Oral Presentation Australian Society for Microbiology Annual Scientific Meeting 2019

Autoinducer 2 and Galactose: How Quorum Sensing Dictates Sugar Metabolism in Streptococcus pneumoniae (#167)

Kimberley T McLean 1 , Victoria G Pederick 1 , Lauren J McAllister 1 , Adrienne W Paton 1 , Christopher A McDevitt 2 , Marco Oggioni 3 , James C Paton 1 , Claudia Trappetti 1
  1. Research Centre for Infectious Diseases, University of Adelaide, Adelaide, South Australia, Australia
  2. Department of Microbiology and Immunology, University of Melbourne, Melbourne, Victoria, Australia
  3. Department of Genetics, University of Leicester, Leicester, United Kingdom

Through an unknown mechanism, Streptococcus pneumoniae (the pneumococcus) is able to switch from asymptomatic coloniser to invasive pathogen whilst colonising the nasopharynx. In this niche, cell-to-cell communication is achieved through the coordinated release and sensing of signalling molecules known as autoinducers in a process known as Quorum Sensing (QS). We have focused on characterising the Autoinducer-2 (AI-2)/LuxS QS system and observed that AI-2 QS signalling is dependent upon the membrane bound protein FruA for the import of AI-2, which enables the pneumococcus to utilise galactose as a carbon source by upregulation of the Leloir pathway. We have been investigating whether AI-2 phosphorylates the Leloir pathway regulator GalR after FruA-mediated uptake. GalR is known to possess three potential phosphorylation sites: S317, T319 and T323. Substitution mutants have been generated to either abrogate (A) or constitutively mimic (D or E) phosphorylation at these sites, either singly or in combination. Subsequent growth assays and transcriptional analyses revealed complex phenotypes. Interestingly, galRAAA, galRDDD and galREEE strains fail to grow in Chemically Defined Media with galactose as the sole carbon source, indicating that these sites are essential for galactose utilisation. However, the presence of endogenous intracellular AI-2 may be having an impact on the Leloir pathway, thereby complicating interpretation of mutant phenotypes. This is being addressed by the introduction of a luxS deletion mutation into each of the galR substitution mutants, rendering these strains unable to produce AI-2. Additionally, the membrane-bound protein FruA, the putative AI-2 receptor, has been successfully over-expressed and purified, enabling direct biochemical characterisation of its interaction with AI-2. The outcomes of this study will provide greater understanding of the relationship between intracellular sugar metabolism and cell-to-cell communication within the pneumococcus, parameters that are known to profoundly impact virulence phenotype. Additionally, greater translational outcomes will be delivered by bettering our understanding of how to control bacterial colonisation and interfere with the community-like behaviours of biofilms.