Regulatory Activities of Four ArsR Proteins in Agrobacterium tumefaciens 5A
Regulatory Activities of Four ArsR Proteins in Agrobacterium tumefaciens 5A
复制标题
DOI：
10.1128/aem.00262-16
复制
发表时间：
2016-06-01
影响因子：
4.4
通讯作者：
McDermott, Timothy R.
中科院分区：
文献类型：
--
作者：
Kang, Yoon-Suk;Brame, Keenan;McDermott, Timothy R.
ArsR 是一种经过充分研究的转录抑制因子，可调节微生物与砷的相互作用。大多数微生物都有一个 arsR 基因，但在存在多个拷贝的情况下，各自的作用或潜在的功能重叠尚未被探索。我们检查了根癌农杆菌 5A 中由 arsR1 和 arsR2（ars1 操纵子）以及 arsR3 和 arsR4（ars2 操纵子）编码的阻遏蛋白。 ArsR1 和 ArsR4 的一级序列非常相似，并且在系统发育上与 ArsR2 和 ArsR3 不同，而 ArsR2 和 ArsR3 也彼此非常相似。 arsR1、arsR2 和 arsR4 的报告基因构建体 (lacZ) 均可被 As(III) 诱导，但 arsR3 的表达（通过逆转录酶 PCR 监测）不受 As(III) 影响，并且似乎在转录上与上游 lysR 型基因相关。使用缺失突变和其他报告基因测定相结合的实验表明，编码的阻遏蛋白（i）并不像通常已知的ArsR蛋白那样都是自动调节的，（ii）表现出对彼此编码基因的可变控制，以及（iii）对先前显示在ArsR1控制下的其他基因施加可变控制。此外，ArsR2、ArsR3 和 ArsR4 似乎对一些被 ArsR1 抑制的基因具有类似激活剂的功能，这偏离了 ArsR 蛋白经过充分研究的抑制作用。差异化的调控活动表明存在一个以前在 ArsR 研究中未观察到的复杂调控网络。结果表明，重复调节蛋白的精细ArsR序列偏差显然会转化为不同的调节作用。 重要性鉴于ArsR阻遏物在调节微生物砷相互作用的各个方面的重要性，当微生物携带多个arsR拷贝时，评估潜在的调节重叠和/或干扰非常重要。本研究探讨了这个问题，并表明根癌农杆菌 5A 中的四个 arsR 基因与两个独立的 ars 操纵子相关，编码的蛋白质在自动调节和交叉调节以及其他功能基因的控制方面表现出不同程度的功能重叠。在某些情况下，调节活性的差异仅与蛋白质一级结构的有限差异相关。本文总结的实验还提供了证据，表明 ArsR 蛋白似乎具有激活剂功能，代表了 ArsR 的新调节活性，而此前人们只知道 ArsR 是一种阻遏蛋白。
ArsR is a well-studied transcriptional repressor that regulates microbe-arsenic interactions. Most microorganisms have an arsR gene, but in cases where multiple copies exist, the respective roles or potential functional overlap have not been explored. We examined the repressors encoded by arsR1 and arsR2 (ars1 operon) and by arsR3 and arsR4 (ars2 operon) in Agrobacterium tumefaciens 5A. ArsR1 and ArsR4 are very similar in their primary sequences and diverge phylogenetically from ArsR2 and ArsR3, which are also quite similar to one another. Reporter constructs (lacZ) for arsR1, arsR2, and arsR4 were all inducible by As(III), but expression of arsR3 (monitored by reverse transcriptase PCR) was not influenced by As(III) and appeared to be linked transcriptionally to an upstream lysR-type gene. Experiments using a combination of deletion mutations and additional reporter assays illustrated that the encoded repressors (i) are not all autoregulatory as is typically known for ArsR proteins, (ii) exhibit variable control of each other's encoding genes, and (iii) exert variable control of other genes previously shown to be under the control of ArsR1. Furthermore, ArsR2, ArsR3, and ArsR4 appear to have an activator-like function for some genes otherwise repressed by ArsR1, which deviates from the well-studied repressor role of ArsR proteins. The differential regulatory activities suggest a complex regulatory network not previously observed in ArsR studies. The results indicate that fine-scale ArsR sequence deviations of the reiterated regulatory proteins apparently translate to different regulatory roles.IMPORTANCEGiven the significance of the ArsR repressor in regulating various aspects of microbe-arsenic interactions, it is important to assess potential regulatory overlap and/or interference when a microorganism carries multiple copies of arsR. This study explores this issue and shows that the four arsR genes in A. tumefaciens 5A, associated with two separate ars operons, encode proteins exhibiting various degrees of functional overlap with respect to autoregulation and cross-regulation, as well as control of other functional genes. In some cases, differences in regulatory activity are associated with only limited differences in protein primary structure. The experiments summarized herein also present evidence that ArsR proteins appear to have activator functions, representing novel regulatory activities for ArsR, previously known only to be a repressor.