Défense de thèse de doctorat en sciences biologiques "erythritol catabolic pathway"

Revision of the erythritol catabolic pathway and of the central metabolism of the pathogen Brucella

Jury

Emile VAN SCHAFTINGEN (UCL), Ignacio MORIYON (Univ. Navarra, Espagne), Olivier NEYROLLE (Univ. Toulouse 3, France), Michel JADOT, président (UNamur), Jean-Jacques LETESSON, promoteur (UNamur)

Résumé

Colonization of mammal hosts by bacterial pathogens requires various adaptation 
mechanisms. One of these, is the fine-tuning of bacterial metabolism according 
to nutrients availability and physical conditions (O_2 , pH, …) that are 
encountered during the infection. Each colonized niche is likely to be 
characterized by specific physico-chemical properties imposing a metabolic 
plasticity to bacterial pathogens.

/Brucella spp. /are facultative intracellular pathogens, which manage to adapt, 
feed and multiply in various mammal hosts, in different organs and tissues, as 
well as in diverse cell-types either professional or non-professional 
phagocytes. Despite this impressive adaptability, little is known on the 
metabolism of these bacteria, especially /in vivo/. Current understanding of 
/Brucella/ metabolism can be summarized as: (i) hexose metabolism is based on 
the pentose phosphate pathway with the TCA cycle as the glycolysis is 
interrupted and the Entner-Doudoroff pathway does not appear to be active; (ii) 
erythritol is a preferential carbon source and is catabolized in 5 steps to 
produce dihydroxyacetone phosphate. Interestingly, this polyol might represent a 
relevant carbon source /in vivo /as it is found in substantial amounts in the 
reproductive tracts of some hosts for which these bacteria have a particular 
tropism. During our investigation on /Brucella spp./ metabolism/, /data 
inconsistent with these models were quickly gathered, however, leading us to 
re-examine what was thought to be known.

First, we have revisited the erythritol catabolic pathway in /B. abortus /2308, 
which was described nearly 40 years ago. We identified and purified five 
proteins that were required for bacterial growth on this polyol as sole carbon 
source. We have demonstrated that these five enzymes work sequentially for the 
conversion of erythritol to erythrose-4-P, a metabolite of the pentose phosphate 
pathway and not to dihydroxyacetone phosphate unlike previously considered. By 
doing so, four reactions out of the five initially described were revised and 
include three new enzymatic reactions. In parallel, we investigated the 
relevancy of erythritol as carbon source during the infection. Our results 
demonstrate that erythritol catabolism is not required for the infection of RAW 
264.7 murine macrophages, human THP-1 macrophage-like cells or BeWo and JEG-3 
human trophoblasts. In the latter cells, we nonetheless demonstrate that the 
polyol is present in substantial amount and is in all likelihood used by 
intracellular bacteria.

Second, we provide isotopic and genetic evidences that the catabolism of glucose 
in /B. suis /bv.5 is not based on the pentose phosphate pathway but rather on 
the Entner-Doudoroff pathway in chemically defined medium. Our data demonstrate 
that almost 70% of the assimilated glucose is indeed catabolized by this pathway 
while only the 30 remaining % are metabolized by the pentose phosphate pathway. 
We also provide first evidences that the TCA cycle operates in an oxidative 
manner and is supplied by both acetyl-CoA and oxaloacetate. Such EDP-dependent 
glucose catabolism appears to be a feature common to several phylogenetically 
close a-proteobacteria. Finally, it appears that glucose and hexose metabolisms 
do not play a major role for bacterial physiology during the infection of RAW 
264.7 macrophages.

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