One ligand, two regulators and three binding sites: How KDPG controls primary carbon metabolism in Pseudomonas.
One ligand, two regulators and three binding sites: How KDPG controls primary carbon metabolism in Pseudomonas.
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DOI：
10.1371/journal.pgen.1006839
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发表时间：
2017-06
期刊：
影响因子：
4.5
通讯作者：
Malone JG
中科院分区：
文献类型：
--
作者：
Campilongo R;Fung RKY;Little RH;Grenga L;Trampari E;Pepe S;Chandra G;Stevenson CEM;Roncarati D;Malone JG
初级碳代谢的有效调节对于细菌成功适应不同的环境至关重要。我们已经确定了一个未表征的转录调控因子; RccR，控制这一过程中响应碳源的可用性。在植物相关微生物荧光假单胞菌中破坏rccR会抑制其在限定培养基中的生长，并损害其在小麦根际定殖的能力。在结构上，RccR与Entner-Doudoroff（艾德）途径调节剂HexR几乎相同，并且两种蛋白质都由相同的ED中间体2-酮基-3-脱氧-6-磷酸葡萄糖酸（KDPG）控制。尽管有这些相似之处，HexR和RccR控制着初级代谢的完全不同的方面，RccR调节丙酮酸代谢（aceEF）、乙醛酸分流（aceA、glcB、pntAA）和丙酮酸生成（pckA、gap）。RccR表现出复杂和不寻常的调节行为;根据可用的碳源，在丙酮酸代谢和乙醛酸分流/乙醛酸生成位点之间切换阻遏。这种调控复杂性是由两个不同的伪回文结合位点实现的，其不同之处仅在于其接头区的长度，KDPG结合增加了对28 bp aceA结合位点的亲和力，但降低了对15 bp aceE位点的亲和力。因此，RccR能够同时抑制和激活基因表达，以响应碳源的可用性。RccR和HexR调节剂一起使初级碳代谢的多个方面能够快速协调，以响应单个关键中间体的水平。在这里，我们展示了假单胞菌如何控制多个不同的主要碳代谢途径的传感水平的KDPG，Entner杜氏（艾德）途径中间体。KDPG与两种高度相似的转录因子结合;艾德调节因子HexR和先前未表征的蛋白质RccR。RccR反向控制乙醛酸分流、丙酮酸生成和丙酮酸代谢，在丙酮酸代谢基因表达时抑制前两种途径，反之亦然。这种复杂的调节是由RccR调节子启动子中的两个不同的RccR结合共有序列实现的。KDPG结合同时增加RccR对乙醛酸分流和丙酮酸生成启动子的亲和力，并释放丙酮酸代谢的抑制。这种优雅的双调节器电路允许假单胞菌通过感测单个关键中间体KDPG来快速响应碳源的可用性。
Effective regulation of primary carbon metabolism is critically important for bacteria to successfully adapt to different environments. We have identified an uncharacterised transcriptional regulator; RccR, that controls this process in response to carbon source availability. Disruption of rccR in the plant-associated microbe Pseudomonas fluorescens inhibits growth in defined media, and compromises its ability to colonise the wheat rhizosphere. Structurally, RccR is almost identical to the Entner-Doudoroff (ED) pathway regulator HexR, and both proteins are controlled by the same ED-intermediate; 2-keto-3-deoxy-6-phosphogluconate (KDPG). Despite these similarities, HexR and RccR control entirely different aspects of primary metabolism, with RccR regulating pyruvate metabolism (aceEF), the glyoxylate shunt (aceA, glcB, pntAA) and gluconeogenesis (pckA, gap). RccR displays complex and unusual regulatory behaviour; switching repression between the pyruvate metabolism and glyoxylate shunt/gluconeogenesis loci depending on the available carbon source. This regulatory complexity is enabled by two distinct pseudo-palindromic binding sites, differing only in the length of their linker regions, with KDPG binding increasing affinity for the 28 bp aceA binding site but decreasing affinity for the 15 bp aceE site. Thus, RccR is able to simultaneously suppress and activate gene expression in response to carbon source availability. Together, the RccR and HexR regulators enable the rapid coordination of multiple aspects of primary carbon metabolism, in response to levels of a single key intermediate. Here we show how Pseudomonas controls multiple different primary carbon metabolism pathways by sensing levels of KDPG, an Entner Doudoroff (ED) pathway intermediate. KDPG binds to two highly similar transcription factors; the ED regulator HexR and the previously uncharacterised protein RccR. RccR inversely controls the glyoxylate shunt, gluconeogenesis and pyruvate metabolism, suppressing the first two pathways as pyruvate metabolism genes are expressed, and vice versa. This complex regulation is enabled by two distinct RccR-binding consensus sequences in the RccR regulon promoters. KDPG binding simultaneously increases RccR affinity for the glyoxylate shunt and gluconeogenesis promoters, and releases repression of pyruvate metabolism. This elegant two-regulator circuit allows Pseudomonas to rapidly respond to carbon source availability by sensing a single key intermediate, KDPG.