Bax-inhibitor-1knockdown phenotypes are suppressed byBuffyand exacerbate degeneration in aDrosophilamodel of Parkinson disease

Abstract

BackgroundBax inhibitor-1 (BI-1) is an evolutionarily conserved cytoprotective transmembrane protein that acts as a suppressor ofBax-induced apoptosis by regulation of endoplasmic reticulum stress-induced cell death. We knocked downBI-1in the sensitivedopa decarboxylase(Ddc) expressing neurons ofDrosophila melanogasterto investigate its neuroprotective functions. We additionally sought to rescue theBI-1-induced phenotypes by co-expression with the pro-survivalBuffyand determined the effect ofBI-1knockdown on the neurodegenerative α-synuclein-induced Parkinson disease (PD) model.MethodsWe used organismal assays to assess longevity of the flies to determine the effect of the altered expression ofBI-1in theDdc-Gal4-expressing neurons by employing two RNAi transgenic fly lines. We measured the locomotor ability of these RNAi lines by computing the climbing indices of the climbing ability and compared them to a control line that expresses thelacZtransgene. Finally, we performed biometric analysis of the developing eye, where we counted the number of ommatidia and calculated the area of ommatidial disruption.ResultsThe knockdown ofBI-1in these neurons was achieved under the direction of theDdc-Gal4transgene and resulted in shortened lifespan and precocious loss of locomotor ability. The co-expression ofBuffy, the Drosophila anti-apoptotic Bcl-2 homologue, withBI-1-RNAiresulted in suppression of the reduced lifespan and impaired climbing ability. Expression of human α-synucleinin Drosophila dopaminergic neurons results in neuronal degeneration, accompanied by the age-dependent loss in climbing ability. We exploited this neurotoxic system to investigate possible BI-1 neuroprotective function. The co-expression of α-synucleinwithBI-1-RNAiresults in a slight decrease in lifespan coupled with an impairment in climbing ability. In supportive experiments, we employed the neuron-rich Drosophila compound eye to investigate subtle phenotypes that result from altered gene expression. The knockdown ofBI-1in the Drosophila developing eye under the direction of theGMR-Gal4transgene results in reduced ommatidia number and increased disruption of the ommatidial array. Similarly, the co-expression ofBI-1-RNAiwithBuffyresults in the suppression of the eye phenotypes. The expression of α-synucleinalong with the knockdown ofBI-1resulted in reduction of ommatidia number and more disruption of the ommatidial array.ConclusionKnockdown ofBI-1in the dopaminergic neurons of Drosophila results in a shortened lifespan and premature loss in climbing ability, phenotypes that appear to be strongly associated with models of PD in Drosophila, and which are suppressed upon overexpression ofBuffyand worsened by co-expression with α-synuclein. This suggests thatBI-1is neuroprotective and its knockdown can be counteracted by the overexpression of the pro-survivalBcl-2homologue.