JIOMICS
JIOMICS Internacional
JIOMICS Internacional
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<strong>JIOMICS</strong> | VOL 5 | ISSUE 2 | DECEMBER 2015 | 1-62<br />
JOURNAL OF INTEGRATED OMICS<br />
Journal of Integrated Omics<br />
A METHODOLOGICAL JOURNAL<br />
HTTP://WWW.<strong>JIOMICS</strong>.COM<br />
Special Issue: Proceeding Abstracts of the 4 th International Congress on Analytical Proteomics (ICAP 2015)<br />
Effect of a quorum sensing inhibitor on the Pseudomonas aeruginosa PAO1<br />
proteome<br />
Shunmugiah Thevar Karutha Pandian*<br />
Department of Biotechnology, Alagappa University, Science Campus, Karaikudi 630 004, India. *Corresponding author:<br />
k_pandian@rediffmail.com<br />
Available Online: 31 December 2015<br />
Abstract<br />
Purpose: Research in the field of quorum sensing (QS) inhibition resulted in the identification of numerous QS inhibitors (QSIs). But<br />
their mode of action and impact on proteome of bacterial pathogens remains unclear. Hence, the aim of the present study is to find out the<br />
molecular mechanism of QSI and its effect on cytoplasmic proteome of P. aeruginosa PAO1.<br />
Experimental description: Cytoplasmic proteins of P. aeruginosa PAO1 grown in the absence and presence of 5 µg of QSI were extracted<br />
by sonication. For 2DE, 600µg each of cytoplasmic proteins from control and treated samples were subjected to isoelectric focusing using immobilized<br />
pH gradient (IPG) strips (18 cm, pH 3-10 NL (GE Healthcare)) at 20°C using standard parameters for 18 h. Then the IPG strips were<br />
reduced and alkylated for 20 min. After this step, strips were placed on 12 to 15% gradient sodium dodecyl sulphate-polyacrylamide gels (Ettan<br />
DALT six, GE Healthcare) for the second dimension and electrophoresis was performed. Protein spots were visualized by colloidal Coomassie<br />
brilliant blue G-250. The gels were scanned with Image scanner III (GE Healthcare) and the raw images were analyzed using ImageMaster 2D<br />
Platinum 7. Experiments were performed in biological triplicates. More than two fold differentially regulated protein spots were selected for<br />
in-gel trypsin digestion and analyzed using MALDI TOF/TOF (AXIMA Performance, Shimadzu.Biotech). The proteins were identified by<br />
peptide mass fingerprinting using the software tool MS-Fit (http://prospector.ucsf.edu).<br />
Results: Among the detected spots (613), 48 (7.8%) spots are downregulated and 31 (5%) spots are upregulated by more than two fold.<br />
Based on statistical significance (ANOVA 0.05), seventeen downregulated and five upregulated protein spots were selected for MALDI TOF/<br />
TOF analysis. Downregulated proteins are involved in iron transport, transcriptional regulation, antibiotic resistance, chemotaxis and two<br />
component signaling systems. QSI treatment upregulated the expression of histidine kinase, ABC-transporter protein, putative transcriptional<br />
regulator, hypothetical proteins G655_17540 and PSPA7_1308.<br />
Conclusions: Since iron transport proteins, two-component response regulators and sensor are involved in the activation of protease, exotoxin<br />
A, and pyoverdin biosynthesis proteins in P. aeruginosa, downregulation of above said proteins by QSI could be a major event in attenuation<br />
of virulence factor production. In addition, QSI also blocked the expression of proteins involved in the pyoveridine and pyochelin synthesis<br />
proteins. Most notably, putative iron-sulfur proteins which are known to express highly in biofilms are significantly downregulated<br />
upon QSI treatment. In conclusion, QSI inhibits the virulence factor production by targeting the iron homeostasis, pyoveridine and pyochelin<br />
biosynthesis pathways.<br />
Keywords: quorum sensing inhibitor; P. aeruginosa PAO1; quorum sensing.<br />
Acknowledgements: The financial support from ICMR and CSIR is thankfully acknowledged. The Instrumentation Facility provided by<br />
DST (FIST & PURSE) and UGC (SAP-DRS1) and Computational/Bioinformatics Facility provided by DBT (BIF) are gratefully acknowledged.<br />
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