Novel ethyl methanesulfonate (EMS)-induced alleles of the Drosophila homolog of LRRK2 reveal a crucial role in endolysosomal functions and autophagy in vivo.
Bottom Line: Using these alleles, we show that lrrk loss-of-function causes striking defects in the endolysosomal and autophagy pathways, including the accumulation of markedly enlarged lysosomes that are laden with undigested contents, consistent with a defect in lysosomal degradation. lrrk loss-of-function also results in the accumulation of autophagosomes, as well as the presence of enlarged early endosomes laden with mono-ubiquitylated cargo proteins, suggesting an additional defect in lysosomal substrate delivery.Interestingly, the lysosomal abnormalities in these lrrk mutants can be suppressed by a constitutively active form of the small GTPase rab9, which promotes retromer-dependent recycling from late endosomes to the Golgi.Collectively, our data provides compelling evidence of a vital role for lrrk in lysosomal function and endolysosomal membrane transport in vivo, and suggests a link between lrrk and retromer-mediated endosomal recycling.
Affiliation: Department of Neurology, University of California, Los Angeles, CA 90095, USA. Molecular Biology Institute, University of California, Los Angeles, CA 90095, USA.Show MeSH
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Mentions: Previously, we and others have reported that lrrke03680 homozygous females show a dramatic reduction in female fertility (Dodson et al., 2012; Imai et al., 2008; Lee et al., 2007), which can be rescued by restoring wild-type lrrk function specifically in follicle cells (Dodson et al., 2012). Follicle cells are a somatic epithelial cell monolayer that surrounds the developing oocyte during oogenesis (Fig. 2A) (King, 1970; Spradling, 1993). Follicle cells have a number of advantages that make them an attractive cell biological system, including their large size, ease of accessibility and squamous morphology that aids in visualization of subcellular structures. As expected, all trans-heterozygous combinations of lrrk NS alleles showed reduced female fertility that could be rescued by follicle-cell-specific expression of wild-type lrrk (Fig. 1C). Expression of lrrkGS, which is analogous to the most common PD-causing mutation in human LRRK2 (G2019S) (Bonifati, 2007; Taylor et al., 2006), also suppressed female infertility in lrrk NS flies (Fig. 1C). This suggests that lrrkGS retains at least some of the functions of wild-type lrrk, consistent with what we have previously reported (Dodson et al., 2012). Moreover, follicle-cell-specific expression of human LRRK2 also restored fertility to lrrk NS flies (Fig. 1C), suggesting that the human and Drosophila proteins are functionally conserved.
Affiliation: Department of Neurology, University of California, Los Angeles, CA 90095, USA. Molecular Biology Institute, University of California, Los Angeles, CA 90095, USA.