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Regulatory Activities of Four ArsR Proteins in Agrobacterium tumefaciens 5A.

ABSTRACT: 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.Given 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.

SUBMITTER: Kang YS

PROVIDER: S-EPMC4959165 | biostudies-literature | 2016 Jun

REPOSITORIES: biostudies-literature

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Regulatory Activities of Four ArsR Proteins in Agrobacterium tumefaciens 5A.

Kang Yoon-Suk YS Brame Keenan K Jetter Jonathan J Bothner Brian B BB Wang Gejiao G Thiyagarajan Saravanamuthu S McDermott Timothy R TR

Applied and environmental microbiology 20160531 12

<h4>Unlabelled</h4>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 fr ...[more]

PMID: 27037117

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Project description:Rotary flagella propel bacteria through liquid and across semisolid environments. Flagella are composed of the basal body that constitutes the motor for rotation, the curved hook that connects to the basal body, and the flagellar filament that propels the cell. Flagellar filaments can be composed of a single flagellin protein, such as in Escherichia coli, or made up of multiple flagellins, such as in Agrobacterium tumefaciens The four distinct flagellins FlaA, FlaB, FlaC, and FlaD produced by wild-type A. tumefaciens are not redundant in function but have specific properties. FlaA and FlaB are much more abundant than FlaC and FlaD and are readily observable in mature flagellar filaments, when either FlaA or FlaB is fluorescently labeled. Cells producing FlaA with any one of the other three flagellins can generate functional filaments and thus are motile, but FlaA alone cannot constitute a functional filament. In flaA mutants that manifest swimming deficiencies, there are multiple ways by which these mutations can be phenotypically suppressed. These suppressor mutations primarily occur within or upstream of the flaB flagellin gene or in the transcription factor sciP regulating flagellin expression. The helical conformation of the flagellar filament appears to require a key asparagine residue present in FlaA and absent in other flagellins. However, FlaB can be spontaneously mutated to render helical flagella in the absence of FlaA, reflecting their overall similarity and perhaps the subtle differences in the specific functions they have evolved to fulfill.IMPORTANCE Flagellins are abundant bacterial proteins comprising the flagellar filaments that propel bacterial movement. Several members of the alphaproteobacterial group express multiple flagellins, in contrast to model systems, such as with Escherichia coli, which has one type of flagellin. The plant pathogen Agrobacterium tumefaciens has four flagellins, the abundant and readily detected FlaA and FlaB, and lower levels of FlaC and FlaD. Mutational analysis reveals that FlaA requires at least one of the other flagellins to function, as flaA mutants produce nonhelical flagella and cannot swim efficiently. Suppressor mutations can rescue this swimming defect through mutations in the remaining flagellins, including structural changes imparting helical shape to the flagella, and putative regulators. Our findings shed light on how multiple flagellins contribute to motility.

Dataset Information

Regulatory Activities of Four ArsR Proteins in Agrobacterium tumefaciens 5A.

Publications

Regulatory Activities of Four ArsR Proteins in Agrobacterium tumefaciens 5A.

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