A Novel DNA-Binding Protein, PhaR, Plays a Central Role in the Regulation of Polyhydroxyalkanoate Accumulation and Granule Formation in the Haloarchaeon Haloferax mediterranei
A Novel DNA-Binding Protein, PhaR, Plays a Central Role in the Regulation of Polyhydroxyalkanoate Accumulation and Granule Formation in the Haloarchaeon Haloferax mediterranei
复制标题
一种新型 DNA 结合蛋白 PhaR 在调节 Haloarchaeon Haloferax mediterranei 的聚羟基脂肪酸酯积累和颗粒形成中发挥核心作用
DOI：
10.1128/aem.02878-14
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发表时间：
2015-01-01
影响因子：
4.4
通讯作者：
Xiang, Hua
中科院分区：
文献类型：
--
作者：
Cai, Shuangfeng;Cai, Lei;Xiang, Hua
摘要聚羟基脂肪酸酯（PHA）是由许多微生物合成并组装成颗粒状的PHA，在许多微生物中形成过程受到很好的调控。然而，这种调节在盐古菌中仍不清楚。在这项研究中，我们确定了一个PHA颗粒相关的调节（PhaR），负调控自己的基因和颗粒结构基因phaP在同一操纵子（phaRP）在地中海盐藻的表达。染色质免疫沉淀-定量PCR（ChIP-qPCR）分析证明PhaR和phaRP启动子之间在体内存在显著的相互作用。phaRP启动子的扫描诱变揭示了一个特定的顺式元件作为PhaR的可能结合位置。PhaR的盐古菌同源物包含一个新的保守结构域，该结构域属于AbrB样蛋白中发现的交换发夹桶折叠家族。氨基酸取代表明，这种AbrB样结构域是至关重要的抑制活性的PhaR。此外，phaRP启动子在PHA阴性菌株中具有较弱的活性，这意味着PHA颗粒在PhaR滴定中的功能。此外，H.缺乏phaR的菌株缺乏PHA积累，并产生具有不规则形状的颗粒。有趣的是，PhaR本身可以以PhaP非依赖性方式促进PHA合成和颗粒形成。总之，我们的研究结果表明，嗜盐古菌PhaR是一种新的双功能蛋白，在调节PHA积累和颗粒形成中起着核心作用。医生。
ABSTRACT Polyhydroxyalkanoates (PHAs) are synthesized and assembled as PHA granules that undergo well-regulated formation in many microorganisms. However, this regulation remains unclear in haloarchaea. In this study, we identified a PHA granule-associated regulator (PhaR) that negatively regulates the expression of both its own gene and the granule structural gene phaP in the same operon (phaRP) in Haloferax mediterranei. Chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR) assays demonstrated a significant interaction between PhaR and the phaRP promoter in vivo. Scanning mutagenesis of the phaRP promoter revealed a specific cis-element as the possible binding position of the PhaR. The haloarchaeal homologs of the PhaR contain a novel conserved domain that belongs to a swapped-hairpin barrel fold family found in AbrB-like proteins. Amino acid substitution indicated that this AbrB-like domain is critical for the repression activity of PhaR. In addition, the phaRP promoter had a weaker activity in the PHA-negative strains, implying a function of the PHA granules in titration of the PhaR. Moreover, the H. mediterranei strain lacking phaR was deficient in PHA accumulation and produced granules with irregular shapes. Interestingly, the PhaR itself can promote PHA synthesis and granule formation in a PhaP-independent manner. Collectively, our results demonstrated that the haloarchaeal PhaR is a novel bifunctional protein that plays the central role in the regulation of PHA accumulation and granule formation in H. mediterranei.