Han Liyuan, Xu Jiaxin, Gu Yuan, Feng Ruiying, Liu Shuo, Lu Qiujie, Lv Shuzhe, Zhang Youming, Yan Fu
State Key Laboratory of Microbial Technology, Shandong University, Qingdao, Shandong 266237, China.
State Key Laboratory of Microbial Technology, Shandong University, Qingdao, Shandong 266237, China.
Int J Biol Macromol. 2025 May;307(Pt 3):142248. doi: 10.1016/j.ijbiomac.2025.142248. Epub 2025 Mar 18.
Genetic manipulation in bacteria, particularly in industrially relevant Streptomyces strains, is often hindered by low DNA transfer efficiency. This study investigates the role of cell envelope components, particularly teichoic acid (TA), in enhancing conjugation efficiency. Using CRISPR-Cpf1 system, we systematically disrupted 26 cell envelope-related genes in Streptomyces coelicolor A3(2), revealing that TA biosynthesis significantly influences DNA uptake. Deletion of SCO1526 (phosphatidylinositol mannoside acyltransferase) and SCO4847 (d-alanyl-d-alanine carboxypeptidase) markedly increased conjugation efficiency, while heterologous expression of TA biosynthetic genes in industrial strains S. hygroscopicus, S. avermitilis and S. venezuelae resulted in a 1300-fold, 4.9-fold and 4.9-fold enhancement, respectively. Strain-specific differences in TA impact probably linked to variations in cell wall structure and TA synthesis capacity. These findings highlight the critical role of TA in bacterial conjugation and offer a robust strategy for improving genetic manipulation in industrially important Streptomyces strains. This work advances our understanding of bacterial genetic tractability and provides a foundation for harnessing the biosynthetic potential of traditionally hard-to-manipulate bacteria.
在细菌中进行基因操作，尤其是在与工业相关的链霉菌菌株中，常常受到DNA转移效率低下的阻碍。本研究调查了细胞壁成分，特别是磷壁酸（TA），在提高接合效率中的作用。利用CRISPR-Cpf1系统，我们系统地破坏了天蓝色链霉菌A3(2)中26个与细胞壁相关的基因，结果表明TA生物合成显著影响DNA摄取。缺失SCO1526（磷脂酰肌醇甘露糖苷酰基转移酶）和SCO4847（D-丙氨酰-D-丙氨酸羧肽酶）可显著提高接合效率，而在工业菌株吸水链霉菌、阿维链霉菌和委内瑞拉链霉菌中异源表达TA生物合成基因，接合效率分别提高了1300倍、4.9倍和4.9倍。TA影响的菌株特异性差异可能与细胞壁结构和TA合成能力的变化有关。这些发现突出了TA在细菌接合中的关键作用，并为提高工业上重要的链霉菌菌株的基因操作提供了一个有力的策略。这项工作增进了我们对细菌遗传易操作性的理解，并为挖掘传统上难以操作的细菌的生物合成潜力奠定了基础。
