GAT3-dependent regulation of glioma invasiveness via a lipid raft-associated PMCA4 Ca2+ transporter and a downstream CaMKII/CREB signaling – implications for compartmentalized signaling in glioma tumors

Emerging evidence underscores the crucial role of compartmentalized Ca²⁺ and GABA signaling in the development and progression of gliomas. Our findings reveal that low GAT3 expression and high PMCA4 levels are strongly associated with poor survival outcomes in glioma patients, suggesting their involvement in tumor progression. Using C6 glioma model, we uncovered a dynamic interaction between GAT3 and PMCA4 within lipid raft microdomains, which plays a key role in fine-tuning of localized Ca²⁺ dynamics in response to GABA stimulation. Knockdown of PMCA4 increased resting Ca²⁺concentration and enhanced Ca²⁺ accumulation in lipid rafts following 3-min pulse GABA stimulation, significantly impairing glioma cell migration and invasion. Interestingly, the expression of Ca²⁺ chelator parvalbumin in rafts abolished both baseline and GABA-stimulated Ca²⁺ rises, effectively restoring the migratory and invasive potential of tumor cells. We further demonstrated that GAT3 interacted with calmodulin, a pivotal regulator of PMCA4, and this interaction was decreased following 24 h GABA treatment. Long-term GABA stimulation also disrupted PMCA4/GAT3 complex, overloaded lipid rafts with Ca²⁺ and decreased glioma invasiveness in the presence of PMCA4. In these conditions, we observed GAT3- and Ca²⁺/calmodulin-dependent protein kinase II-dependent CREB phosphorylation at Ser133, which was controlled by Ca²⁺ events in lipid rafts and required to maintain glioma invasiveness. Our study uncovers a previously unrecognized GAT3-dependent mechanism of Ca²⁺compartmentalization in membrane microdomains, shedding new light on its potential role in tumor behavior. Understanding these local Ca²⁺ signaling partnerships will offer valuable insights into gliomagenesis and could lead to the development of novel therapeutic strategies for glioma treatment.