通过下调副酸乳杆菌中的 ackA,揭示 PhoP 主调节因子在亚砷酸盐抗性中的作用

IF 4.8 Q1 MICROBIOLOGY
Daniela Corrales , Cristina Alcántara , Dinoraz Vélez , Vicenta Devesa , Vicente Monedero , Manuel Zúñiga
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引用次数: 0

摘要

在细菌中,双组分系统PhoPR在调节许多与磷酸盐摄取和代谢相关的基因中起重要作用。在副干酪乳杆菌中,反应调节因子PhoP的失活导致对亚砷酸盐的抗性增加[As(III)]。比较转录组学分析显示PhoP的缺失对转录组有很强的影响,约占Lc的57.5%。Paracasei基因存在差异表达,但只有92个上调基因和23个下调基因达到了大于2倍的变化。其中,编码ABC膦酸转运蛋白的phnDCEB簇和编码乙酸激酶基因ackA (LCABL_01600)的phnDCEB簇分别下调了10倍和7倍。与选定的PhoP调控基因的体外结合实验表明,PhoP的磷酸化刺激其与pstS(磷酸ABC转运体结合亚基)、phnD和glnA谷氨酰胺合成酶的启动子区域结合,而未检测到与poxL(丙酮酸氧化酶)或ackA推定启动子区域的结合。这一结果首次在乳杆菌科物种中鉴定出三个属于Pho调控子的基因/操纵子。转录组学分析中获得的reads图谱显示,在PhoP突变体中,ackA的转录在ackA编码区出现发夹结构后严重减少。phnD的失活不影响对As(III)的抗性,而ackA的失活导致与PhoP突变体相同水平的抗性。这些发现强烈提示PhoP突变体As(III)的抗性是由于ackA的下调。讨论了可能的作用机制。
本文章由计算机程序翻译,如有差异,请以英文原文为准。

Unveiling the role of the PhoP master regulator in arsenite resistance through ackA downregulation in Lacticaseibacillus paracasei

Unveiling the role of the PhoP master regulator in arsenite resistance through ackA downregulation in Lacticaseibacillus paracasei
In bacteria, the two-component system PhoPR plays an important role in regulating many genes related to phosphate uptake and metabolism. In Lacticaseibacillus paracasei inactivation of the response regulator PhoP results in increased resistance to arsenite [As(III)]. A comparative transcriptomic analysis revealed that the absence of PhoP has a strong effect on the transcriptome, with about 57.5 % of Lc. paracasei genes being differentially expressed, although only 92 of the upregulated genes and 23 of the downregulated genes reached a fold change greater than 2. Among them, the phnDCEB cluster, encoding a putative ABC phosphonate transporter and the acetate kinase encoding gene ackA (LCABL_01600) were downregulated tenfold and sevenfold, respectively. In vitro binding assays with selected PhoP-regulated genes showed that phosphorylation of PhoP stimulated its binding to the promoter regions of pstS (phosphate ABC transporter binding subunit), phnD and glnA glutamine synthetase) whereas no binding to the poxL (pyruvate oxidase) or ackA putative promoter regions was detected. This result identified for the first time three genes/operons belonging to the Pho regulon in a Lactobacillaceae species. Mapping of the reads obtained in the transcriptomic analysis revealed that transcription of ackA was severely diminished in the PhoP mutant after a hairpin structure located within the ackA coding region. Inactivation of phnD did not affect As(III) resistance whereas inactivation of ackA resulted in the same level of resistance as that observed in the PhoP mutant. These finding strongly suggests that PhoP mutant As(III) resistance is due to downregulation of ackA. Possible mechanisms of action are discussed.
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来源期刊
Current Research in Microbial Sciences
Current Research in Microbial Sciences Immunology and Microbiology-Immunology and Microbiology (miscellaneous)
CiteScore
7.90
自引率
0.00%
发文量
81
审稿时长
66 days
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