Global response of Porphyromonas gingivalis to heme-limitation

Porphyromonas gingivalis is an anaerobic, asaccharolytic Gram-negative bacterium that has essential requirements for both iron and protoporphyrin IX, which it preferentially obtains in the form of heme from hemoglobin. Heme availability has been linked to the virulence of this bacterium. Objectives: To determine changes in the P. gingivalis global pattern of protein and transcript abundances in response to heme-limitation Methods: P. gingivalis W50 was grown in continuous culture in heme-excess and heme–limitation. A combination of large-scale, quantitative proteomic analysis using stable isotope labeling strategies (ICAT) and mass spectrometry together with transcriptomic analysis using custom made DNA microarrays was employed to compare heme-limitation with heme-excess. Results: The transcriptomic analysis identified 160 significantly differentially expressed genes whilst in total 142 proteins were identified by the ICAT analysis, of which 70 exhibited a two fold or greater change in abundance between growth conditions. There was broad agreement between the transcriptomic and proteomic analyses. Both analyses detected an up-regulation of known and putative heme transport systems including the Hmu and Htr systems and enzymes involved in the oxidative stress response including alkyl hydroperoxide reductase. There was an increase in abundance of three proteins believed to be involved in invasion of human epithelial cells. (PG0159, PG0350 and PG1374). The role of an invasin-related protein, PG1374 (IrpI) in cell invasion was confirmed by determining epithelial cell invasion of an isogenic IrpI mutant. There was a coordinated change in the abundances of the enzymes of the aspartate and glutamate catabolic pathways. Conclusion: The differential regulation of a significant number of proteins linked to iron/heme transport, invasion of host cells, oxidative stress response and virulence of the bacterium indicate there is a coordinated response by P. gingivalis to heme availability. This study was funded by the Australian National Health and Medical Research Council.