Study of the putative role of HIF-1alpha and BNIP3L in the biology of mitochondria in Brucella abortus-infected cells

Abstract

Brucella abortus are facultative intracellular pathogen, causing Brucellosis, known for their interactions and the subversion of multiple organelles in infected host cells, including mitochondria. The mitochondrion is a dynamic organelle that plays a central role in the cell, involving ATP production, cellular immunity, many syntheses and degradation pathways, regulation of calcium homeostasis and the integration of life-or-death signals. These important roles of mitochondria in the regulation of many biological processes also make the mitochondria an important target for pathogens. B. abortus is able to corrupt several functions of the mitochondria to its advantage. Indeed, a glycolytic shift, the fragmentation of the mitochondrial population and induction of mitophagy are observed in both myeloid and non-myeloid B. abortus-infected cells. More recently, in the host laboratory, a nuclear accumulation of HIF-1α correlated with Parkin translocation have been observed in Brucella-infected HeLa cells or macrophages. In addition, at 48 h post-infection (p.i) time, the induction of mitophagy correlates with the expression of the HIF-1α-target gene encoding BNIP3L was demonstrated.
The major aim of the master thesis was therefore to better understand the putative role of these molecular actors/effectors (HIF-1α, Parkin and BNIP3L) in the induction of mitochondrial fragmentation and/or mitophagy in B. abortus-infected HeLa and macrophages.
In this work, we observed a decrease in CFU numbers in macrophages in which HIF-1α is stabilised by a chemical hypoxia (CoCl2 treatment) when the stabilisation of the transcription factor occurs prior to or during the early phase of infection. These observations suggest that the entry and/or survival of the bacteria may be disrupted even if the response is not observed for HeLa infected cells. We also show that the translocation of Parkin in the nucleus of Brucella-infected cells is not a robust phenotype while a knock-down of BNIP3L expression in B. abortus-infected HeLa cells prevent, at least partially, but significantly, the fragmentation of mitochondria observed in Brucella-infected cells. A protective effect of BNIP3L expression silencing was also observed on B. abortus-induced mitophagy as assessed by a decreased co-localisation between LC3 and TOM20 in B. abortus-infected HeLa cells. Finally, we focused on the erUPR response potentially involved in the upregulation of autophagic/mitaphagic pathways. We found that activation of the IRE1 pathway occurs in HeLa cells infected with B. abortus at 24 and 48 h p.i. However, the use of siRNA directed against IRE1 did not have any effect on the induction of BNIP3L nor on mitochondrial fragmentation induced by B. abortus in HeLa cells.
In conclusion, in this work, we showed that the mitochondrial fragmentation and mitophagy triggered by Brucella during its intracellular trafficking in the host cells might be mediated by BNIP3L but not Parkin.

Date of Award	Jan 2022
Original language	English
Awarding Institution	University of Namur
Supervisor	Thierry Arnould (Supervisor) & Xavier De Bolle (Co-Supervisor)