Department of Biological Sciences, Columbia University, New York, New York, USA.
Center for Genomics and Systems Biology, Department of Biology, New York University, New York, New York, USA.
J Bacteriol. 2018 Apr 9;200(9). doi: 10.1128/JB.00031-18. Print 2018 May 1.
Microbes in biofilms face the challenge of substrate limitation. In particular, oxygen often becomes limited for cells in biofilms growing in the laboratory or during host colonization. Previously we found that phenazines, antibiotics produced by , balance the intracellular redox state of cells in biofilms. Here, we show that genes involved in denitrification are induced in phenazine-null (Δ) mutant biofilms grown under an aerobic atmosphere, even in the absence of nitrate. This finding suggests that resident cells employ a bet-hedging strategy to anticipate the potential availability of nitrate and counterbalance their highly reduced redox state. Consistent with our previous characterization of aerobically grown colonies supplemented with nitrate, we found that the pathway that is induced in Δ mutant colonies combines the nitrate reductase activity of the periplasmic enzyme Nap with the downstream reduction of nitrite to nitrogen gas catalyzed by the enzymes Nir, Nor, and Nos. This regulatory relationship differs from the denitrification pathway that functions under anaerobic growth, with nitrate as the terminal electron acceptor, which depends on the membrane-associated nitrate reductase Nar. We identified the sequences in the promoter regions of the and operons that are required for the effects of phenazines on expression. We also show that specific phenazines have differential effects on gene expression. Finally, we provide evidence that individual steps of the denitrification pathway are catalyzed at different depths within aerobically grown biofilms, suggesting metabolic cross-feeding between community subpopulations. An understanding of the unique physiology of cells in biofilms is critical to our ability to treat fungal and bacterial infections. Colony biofilms of the opportunistic pathogen grown under an aerobic atmosphere but without nitrate express a denitrification pathway that differs from that used for anaerobic growth. We report that the components of this pathway are induced by electron acceptor limitation and that they are differentially expressed over the biofilm depth. These observations suggest that (i) exhibits "bet hedging," in that it expends energy and resources to prepare for nitrate availability when other electron acceptors are absent, and (ii) cells in distinct biofilm microniches may be able to exchange substrates to catalyze full denitrification.
生物膜中的微生物面临着基质限制的挑战。特别是，在实验室或宿主定植过程中，生物膜中生长的细胞中的氧气通常会变得有限。我们之前发现，吩嗪类物质是由 产生的抗生素，可以平衡生物膜中细胞的细胞内氧化还原状态。在这里，我们表明，在有氧气氛下生长的吩嗪缺失（Δ）突变体生物膜中诱导了参与反硝化的基因，即使没有硝酸盐也是如此。这一发现表明，常驻细胞采用了一种“赌博”策略来预测硝酸盐的潜在可用性，并平衡其高度还原的氧化还原状态。与我们之前对补充有硝酸盐的有氧生长菌落的特征一致，我们发现，在 Δ 突变菌落中诱导的途径结合了周质酶 Nap 的硝酸盐还原酶活性，以及随后由 Nir、Nor 和 Nos 酶催化的将亚硝酸盐还原为氮气的下游反应。这种调控关系与作为末端电子受体的厌氧生长下的反硝化途径不同，后者依赖于膜相关的硝酸盐还原酶 Nar。我们确定了 和 操纵子启动子区域中对吩嗪类物质对表达影响所需的序列。我们还表明，特定的吩嗪类物质对 基因表达有不同的影响。最后，我们提供了证据表明，有氧生长生物膜中反硝化途径的各个步骤在不同深度处被催化，表明群落亚群之间存在代谢交叉喂养。了解生物膜中细胞的独特生理学对于我们治疗真菌和细菌感染的能力至关重要。在有氧但没有硝酸盐的条件下生长的机会性病原体 的菌落生物膜表达了一种与厌氧生长不同的反硝化途径。我们报告说，该途径的组成部分是由电子受体限制诱导的，并且它们在生物膜深度上差异表达。这些观察结果表明，（i） 表现出“赌博”，即在其他电子受体不存在时，它会消耗能量和资源为硝酸盐的可用性做准备，以及（ii）不同生物膜微生境中的细胞可能能够交换底物以催化完全反硝化。
