AUTHOR=Wang Zhao-Bao , Li Ya-Qing , Lin Jian-Qun , Pang Xin , Liu Xiang-Mei , Liu Bing-Qiang , Wang Rui , Zhang Cheng-Jia , Wu Yan , Lin Jian-Qiang , Chen Lin-Xu 

TITLE=The Two-Component System RsrS-RsrR Regulates the Tetrathionate Intermediate Pathway for Thiosulfate Oxidation in Acidithiobacillus caldus

JOURNAL=Frontiers in Microbiology

VOLUME=7

YEAR=2016

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2016.01755

DOI=10.3389/fmicb.2016.01755

ISSN=1664-302X

ABSTRACT=<p><italic>Acidithiobacillus caldus</italic> (<italic>A. caldus</italic>) is a common bioleaching bacterium that possesses a sophisticated and highly efficient inorganic sulfur compound metabolism network. Thiosulfate, a central intermediate in the sulfur metabolism network of <italic>A. caldus</italic> and other sulfur-oxidizing microorganisms, can be metabolized via the tetrathionate intermediate (S<sub>4</sub>I) pathway catalyzed by thiosulfate:quinol oxidoreductase (Tqo or DoxDA) and tetrathionate hydrolase (TetH). In <italic>A. caldus</italic>, there is an additional two-component system called RsrS-RsrR. Since <italic>rsrS</italic> and <italic>rsrR</italic> are arranged as an operon with <italic>doxDA</italic> and <italic>tetH</italic> in the genome, we suggest that the regulation of the S<sub>4</sub>I pathway may occur via the RsrS-RsrR system. To examine the regulatory role of the two-component system RsrS-RsrR on the S<sub>4</sub>I pathway, Δ<italic>rsrR</italic> and Δ<italic>rsrS</italic> strains were constructed in <italic>A. caldus</italic> using a newly developed markerless gene knockout method. Transcriptional analysis of the <italic>tetH</italic> cluster in the wild type and mutant strains revealed positive regulation of the S<sub>4</sub>I pathway by the RsrS-RsrR system. A 19 bp inverted repeat sequence (IRS, <underline>AACACCTGT</underline>T<underline>ACACCTGTT</underline>) located upstream of the <italic>tetH</italic> promoter was identified as the binding site for RsrR by using electrophoretic mobility shift assays (EMSAs) <italic>in vitro</italic> and promoter-probe vectors <italic>in vivo</italic>. In addition, Δ<italic>rsrR</italic>, and Δ<italic>rsrS</italic> strains cultivated in K<sub>2</sub>S<sub>4</sub>O<sub>6</sub>-medium exhibited significant growth differences when compared with the wild type. Transcriptional analysis indicated that the absence of <italic>rsrS</italic> or <italic>rsrR</italic> had different effects on the expression of genes involved in sulfur metabolism and signaling systems. Finally, a model of tetrathionate sensing by RsrS, signal transduction via RsrR, and transcriptional activation of <italic>tetH</italic>-<italic>doxDA</italic> was proposed to provide insights toward the understanding of sulfur metabolism in <italic>A. caldus</italic>. This study also provided a powerful genetic tool for studies in <italic>A. caldus</italic>.</p>