Final Report Summary - MONOGAD (Elucidating novel roles and mechanisms of the GAD system in stress resistance and virulence of Listeria monocytogenes.)
Listeria monocytogenes is a bacterial foodborne pathogen that causes listeriosis, a life-threatening disease in humans. It is the most deadly foodborne disease due to its high mortality rates (30-40%) affecting mainly immunocompromised individuals. Elimination of this pathogen relies significantly on acidic conditions, in premises (acid-based disinfectants), in foods (acidic preservatives) and even in our body (acidity in stomach). Therefore, our ability to eliminate L. monocytogenes relies on our understanding of its acid resistance mechanisms of which, the most important is the glutamate decarboxylase (GAD) system. This system operates by consuming protons through the decarboxylation of glutamate (Glu) to γ-aminobutyric acid (GABA). In light of discoveries made by the grant holder such as the existence of the intracellular GAD system (GADi) working on intracellular glutamate in parallel with the extracellular one (GADe) and the existence of unknown compounds that affect the activity and/or transcription of the GAD system in this organism this grant was set to increase our understanding of the function of the GAD system.
Common belief holds that the GABA produced by the GAD system protects cells under acidic conditions through its high buffering capacity. In this work we showed that GABA has similar buffering capacity with glutamate suggesting that the GAD system protects mainly through the consumption of protons. Furthermore, we have shown that GADi contributes to the acid resistance in EGD strain that uses only this system and not GADe. In addition, we demonstrated that in different strains various decarboxylases could contribute to the GADi activity as the GadD3 is the major determinant of GADi in EGD while in 10403S this role is fulfilled by GadD2. We have also investigated various compounds for their ability to activate the GAD system under conditions that do not allow that (in Defined medium). In short, rich sources of aminoacids (casaminoacids, Tryptone Soy Broth, Trypticase or Proteose Peptone) resulted in significant activation of the GAD system when added in Defined Medium through transcription during early stationary phase and not through increased GAD activity. Furthermore, none of various individual aminoacids was able to activate the GAD system in Defined Medium, but a lower level of activation was observed with L-cysteine, L-tryptophane, cyclic di-AMP and NaCl. Overall the work enhanced our understanding of the function of the GAD system.
This grant has contributed greatly to my career integration and scientific independence. As a transferable grant it was seen by the Host Institution as a good complementation to a start-up package. This grant has proven key towards obtaining a permanent position as a lecturer in Food Microbiology at UoR and building my own research group through grants of an overall value of €750,000. The group comprises/ed 7 PhD students (2 more to join shortly) and 4 more PhD students as co-PI while 2 more PhD students and 2 post docs will be joining shortly (1 is jointly supervised). Furthermore, it contributed significantly to collaborations with various companies and other colleagues within UoR and other Universities within and outside the UK.
Common belief holds that the GABA produced by the GAD system protects cells under acidic conditions through its high buffering capacity. In this work we showed that GABA has similar buffering capacity with glutamate suggesting that the GAD system protects mainly through the consumption of protons. Furthermore, we have shown that GADi contributes to the acid resistance in EGD strain that uses only this system and not GADe. In addition, we demonstrated that in different strains various decarboxylases could contribute to the GADi activity as the GadD3 is the major determinant of GADi in EGD while in 10403S this role is fulfilled by GadD2. We have also investigated various compounds for their ability to activate the GAD system under conditions that do not allow that (in Defined medium). In short, rich sources of aminoacids (casaminoacids, Tryptone Soy Broth, Trypticase or Proteose Peptone) resulted in significant activation of the GAD system when added in Defined Medium through transcription during early stationary phase and not through increased GAD activity. Furthermore, none of various individual aminoacids was able to activate the GAD system in Defined Medium, but a lower level of activation was observed with L-cysteine, L-tryptophane, cyclic di-AMP and NaCl. Overall the work enhanced our understanding of the function of the GAD system.
This grant has contributed greatly to my career integration and scientific independence. As a transferable grant it was seen by the Host Institution as a good complementation to a start-up package. This grant has proven key towards obtaining a permanent position as a lecturer in Food Microbiology at UoR and building my own research group through grants of an overall value of €750,000. The group comprises/ed 7 PhD students (2 more to join shortly) and 4 more PhD students as co-PI while 2 more PhD students and 2 post docs will be joining shortly (1 is jointly supervised). Furthermore, it contributed significantly to collaborations with various companies and other colleagues within UoR and other Universities within and outside the UK.