Relationship of the CreBC two-component regulatory system and inner membrane protein CreD with swimming motility in Stenotrophomonas maltophilia

Hsin-Hui Huang; Wei-Ching Chen; Cheng-Wen Lin; Yi-Tsung Lin; Hsiao-Chen Ning; Yi-Chih Chang; Tsuey-Ching Yang

doi:10.1371/journal.pone.0174704

Relationship of the CreBC two-component regulatory system and inner membrane protein CreD with swimming motility in Stenotrophomonas maltophilia

PLoS One. 2017 Apr 24;12(4):e0174704. doi: 10.1371/journal.pone.0174704. eCollection 2017.

Authors

Hsin-Hui Huang¹, Wei-Ching Chen^{1

2}, Cheng-Wen Lin³, Yi-Tsung Lin^{4

5}, Hsiao-Chen Ning^{6

7}, Yi-Chih Chang³, Tsuey-Ching Yang¹

Affiliations

¹ Department of Biotechnology and Laboratory Science in Medicine, National Yang-Ming University, Taipei, Taiwan.
² Super Laboratory Co. Ltd., New Taipei City, Taiwan.
³ Department of Medical Laboratory Science and Biotechnology, China Medical University, Taichung, Taiwan.
⁴ Division of Infectious Diseases, Department of Medicine, Taipei Veterans General Hospital, Taipei, Taiwan.
⁵ School of Medicine, National Yang-Ming University, Taipei, Taiwan.
⁶ Department of Laboratory Medicine, Chang Gung Memorial Hospital Linkou Branch, Taoyuan, Taiwan.
⁷ Department of Medical Biotechnology and Laboratory Science, Chang Gung University, Taoyuan, Taiwan.

Abstract

The CreBC two-component system (TCS) is a conserved regulatory system found in Escherichia coli, Aeromonas spp., Pseudomonas aeruginosa, and Stenotrophomonas maltophilia. In this study, we determined how CreBC TCS regulates secreted protease activities and swimming motility using creB, creC, and creBC in-frame deletion mutants (KJΔCreB, KJΔCreC, and KJΔBC) of S. maltophilia KJ. Compared to wild-type KJ, KJΔCreB had a comparable secreted protease activity; however, the secreted protease activities were obviously reduced in KJΔCreC and KJΔBC, suggesting that CreC works together with another unidentified response regulator (not CreB) to regulate secreted protease activity. Single gene inactivation of creB or creC resulted in mutants with an enhanced swimming motility, and this phenotype was exacerbated in a double mutant KJΔBC. To elucidate the underlying mechanism responsible for the ΔcreBC-mediated swimming enhancement, flagella morphology observation, RNA-seq based transcriptome assay, qRT-PCR, and membrane integrity and potential assessment were performed. Flagella morphological observation ruled out the possibility that swimming enhancement was due to altered flagella morphology. CreBC inactivation upregulated the expression of creD and flagella-associated genes encoding the basal body- and motor-associated proteins. Furthermore, KJΔBC had an increased membrane susceptibility to Triton X-100 and CreD upregulation in KJΔBC partially alleviated the compromise of membrane integrity. The impact of creBC TCS on bacterial membrane potential was assessed by carbonyl cyanide m-chlorophenyl hydrazine (CCCP50) concentration at which 50% of bacterial swimming is inhibited. CCCP50 of wild-type KJ increased when creBC was deleted, indicating an association between the higher membrane potential of KJΔBC cells and enhanced motility. Upregulation of the basal body- and motor-associated genes of flagella in KJΔBC cells may explain the increased membrane potential. Collectively, inactivation of creBC increased swimming motility through membrane potential increase and creD upregulation in S. maltophilia. The increased membrane potential may supply more energy for flagella propelling and CreD upregulation supports membrane stability, providing a strong membrane for flagellum function.

MeSH terms

Bacterial Proteins / genetics
Bacterial Proteins / metabolism*
Flagella / metabolism*
Gene Expression Regulation, Bacterial*
Stenotrophomonas maltophilia / genetics
Stenotrophomonas maltophilia / metabolism*
Swimming

Substances

Bacterial Proteins

Grants and funding

This work was supported by grant MOST 104-2320-B-010-023-MY3 from Ministry of Science and Technology of Taiwan and grant 40419001 from Professor Tsuei-Chu Mong Merit Scholarship. The authors acknowledge the High-throughput Genome and Big Data Analysis Core Facility of National Core Facility Program for Biotechnology, Taiwan (MOST 104-2319-B-010-001), for sequencing. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.