黄色链霉菌耐受铁离子的胁迫机制

Iron tolerance mechanism of Streptomyces flavus

摘要:

【背景】 铁是微生物生长与代谢所必需的一种微量元素，在生物体中参与呼吸、DNA前体合成、基因调节及抗氧化应激等重要生理活动。黄色链霉菌(Streptomyces flavus) TRM45540是一株分离自新疆罗布泊的放线菌，前期研究表现出对铁离子具有较强耐受性，探索菌株TRM45540耐受铁离子胁迫的响应机制具有重要意义。【目的】 揭示菌株TRM45540耐受铁离子胁迫的机制。【方法】 以菌株TRM45540为研究对象，通过向ISP4液体培养基中添加不同质量的硫酸亚铁来胁迫菌株TRM45540，并对胁迫后的样本进行转录组测序与分析，探索在铁离子胁迫条件下菌株TRM45540耐受铁离子的响应机制。同时通过扫描电子显微镜(scanning electron microscopy, SEM)和能谱分析观察并检测铁离子胁迫下菌丝表面形态、元素种类及含量变化。【结果】 低硫酸亚铁添加量(1-500 mg/L)和高硫酸亚铁添加量(2 500-4 500 mg/L)均不利于菌株TRM45540生长，在硫酸亚铁添加量为500-2 500 mg/L时可以促进菌株TRM45540生物量的积累。扫描电子显微镜观察到随着硫酸亚铁添加量的增加，菌株TRM45540菌丝体外有大量胞外聚合物(extracellular polymeric substances, EPS)生成；能谱分析结果显示，EPS中氧、铁元素含量升高，氮元素含量降低，推测EPS为多糖；转录组分析结果表明，受Fe(Ⅲ/Ⅱ)胁迫后差异表达基因涉及ABC转运系统、二元信号系统、铁离子运输、氨基酸代谢、谷胱甘肽表达、卟啉类化合物代谢等多个代谢通路。【结论】 菌株TRM45540受到Fe(Ⅲ/Ⅱ)离子胁迫后，触发了生物大分子分泌并输出到细胞表面形成多糖类EPS，菌体表面的EPS络合大量铁离子，阻止铁离子内流。同时触发二元信号系统，启动一系列耐受铁离子胁迫的响应机制，酸性氨基酸大量表达，络合铁离子的卟啉类化合物合成增加，参与解毒的半胱氨酸和谷胱甘肽的生物合成加剧，从而多途径减少高铁离子的伤害。

关键词:铁离子胁迫;黄色链霉菌;转录组分析;能谱分析

Abstract:

[Background] Iron is a microelement essential for the growth and metabolism of microorganisms, playing a role in vital physiological activities like respiration, DNA precursor synthesis, gene regulation, and response to oxidative stress. Studies have confirmed the tolerance of Streptomyces flavus TRM45540, an actinomycete strain isolated from Lop Nur in Xinjiang, to ferric ions. It is of great significance to elucidate the response mechanism of S. flavus TRM45540 to iron stress. [Objective] To reveal the iron tolerance mechanism of S. flavus TRM45540. [Methods] S. flavus TRM45540 was cultured in the ISP4 liquid media supplemented with different levels of ferrous sulfate. Transcriptome sequencing was employed to explore the response mechanism to iron. Additionally, scanning electron microscopy and energy spectrum analysis were employed to observe changes in mycelial surface morphology as well as elemental species and content under ferric ion stress. [Results] The supplementation of ferrous sulfate at low (1-500 mg/L) and high (2 500-4 500 mg/L) levels had negative effects on the growth of S. flavus TRM45540. However, supplementing ferrous sulfate at 500-2 500 mg/L promoted the biomass accumulation of S. flavus TRM45540. With the increase in the supplementing level of ferrous sulfate, a large amount of extracellular polymers substances (EPSs) were generated outside the mycelia of S. flavus TRM45540. The energy spectrum analysis showed increases in the oxygen and iron content and a decrease in the nitrogen content of EPS, suggesting a polysaccharide composition. The transcriptome analysis showed that differentially expressed genes post ferric ion stress were associated with multiple metabolic pathways including the ABC transporter system, two component system, ferric ion transport, amino acid metabolism, glutathione expression, and porphyrin metabolism. [Conclusion] The ferric ion stress induced the secretion and export of biomolecules to the cell surface of S. flavus TRM45540, leading to the formation of polysaccharide-based EPS. The EPS was complexed with a large amount of ferric ions on the bacterial cell surface, thereby impeding the influx of ferric ions. Additionally, the activation of the two component system triggered a cascade of responses to ferric ion stress. This included upregulation of acidic amino acids, enhanced synthesis of porphyrins binding to ferric ions, and increased biosynthesis of cysteine and glutathione involved in ferric ion removal. These pathways coordinated to reduce the cellular damage caused by high ferric ion levels.

Key words:ferric ion stress;Streptomyces flavus;transcriptome analysis;energy spectrum analysis