Identifying cellular pathways modulated by Drosophila palmitoyl-protein thioesterase 1 function

Abstract

Infantile-onset Neuronal Ceroid Lipofuscinosis (INCL) is a severe pediatric neurodegenerative disorder produced by mutations in the gene encoding palmitoyl-protein thioesterase 1 (Ppt1). This enzyme is responsible for the removal of a palmitate post-translational modification from an unknown set of substrate proteins. To better understand the function of Ppt1 in neurons, we performed an unbiased dominant loss-of-function genetic modifier screen in Drosophila using a previously characterized Ppt1 gain-of-function system. The enhancers and suppressors identified in our screen make novel connections between Ppt1 and genes involved in cellular trafficking and the modulation of synaptic growth. We further support the relevance of our screen by demonstrating that Garland cells from Ppt1 loss-of-function mutants have defects in endocytic trafficking. Endocytic tracer uptake and ultrastructural analysis of these non-neuronal cells points to Ppt1 playing a role in modulating the early stages of vesicle formation. This work lays the groundwork for further experimental exploration of these processes to better understand their contributions to the INCL disease process.

Mutations in palmitoyl-protein thioesterase 1 alter exocytosis and endocytosis at synapses in Drosophila larvae

“…This confirms our previous work showing gain-of-function genetic interactions between these 2 loci and electron microscopic analysis of Ppt1 A179T garland cells that revealed ultrastructural defects in endocytic processes similar to those observed in dynamin mutants. 23 Taken together, these data support a direct role for Ppt1 and de-palmitoylation in modulating endocytosis. A reduced synaptic vesicle pool, produced by disruption of dynamin function and reduced synaptic vesicle recycling, could account for the genetic interaction between Ppt1 and shi temperature sensitive paralysis mutants.…”

“…21 Gain-of-function genetic screens in the fly have implicated Ppt1 function in the regulation of several neuronal processes, including synaptic growth/homeostasis and synaptic vesicle endo/exocytosis. 22,23 An examination of non-neuronal larval garland cells in Ppt1 mutants showed reduced endocytosis rates, without alterations in endo-lysosomal trafficking. 23 Similarly, alterations in endocytosis have been observed in fibroblasts from INCL patients.…”

“…22,23 An examination of non-neuronal larval garland cells in Ppt1 mutants showed reduced endocytosis rates, without alterations in endo-lysosomal trafficking. 23 Similarly, alterations in endocytosis have been observed in fibroblasts from INCL patients. 24 In neural tissue, changes in endocytosis have been reported in cultured neurons of Ppt1-deficient mice and humans in the context of a reduction in the synaptic vesicle pool size, but the functional consequences of Ppt1 mutations on endocytosis remains to be characterized.…”

A canine Arylsulfatase G ( ARSG ) mutation leading to a sulfatase deficiency is associated with neuronal ceroid lipofuscinosis

“…To date, the three main cellular mechanisms likely impaired in NCLs are intracellular and membrane trafficking, autophagy, and calcium storage. First, cytoskeletonassociated proteins display abnormalities in CLN1-and CLN5-deficient neurons that are associated with changes in the growth cone assembly (7), and CLN1 modulates the early stages of endocytic vesicule formation (8). Second, reduced autophagy through inhibition of fusion between autophagosomes and lysosomes is observed in most NCLs (5).…”

La céroïde-lipofuscinose de l'American Staffordshire Terriers

“…-il a été montré que des protéines associées au cytosquelette étaient anormales dans les neurones atteints des formes CLN1 et CLN5 (von Schantz et al 2008) et que la protéine de la forme CLN1 (ou protéine CLN1) modulerait les premiers stades de l'endocytose (Saja et al 2010) ; -une diminution de l'autophagie, résultant d'une inhibition de la fusion entre autophagosomes et lysosomes a été observée dans la plupart des CLNs (Bellettato & Scarpa, 2010). En conséquence, des protéines polyubiquitinylées et des mitochondries lésées (sources potentielles de radicaux libres) s'accumulent dans les neurones, le stress du réticulum endoplasmique est activé et ces phénomènes convergent vers l'apoptose du neurone (Zhao & Ackerman, 2006 ;Bellettato & Scarpa, 2010) ;…”