Rett syndrome (RTT) is a severe neurodevelopmental disorder primarily caused by mutations in the methyl-CpG binding protein 2 (MECP2) gene. Despite advancements in research, no cure exists due to an incomplete understanding of the molecular effects of MeCP2 deficiency. Previous studies have identified impaired tropomyosin receptor kinase (Trk) neurotrophin (NTP) signaling and mitochondrial redox imbalances as key drivers of the pathology. Moreover, altered glycosphingolipid metabolism has been reported in RTT. GM1 ganglioside is a known regulator of the nervous system, and growing evidence indicates its importance in maintaining neuronal homeostasis via its oligosaccharide chain, coded as GM1-OS. GM1-OS directly interacts with the Trk receptors on the cell surface, triggering neurotrophic and neuroprotective pathways in neurons. In this study, we demonstrate that GM1-OS ameliorates RTT deficits in the Mecp2-null model. GM1-OS restored synaptogenesis and reduced mitochondrial oxidative stress of Mecp2-knock-out (ko) cortical neurons. When administered in vivo, GM1-OS mitigated RTT-like symptoms. Our findings indicate that GM1-OS effects were mediated by Trk receptor activation on the neuron’s plasma membrane. Overall, our results highlight GM1-OS as a promising candidate for RTT treatment.

Background: GM3 Synthase Deficiency (GM3SD) is a rare autosomal recessive neurodevelopmental disease characterized by recurrent seizures and neurological deficits. The disorder stems from mutations in the ST3GAL5 gene, encoding GM3 synthase (GM3S), a key enzyme in ganglioside biosynthesis. While enzyme deficiencies affecting ganglioside catabolism are well-documented, the consequences of impaired ganglioside biosynthesis remain less explored. Methods: To investigate GM3SD, we used a Human Embryonic Kidney 293-T (HEK293-T) knockout (KO) cell model generated via CRISPR/Cas9 technology. Lipid composition was assessed via high-performance thin-layer chromatography (HPTLC); glycohydrolase activity in lysosomal and plasma membrane (PM) fractions was enzymatically analyzed. Lysosomal homeostasis was evaluated through protein content analysis and immunofluorescence, and cellular bioenergetics was measured using a luminescence-based assay. Results: Lipidome profiling revealed a significant accumulation of lactosylceramide (LacCer), the substrate of GM3S, along with increased levels of monosialyl-globoside Gb5 (MSGb5), indicating a metabolic shift in glycosphingolipid biosynthesis. Lipid raft analysis revealed elevated cholesterol levels, which may impair microdomain fluidity and signal transduction. Furthermore, altered activity of lysosomal and plasma membrane (PM)-associated glycohydrolases suggests secondary deregulation of glycosphingolipid metabolism, potentially contributing to abnormal lipid patterns. In addition, we observed increased lysosomal mass, indicating potential lysosomal homeostasis dysregulation. Finally, decreased adenosine triphosphate (ATP) levels point to impaired cellular bioenergetics, emphasizing the metabolic consequences of GM3SD. Conclusions: Together, these findings provide novel insights into the molecular alterations associated with GM3SD and establish the HEK293-T KO model as a promising platform for evaluating potential therapeutic strategies.