Molecular mechanism for sphingosine-induced Pseudomonas ceramidase expression through the transcriptional regulator SphR

Nozomu Okino, Makoto Ito

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

Pseudomonas aeruginosa, an opportunistic, but serious multidrug-resistant pathogen, secretes a ceramidase capable of cleaving the N-acyl linkage of ceramide to generate fatty acids and sphingosine. We previously reported that the secretion of P. aeruginosa ceramidase was induced by host-derived sphingolipids, through which phospholipase C-induced hemolysis was significantly enhanced. We herein investigated the gene(s) regulating sphingolipid-induced ceramidase expression and identified SphR, which encodes a putative AraC family transcriptional regulator. Disruption of the sphR gene in P. aeruginosa markedly decreased the sphingomyelin-induced secretion of ceramidase, reduced hemolytic activity, and resulted in the loss of sphingomyelin-induced ceramidase expression. A microarray analysis confirmed that sphingomyelin significantly induced ceramidase expression in P. aeruginosa. Furthermore, an electrophoretic mobility shift assay revealed that SphR specifically bound free sphingoid bases such as sphingosine, dihydrosphingosine, and phytosphingosine, but not sphingomyelin or ceramide. A β-galactosidase-assisted promoter assay showed that sphingosine activated ceramidase expression through SphR at a concentration of 100 nM. Collectively, these results demonstrated that sphingosine induces the secretion of ceramidase by promoting the mRNA expression of ceramidase through SphR, thereby enhancing hemolytic phospholipase C-induced cytotoxicity. These results facilitate understanding of the physiological role of bacterial ceramidase in host cells.

Original languageEnglish
Article number38797
JournalScientific reports
Volume6
DOIs
Publication statusPublished - Dec 12 2016

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Ceramidases
Sphingosine
Pseudomonas
Sphingomyelins
Pseudomonas aeruginosa
Sphingolipids
phytosphingosine
Ceramides
Type C Phospholipases
AraC Transcription Factor
Galactosidases
Electrophoretic Mobility Shift Assay
Microarray Analysis
Hemolysis
Genes

All Science Journal Classification (ASJC) codes

  • General

Cite this

Molecular mechanism for sphingosine-induced Pseudomonas ceramidase expression through the transcriptional regulator SphR. / Okino, Nozomu; Ito, Makoto.

In: Scientific reports, Vol. 6, 38797, 12.12.2016.

Research output: Contribution to journalArticle

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