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AMB Express
2019 Dec 05;91:197. doi: 10.1186/s13568-019-0917-z.
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Effect of the luxI/R gene on AHL-signaling molecules and QS regulatory mechanism in Hafnia alvei H4.
Li X
,
Zhang G
,
Zhu Y
,
Bi J
,
Hao H
,
Hou H
.
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Hafnia alvei H4 is a bacterium subject to regulation by a N-acyl-l-homoserine lactone (AHL)-mediated quorum sensing system and is closely related to the corruption of instant sea cucumber. Studying the effect of Hafnia alvei H4 quorum sensing regulatory genes on AHLs is necessary for the quality and preservation of instant sea cucumber. In this study, the draft genome of H. alvei H4, which comprises a single chromosome of 4,687,151 bp, was sequenced and analyzed and the types of AHLs were analyzed employing thin-layer chromatography (TLC) and high resolution triple quadrupole liquid chromatography/mass spectrometry (LC/MS). Then the wild-type strain of H. alvei H4 and the luxI/R double mutant (ΔluxIR) were compared by transcriptome sequencing (RNA-seq). The results indicate that the incomplete genome sequence revealed the presence of one quorum-sensing (QS) gene set, designated as lasI/expR. Three major AHLs, N-hexanoyl-L-homoserine lactone (C6-HSL), N-butyryl-L-homoserine lactone (C4-HSL), and N-(3-oxo-octanoyl)-L-homoserine lactone (3-oxo-C8-HSL) were found, with C6-HSL being the most abundant. C6-HSL was not detected in the culture of the luxI mutant (ΔluxI) and higher levels of C4-HSL was found in the culture of the luxR mutant (ΔluxR), which suggested that the luxR gene may have a positive effect on C4-HSL production. It was also found that AHL and QS genes are closely related in the absence of luxIR double deletion. The results of this study can further elucidate at the genetic level that luxI and luxR genes are involved in the regulation of AHL.
Fig. 1. TLC bioassay of AHLs produced by H. alvei H4 wt and ΔluxR with C. violaceum CV026 as a sensor strain. Lane 1: C4-HSL standard AHL; Lane 2: C6-HSL standard AHL; Lane 3–Lane 9: 0–36 h every 6 h of AHL extracted from H. alvei H4 wild type and mutant strain culture supernatant
Fig. 2. The AHL extracts of wild-type H. alvei H4 and its ΔluxI,ΔluxR,ΔluxIR mutants at different times were detected by LC/MS
Fig. 3. Arrows indicate the direction of transcription of the gene. The number below the gene arrow is the locus tag corresponding to the genome. Different colors represent the predictive function of different genes. LuxI family AHL synthase gene: lasI (gene0750), brown; luxR family transcription factor gene: expR (gene0751), gray; another LuxR family transcription factor gene “solo R”: gene2065, pink
Fig. 4. KEGG “quorum sensing” pathway based on RNA high-throughput sequencing analysis. Proteins appearing in the green boxes are down-regulated at the mRNA level
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