👤 Talal Mousallem

Juvenile neuronal ceroid lipofuscinosis (JNCL) belongs to the neuronal ceroid lipofuscinoses characterized by blindness/seizures/motor/cognitive decline and early death. JNCL is caused by CLN3 gene mutations that negatively modulate cell growth/apoptosis. CLN3 protein (CLN3p) localizes to Golgi/Rab4-/Rab11-positive endosomes and lipid rafts, and harbors a galactosylceramide (GalCer) lipid raft-binding domain. Goals are proving CLN3p participates in GalCer transport from Golgi to rafts, and GalCer deficits negatively affect cell growth/apoptosis. GalCer/mutant CLN3p are retained in Golgi, with CLN3p rescuing GalCer deficits in rafts. Diminishing GalCer in normal cells by GalCer synthase siRNA negatively affects cell growth/apoptosis. GalCer restores JNCL cell growth. WT CLN3p binds GalCer, but not mutant CLN3p. Sphingolipid content of rafts/Golgi is perturbed with diminished GalCer in rafts and accumulation in Golgi. CLN3-deficient raft vesicular structures are small by transmission electron microscopy, reflecting altered sphingolipid composition of rafts. CLN1/CLN2/CLN6 proteins bind to lysophosphatidic acid/sulfatide, CLN6/CLN8 proteins to GalCer, and CLN8 protein to ceramide. Sphingolipid composition/morphology of CLN1-/CLN2-/CLN6-/CLN8- and CLN9-deficient rafts are altered suggesting changes in raft structure/lipid stoichiometry could be common themes underlying these diseases. Show less

The neuronal ceroid lipofuscinoses are pediatric neurodegenerative diseases with common clinical features. Of the nine clinical variants (CLN1-CLN9), six have been genetically identified. Most variants manifest cell death and dysregulated sphingolipid metabolism, suggesting the proteins defective in these disorders may interact along one pathway. NCL patient-derived cell lines exhibit cell growth and apoptotic defects that reverse following transfection with the wild-type gene. The membrane-bound proteins CLN3, CLN6, and CLN8 complement each other, as do CLN1 and CLN2 proteins, with respect to growth and apoptosis. The CLN2 protein also corrects growth and apoptosis in CLN3-, CLN6-, and CLN8-deficient cell lines. Neither CLN1-deficient nor CLN2-deficient growth defects are corrected by CLN3, CLN6, and CLN8 proteins. CLN2, CLN3, CLN6, and CLN8 proteins co-immunoprecipitate and co-localize to early and/or recycling endosomes and lipid rafts. Additionally, CLN2p and CLN1p co-immunoprecipitate. The work presented supports interactions between NCL proteins occurring at multiple points along one pathway. Show less