The complex cytopathology of mitochondrial diseases is usually attributed to insufficient ATP. AMP-activated protein kinase (AMPK) is a highly sensitive cellular energy sensor that is stimulated by ATP-depleting stresses. By antisense-inhibiting chaperonin 60 expression, we produced mitochondrially diseased strains with gene dose-dependent defects in phototaxis, growth, and multicellular morphogenesis. Mitochondrial disease was phenocopied in a gene dose-dependent manner by overexpressing a constitutively active AMPK alpha subunit (AMPKalphaT). The aberrant phenotypes in mitochondrially diseased strains were suppressed completely by antisense-inhibiting AMPKalpha expression. Phagocytosis and macropinocytosis, although energy consuming, were unaffected by mitochondrial disease and AMPKalpha expression levels. Consistent with the role of AMPK in energy homeostasis, mitochondrial "mass" and ATP levels were reduced by AMPKalpha antisense inhibition and increased by AMPKalphaT overexpression, but they were near normal in mitochondrially diseased cells. We also found that 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside, a pharmacological AMPK activator in mammalian cells, mimics mitochondrial disease in impairing Dictyostelium phototaxis and that AMPKalpha antisense-inhibited cells were resistant to this effect. The results show that diverse cytopathologies in Dictyostelium mitochondrial disease are caused by chronic AMPK signaling not by insufficient ATP.
SUMMARYHuman patients with mitochondrial diseases are more susceptible to bacterial infections, particularly of the respiratory tract. To investigate the susceptibility of mitochondrially diseased cells to an intracellular bacterial respiratory pathogen, we exploited the advantages of Dictyostelium discoideum as an established model for mitochondrial disease and for Legionella pneumophila pathogenesis. Legionella infection of macrophages involves recruitment of mitochondria to the Legionella-containing phagosome. We confirm here that this also occurs in Dictyostelium and investigate the effect of mitochondrial dysfunction on host cell susceptibility to Legionella. In mitochondrially diseased Dictyostelium strains, the pathogen was taken up at normal rates, but it grew faster and reached counts that were twofold higher than in the wild-type host. We reported previously that other mitochondrial disease phenotypes for Dictyostelium are the result of the activity of an energy-sensing cellular alarm protein, AMP-activated protein kinase (AMPK). Here, we show that the increased ability of mitochondrially diseased cells to support Legionella proliferation is suppressed by antisense-inhibiting expression of the catalytic AMPKα subunit. Conversely, mitochondrial dysfunction is phenocopied, and intracellular Legionella growth is enhanced, by overexpressing an active form of AMPKα in otherwise normal cells. These results indicate that AMPK signalling in response to mitochondrial dysfunction enhances Legionella proliferation in host cells.
There was an error in the first ePress version of the article Dis. Model. Mech. 2, 479-489.In the ePress version of 28 July 2009, the Clinical issue section of the Translational Impact text had a missing word, which altered the meaning. The correct sentence should read: 'In many cases, the precise genetic defect causing mitochondrial dysfunction is known, but the downstream pathological outcomes for the mitochondrially diseased cell remain poorly understood. ' The correct version was published on ePress on 30 July 2009. The print version is correct.We apologise for this mistake.
Mitochondrial biogenesis is primarily a critical adaptation aimed to balance an increased workload in an attempt to maintain appropriate body perfusion. Until recently, the signaling mechanisms responsible for this response are poorly understood. To examine the role of AMP-activated protein kinase (AMPK), an evolutionarily conserved fuel sensor, in mitochondrial biogenesis, we used constitutively active and antisense inhibition genetic tools in Dictyostelium discoideum. Constitutive activation of AMPK culminated in mitochondrial proliferation and elevated ATP generation; this became marked with higher plasmid copies. Antisense inhibition of AMPK yielded non-significant decrease in the mitochondrial content at low levels. However, the more severe the antisense inhibition, the more significant the diminution of AMPK function, resulting in the more apparent decrease in the Advanced Technology Program (ATP) and mitotracker fluorescence. This finding provides direct genetic evidence that AMPK plays a significant role in ameliorating the effects of cellular energy deficit through mitochondrial proliferation. Thus, the constitutive activation of AMPK initiates signalling to downstream targets. The result perturbation of these pathways would culminate in the mitochondrial biogenesis. Taken together, these findings show the constitutive activation of AMPK propels in vivo mitochondrial biogenesis and ATP generation in D. discoideum as in other organisms.
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