University of Technology Sydney, Climate Change Cluster, Broadway Campus, Ultimo, Sydney, NSW, 2007, Australia.
School of Life Sciences and Proteomics Core Facility, Faculty of Science, University of Technology Sydney, Ultimo, Sydney, NSW, 2007, Australia.
BMC Microbiol. 2024 Oct 8;24(1):392. doi: 10.1186/s12866-024-03554-4.
Chlamydomonas reinhardtii is gaining recognition as a promising expression system for the production of recombinant proteins. However, its performance as a cellular biofactory remains suboptimal, especially with respect to consistent expression of heterologous genes. Gene silencing mechanisms, position effect, and low nuclear transgene expression are major drawbacks for recombinant protein production in this model system. To unveil the molecular changes following transgene insertion, retention, and expression in this species, we genetically engineered C. reinhardtii wild type strain 137c (strain cc-125 mt+) to express the fluorescent protein mVenus and subsequently analysed its intracellular proteome.
The obtained transgenic cell lines showed differences in abundance in more than 400 proteins, with multiple pathways altered post-transformation. Proteins involved in chromatin remodelling, translation initiation and elongation, and protein quality control and transport were found in lower abundance. On the other hand, ribosomal proteins showed higher abundance, a signal of ribosomal stress response.
These results provide new insights into the modifications of C. reinhardtii proteome after transformation, highlighting possible pathways involved in gene silencing. Moreover, this study identifies multiple protein targets for future genetic engineering approaches to improve the prospective use of C. reinhardtii as cell biofactory for industrial applications.
莱茵衣藻正逐渐成为生产重组蛋白的一种很有前途的表达系统。然而，其作为细胞生物工厂的性能仍不理想，尤其是在异源基因的稳定表达方面。基因沉默机制、位置效应和低核转基因表达是该模型系统中重组蛋白生产的主要缺点。为了揭示转基因插入、保留和表达后该物种的分子变化，我们对莱茵衣藻野生型 137c 菌株（cc-125 mt+ 品系）进行了基因工程改造，使其表达荧光蛋白 mVenus，随后分析了其细胞内蛋白质组。
获得的转基因细胞系中，超过 400 种蛋白的丰度存在差异，转化后多个途径发生改变。涉及染色质重塑、翻译起始和延伸以及蛋白质质量控制和运输的蛋白丰度较低。另一方面，核糖体蛋白的丰度较高，这是核糖体应激反应的信号。
这些结果为莱茵衣藻转化后蛋白质组的修饰提供了新的见解，突出了可能参与基因沉默的途径。此外，本研究确定了多个蛋白质靶标，可用于未来的遗传工程方法，以提高莱茵衣藻作为工业应用的细胞生物工厂的预期用途。
