Tioli 2019 MiP2019

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Involvement of mitochondrial respiratory supercomplexes in anti-bacterial innate immunity.

Link: MiP2019

Tioli G, Lenaz G, Falasca AI, Garaude J, Genova ML (2019)

Event: MiP2019

COST Action MitoEAGLE

Metabolic reprogramming has recently emerged as a major feature of innate immune cells following bacterial infection. At the core of this physiological process is the mitochondrion, a bioenergetic organelle that also serves as an immune signalling platform and a critical rheostat for most catabolic processes converging on the mitochondrial electron transport system (ETS) [1]. The dynamic supramolecular organization of the ETS complexes forming the respiratory supercomplexes (SCs) may confer functional advantages to the cells but the precise role of Complex I (NADH:ubiquinone oxidoreductase, CI) within SCs is still little known [2]. Indications exist that organizational adaptations occur in the ETS of macrophages to allow metabolic plasticity during engulfment of live gram-negative bacteria (e.g. Escherichia coli) [3].

The aim of our study is to investigate the role of CI-containing SCs and to characterize the architectural and functional adaptations in cultured bone marrow-derived macrophages (BMDMs) and in isolated mitochondria from C57BL/6N mouse tissues. To determine whether the infection affects mitochondrial respiration, we have approached the issue by performing electrophoretic (2D BN/SDS PAGE) and kinetic analysis in spleen, liver and heart mitochondria from control and infected mice, sacrificed at 72h post intraperitoneal E. coli injection. Our data show a large amount of CI forming SC I1III2 in the control samples: 97-100% of Complex I is bound to dimeric Complex III in spleen, liver and heart mitochondria. To assess the possible functional advantage of the presence of the SC I1III2 [4], we have evaluated the specific activity of Complex I (NADH:ubiquinone oxidoreductase), the NADH-dependent integrated activity by Complex I and Complex III (NADH:cytochrome c oxidoreductase) and the overall respiratory activity (NADH:oxidase), as well as the succinate-dependent integrated activity by Complex II and Complex III (succinate:cytochrome c oxidoreductase). Interestingly, we report high rates of rotenone-insensitive NADH:cytochrome c reductase activity in spleen and liver mitochondria compared to heart samples. We suspect that such activity be associated with the presence of Cytocrome-b5 reductase in the mitochondrial outer membrane and with the production of reactive oxygen species (ROS) that chemically interact with cytochrome c in the assay medium. Our samples of isolated mitochondria from infected animals still show both high percentages of CI bound to SC I1III2 and enzyme activity values comparable to control samples, thus we do not appreciate significant structural/functional differences in the ETS of the investigated tissue mitochondria from infected mice.

Work is in progress for the 2D BN/SDS PAGE analysis in mice BMDMs, both untreated (control) and infected cells co-cultured with live E. coli. Our preliminary results confirm high levels of SC I1III2 in the control samples, which can be fully disassembled in vitro by detergent treatment (1,7% dodecyl maltoside, DDM). The presence of the SC I1III2 seems not to be altered by 12h exposure to E. coli, in accordance with previous results from our laboratory suggesting that BMDM SCs are transiently decreased upon cell infection but come back at 12h, although the whole ETS components seem decreased at the expression level [3]. Further studies are currently underway aimed at analyzing mitochondrial adaptations of BMDM at short time exposure to E. coli.


Bioblast editor: Plangger M, Tindle-Solomon L O2k-Network Lab: IT Bologna Genova ML


Labels: MiParea: Respiration  Pathology: Infectious 

Organism: Mouse  Tissue;cell: Heart, Liver, Blood cells, Macrophage-derived  Preparation: Isolated mitochondria  Enzyme: Complex I, Complex II;succinate dehydrogenase, Complex III, Supercomplex 


HRR: Oxygraph-2k 


Affiliations

Tioli G(1,3), Lenaz G(1), Falasca AI(2), Garaude J(3) and Genova ML(1)
  1. Dept Biomedical Neuromotor Sciences, Univ Bologna, Italy
  2. Dept Food Drug, Univ Parma, Italy
  3. INSERM U1211, Univ Bordeaux, CHU Pellegrin, École de Sages-Femmes, France

References

  1. Sander LE and Garaude J (2018) The mitochondrial respiratory chain: A metabolic rheostat of innate immune cell-mediated antibacterial responses. Mitochondrion 41:28-36.
  2. Lenaz G, Genova ML (2010) Structure and organization of mitochondrial respiratory complexes: a new understanding of an old subject. Antioxid Redox Signal 12:961- 1008.
  3. Garaude J, Acín-Pérez R, Martínez-Cano S, Enamorado M, Ugolini M, Nistal-Villán E, Hervás-Stubbs S, Pelegrín P, Sander LE, Enríquez JA, Sancho D (2016) Mitochondrial respiratory-chain adaptations in macrophages contribute to antibacterial host defense. Nat Immunol 17:1037-45.
  4. Lenaz G, Tioli G, Falasca AI, Genova ML (2016) Complex I function in mitochondrial supercomplexes. Biochim Biophys Acta 1857(7):991-1000.