The Small Protein HemP Is a Transcriptional Activator for the Hemin Uptake Operon in Burkholderia multivorans ATCC 17616
The Small Protein HemP Is a Transcriptional Activator for the Hemin Uptake Operon in Burkholderia multivorans ATCC 17616
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DOI：
10.1128/aem.00479-17
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发表时间：
2017-08-01
影响因子：
4.4
通讯作者：
Tsuda, Masataka
中科院分区：
文献类型：
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作者：
Sato, Takuya;Nonoyama, Shouta;Tsuda, Masataka
铁和血红素在细菌的各种代谢功能中起着非常重要的作用，并且它们的细胞内稳态得以维持，因为这些分子的高浓度游离形式极大地促进了芬顿反应介导的大量活性氧物质的产生，这些活性氧物质严重地破坏各种生物分子。多食伯克霍尔德氏菌ATCC 17616的铁摄取调节因子（Fur）是一种铁响应性的全局转录调节因子，其Fur缺失体表现出多效性表型。在这项研究中，我们发现毛皮缺失者的表型受到hemP额外突变的抑制。铁缺乏条件下，血红素氧合酶的转录受毛皮的负调控，在毛皮缺失者中是组成性的。生长的hemP deletant严重受损，在培养基中含有氯化血红素作为唯一的铁源，证明了重要的作用，HemP氯化血红素的利用。HemP需要作为转录激活因子，其特异性结合含启动子的区域上游的毛皮抑制hmuRSTUV操纵子，其编码用于氯化血红素摄取的蛋白质。hmuR缺失体仍然能够使用氯化血红素作为唯一铁源生长，尽管速率明显低于野生型菌株。这些结果强烈表明（i）HmuR参与氯化血红素摄取，以及（ii）ATCC 17616中存在至少部分其他未知的氯化血红素摄取系统，其表达取决于HemP功能。我们的体外分析还表明，高亲和力结合血红素血红素，这样的属性可能会调节转录激活的hmu operon.IMPORTANCE虽然hmuRSTUV基因的利用血红素作为唯一的铁源已被确定在一些伯克霍尔德氏菌菌株，这些基因的调控表达仍然未知。我们在这项研究中使用B.多噬菌ATCC 17616表明其HemP蛋白是hmuRSTUV操纵子表达所需的，并且据我们所知，在本研究中首次阐明了HemP在β-变形菌属中的作用。HemP蛋白还被发现有两个额外的属性，尚未报告的功能同源物在其他物种;一个是，HemP是能够结合到hmu操纵子的启动子包含区域直接激活其转录，另一个是，HemP也需要一个未知的氯化血红素摄取系统的表达。
Iron and heme play very important roles in various metabolic functions in bacteria, and their intracellular homeostasis is maintained because high concentrations of free forms of these molecules greatly facilitate the Fenton reaction-mediated production of large amounts of reactive oxygen species that severely damage various biomolecules. The ferric uptake regulator (Fur) from Burkholderia multivorans ATCC 17616 is an iron-responsive global transcriptional regulator, and its fur deletant exhibits pleiotropic phenotypes. In this study, we found that the phenotypes of the fur deletant were suppressed by an additional mutation in hemP. The transcription of hemP was negatively regulated by Fur under iron-replete conditions and was constitutive in the fur deletant. Growth of a hemP deletant was severely impaired in a medium containing hemin as the sole iron source, demonstrating the important role of HemP in hemin utilization. HemP was required as a transcriptional activator that specifically binds the promoter-containing region upstream of a Fur-repressive hmuRSTUV operon, which encodes the proteins for hemin uptake. A hmuR deletant was still able to grow using hemin as the sole iron source, albeit at a rate clearly lower than that of the wild-type strain. These results strongly suggested (i) the involvement of HmuR in hemin uptake and (ii) the presence in ATCC 17616 of at least part of other unknown hemin uptake systems whose expression depends on the HemP function. Our in vitro analysis also indicated high-affinity binding of HemP to hemin, and such a property might modulate transcriptional activation of the hmu operon.IMPORTANCE Although the hmuRSTUV genes for the utilization of hemin as a sole iron source have been identified in a few Burkholderia strains, the regulatory expression of these genes has remained unknown. Our analysis in this study using B. multivorans ATCC 17616 showed that its HemP protein is required for expression of the hmuRSTUV operon, and the role of HemP in betaproteobacterial species was elucidated for the first time, to our knowledge, in this study. The HemP protein was also found to have two additional properties that have not been reported for functional homologues in other species; one is that HemP is able to bind to the promotercontaining region of the hmu operon to directly activate its transcription, and the other is that HemP is also required for the expression of an unknown hemin uptake system.