Parkin protects against LRRK2 G2019S mutant-induced dopaminergic neurodegeneration in Drosophila - PubMed (original) (raw)

Comparative Study

Parkin protects against LRRK2 G2019S mutant-induced dopaminergic neurodegeneration in Drosophila

Chee-Hoe Ng et al. J Neurosci. 2009.

Abstract

Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are currently recognized as the most common genetic cause of parkinsonism. Among the large number of LRRK2 mutations identified to date, the G2019S variant is the most common. In Asia, however, another LRRK2 variant, G2385R, appears to occur more frequently. To better understand the contribution of different LRRK2 variants toward disease pathogenesis, we generated transgenic Drosophila over-expressing various human LRRK2 alleles, including wild type, G2019S, Y1699C, and G2385R LRRK2. We found that transgenic flies harboring G2019S, Y1699C, or G2385R LRRK2 variant, but not the wild-type protein, exhibit late-onset loss of dopaminergic (DA) neurons in selected clusters that is accompanied by locomotion deficits. Furthermore, LRRK2 mutant flies also display reduced lifespan and increased sensitivity to rotenone, a mitochondrial complex I inhibitor. Importantly, coexpression of human parkin in LRRK2 G2019S-expressing flies provides significant protection against DA neurodegeneration that occurs with age or in response to rotenone. Together, our results suggest a potential link between LRRK2, parkin, and mitochondria in the pathogenesis of LRRK2-related parkinsonism.

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Figures

Figure 1.

Figure 1.

Expression of LRRK2 transgenes in Drosophila neither affects its overall brain architecture nor triggers obvious eye abnormalities. A, Left, Anti-elav (green) and anti-LRRK2 (red) immunostaining of whole-mount adult brains derived from 2-d-old control or transgenic flies expressing wild-type human LRRK2, as indicated. Right, Enlarged images (and inset) show the localization of elav and LRRK2 signals to the nucleus and cytoplasm, respectively, in the brain of a wild-type LRRK2-expressing fly. An anti-myc immunoblot of brain lysates prepared from 2-d-old control or transgenic flies expressing various LRRK2 species is shown below (genotype: elav-Gal4/+ or elav–Gal4–hLRRK2). B, SEM eye images of 20-d-old GMR-Gal4/+ or GMR–Gal4–hLRRK adult flies. No retinal degeneration was observed in all flies examined.

Figure 2.

Figure 2.

Expression of LRRK2 mutants in flies promotes DA neurodegeneration and concomitant locomotion deficits. A, B, Bar graph showing the number of TH-positive DA neurons in different clusters of various fly species at 20 or 60 d after eclosion, as indicated (n = 10). C, Representative confocal microscopy images showing TH-positive (red) DA neurons in the PPM1 cluster (boxed) of 60-d-old control or LRRK2-expressing flies. Inset, Higher magnification of boxed regions. D, E, Bar graph showing the percentage of various male (D) and female (E) flies at different days after eclosion that reached the top of assay column after 1 min (n = 20) (genotype: ddc–Gal4/+ or ddc–Gal4–hLRRK2).

Figure 3.

Figure 3.

Exposure to rotenone accelerates DA degeneration in LRRK2 G2019 and G2385R mutant flies. A, Bar graph showing the number of TH-positive DA neurons in different clusters of various fly species at 15 d after rotenone treatment (n = 15). B, Representative confocal microscopy images showing TH-positive (red) DA neurons in the PPM3 cluster of various, rotenone-treated fly species, as indicated (genotype: ddc–Gal4/+ or ddc–Gal4–hLRRK2).

Figure 4.

Figure 4.

Parkin coexpression mitigates DA degeneration in LRRK2 G2019S-expressing flies in the presence or absence of rotenone. A, Bar graph showing the number of TH-positive DA neurons in different clusters of various fly species at 15 d after rotenone treatment (n = 15). B, Representative confocal microscopy images showing TH-positive (red) DA neurons in whole-mount adult brains derived from rotenone-treated flies expressing either LRRK2 G2019S alone or in the presence of parkin coexpression. Right, PPM3 cluster (circled) are shown at higher magnification. C, Bar graph showing the number of TH-positive DA neurons in different clusters of the various fly species at 60 d after eclosion, as indicated (n = 8) (genotype: ddc–Gal4/+ or ddc–Gal4–hLRRK2 or ddci-Gal4–hLRRK2;ddc–Gal4–hparkin).

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