2. Institute of Clinical Physiology-CNR, Pisa, Italy.
3. Laboratory of Microbial and Molecular Evolution, Department of Evolutionary Biology,University of Florence, Florence, Italy.
4. Department of Biology and Biotechnology L. Spallanzani, Pavia, Italy.
Background: The Burkholderia cenocepacia J2315 is Gram-negative bacterium that is a pathogen for cystic fibrosis (CF) patients. It displays a high-level of resistance to most antimicrobial drugs. In Gram-negative bacteria, the Resistance- Nodulation-Cell Division (RND) transporter family has a poorly understood role in multidrug resistance. In a previous publication we analysed the RND-4 and RND-9 transporters by microarray analysis. The obtained results suggested that only RND-4 contributes to the antibiotic resistance. The aim of this study was to investigate the role of this efflux transporter from a proteomic point of view.
Methods: We quantitatively compared the intracellular proteome of the deletion mutant B. cenocepacia J2315 impaired in RND-4 transporter (BCAL2820-22) with that of the wild type strain using two-dimensional electrophoresis.
Results: The results pointed out 70 differentially expressed proteins, of which 49 were identified by mass spectrometry. We found that in RND-4 mutant strain, 13 protein spots were up-regulated whilst 35 were down-regulated. One spot was detected only in wild type J2315. Fifty percent of the 35 down-regulated proteins belong to the following functional categories: "amino acids transport and metabolism", "nucleotides transport and metabolism", "lipid transport and metabolism", "translation", "ribosomal structure and biogenesis". Conversely, forty-six percent of 13 the up-regulated proteins belong to the following functional categories: "energy production and conversion", "post-translational modification", "protein turnover, chaperones".
Conclusions: These results indicate that in B. cenocepacia J2315 the RND- 4 gene deletion affects, directly or indirectly, some traits of cell physiology, suggesting for this transporter a wider role than just in drug resistance.
Keywords: Burkholderia cenocepacia J2315, Cystic Fibrosis, RND efflux pumps, two- dimensional electrophoresis, mass spectrometry