the destabilization domain to induce rapid degradation of target protein unless stabilised by Shield-1, a synthetic ligand. Upon the addition of Shield-1,the degradation is halted, leading to the accumulation and stabilisation of the target protein. This system has demonstrated successful regulation of foreign protein expression in mammals, parasites, and plants. In this study, the DD/Shield-1 system was harnessed to regulate the expression of the paromomycin resistance gene and luciferase encoding gene in Chlamydomonas, revealing its capability for rapid, stable, and reversible protein expression regulation in microalgae, serving as a molecular switch. Furthermore, this regulation exhibits reagent dependency, enhancing its applicability in practical production. A strain with induced expression of the gene-editing protein, LbCas12a, was successfully constructed and then tested for gene editing. The findings not only enrich the toolkit for Chlamydomonas molecular studies but offer a promising technique for regulating the expression and validating the functionality of exogenous proteins in microalgae.
莱茵衣藻是合成生物学中一种重要的底盘生物，但在调控外源基因表达方面存在局限性。一种最初源自哺乳动物的去稳定结构域（DD）/Shield-1系统，提供了一种依赖配体的稳定性控制，使其成为一种有价值的工具。该系统利用去稳定结构域诱导靶蛋白快速降解，除非被合成配体Shield-1稳定。添加Shield-1后，降解停止，导致靶蛋白积累并稳定。该系统已在哺乳动物、寄生虫和植物中成功调控外源蛋白表达。在本研究中，利用DD/Shield-1系统调控莱茵衣藻中对氨基糖苷类抗生素抗性基因和荧光素酶编码基因的表达，揭示了其在微藻中快速、稳定和可逆地调控蛋白表达的能力，可作为一种分子开关。此外，这种调控表现出对试剂的依赖性，增强了其在实际生产中的适用性。成功构建了基因编辑蛋白LbCas12a诱导表达的菌株，并对其进行了基因编辑测试。这些发现不仅丰富了莱茵衣藻分子研究的工具库，还为调控微藻中外源蛋白的表达和验证其功能提供了一种有前景的技术。
