In a groundbreaking study published in Science Advances, researchers from the CeMM Research Center for Molecular Medicine have unveiled comprehensive insights into the critical role of the SLC13A5 membrane transporter in neuronal metabolism and its connection to a severe epileptic disorder. Citrate, a central metabolite in cellular biochemistry, is intricately involved in energy production and cellular signaling within neurons. This study elucidates how mutations in the SLC13A5 gene disrupt citrate transport, ultimately leading to developmental epileptic encephalopathy (DEE), a rare but devastating neurological condition.
Citrate serves multiple vital functions in cells, acting primarily as an intermediary in the citric acid cycle, a foundational metabolic pathway responsible for generating energy in the form of ATP. Beyond energy production, citrate contributes to biosynthetic processes essential for cell growth and maintenance. Notably, in neurons, citrate also functions as a neuromodulator, influencing synaptic activity. This dual role heightens the necessity for precise regulation of citrate uptake in the brain, a task mediated predominantly by the SLC13A5 transporter situated in the neuronal cell membranes.
The SLC13A5 protein belongs to a family of solute carrier (SLC) transporters that facilitate the translocation of various substrates across cellular membranes, playing critical roles in maintaining cellular homeostasis. In the brain, high levels of SLC13A5 expression ensure adequate citrate influx from the cerebrospinal fluid into neurons. When mutations impair this transporter’s function, citrate levels become dysregulated, which has been directly linked to the onset of DEE, a condition characterized by early-life seizures and neurodevelopmental impairment.
Despite the clinical significance, the molecular mechanisms governing how distinct SLC13A5 mutations lead to disease phenotypes were poorly understood until now. To address this, the CeMM team employed an advanced technique called deep mutational scanning (DMS), enabling the systematic evaluation of almost ten thousand possible genetic variants of SLC13A5 for their functional impact. This unprecedented scale of analysis allowed for the identification of critical mutations affecting transporter stability, cellular localization, and citrate uptake efficiency.
From this massive dataset, 38 mutant variants were further subjected to experimental interrogation to validate computational predictions and to dissect the biophysical alterations caused by these mutations. This integrative approach revealed that certain mutations lead to reduced protein expression at the membrane, while others compromise the transport kinetics of citrate, decreasing its cellular availability. Such molecular impairments collectively result in defective metabolic processes in neurons, thereby underpinning the pathological basis of SLC13A5 transporter disorder.
Moreover, the researchers introduced a novel framework to assess protein stability across distinct conformational states of SLC13A5, coupled with evolutionary conservation scoring to prioritize variants with probable pathogenicity. These innovative computational tools serve not only in characterizing rare disease mutations but also in expanding our understanding of population-level genetic diversity and its subtle impacts on protein function.
The implications of these findings extend far beyond the narrow confines of a single rare disease. Understanding how membrane transporters like SLC13A5 operate and fail at a molecular level provides essential insights into neuronal biochemistry and paves the way for rational drug design. Precision medicine approaches can now leverage this data to better diagnose and potentially develop targeted therapies for individuals afflicted by SLC13A5-associated epileptic encephalopathy.
“Systematic functional characterization of genetic variants is a powerful strategy, particularly to elucidate the molecular underpinnings of rare and complex human diseases,” notes co-first author Wen-An Wang. His colleague Evandro Ferrada adds that combining experimental data with computational modeling bridges the gap between genotype and phenotype, offering a comprehensive picture of variant effects that can inform clinical interpretation.
This work was made possible through synergy with the RESOLUTE and REsolution consortia, multi-institutional efforts geared towards decoding the entire family of SLC transporters and understanding their roles in cellular logistics. Patient-derived data, obtained from the TESS Research Foundation, further grounded the molecular findings within a clinical context aligned with patient needs.
Giulio Superti-Furga, senior author and scientific director at CeMM, emphasizes that this study exemplifies how blending large-scale mutational analysis with structural and functional elucidation can dramatically enhance our grasp of transporter biology. It underscores the broader principle that precision functional mapping of membrane proteins is essential for translating genetic variation into mechanistic insights and clinical solutions.
As the SLC13A5 transporter’s malfunction is implicated not only in epilepsy but might also be linked indirectly to other neurological and metabolic disorders, future investigations building on this work could unlock new therapeutic avenues. The potential to modulate transporter activity pharmacologically or through gene therapy offers hope for conditions that currently have no effective treatments.
In conclusion, this landmark study sets a high bar for variant effect mapping in membrane proteins and establishes a foundational knowledge base for rare disease research. By integrating deep mutational scans with computational and biochemical methodologies, the investigators have not only clarified the pathogenesis of SLC13A5 Citrate Transporter Disorder but have also broadened the horizon for understanding metabolic control in neuronal health and disease.
Subject of Research: Cells
Article Title: Large-scale experimental assessment of variant effects on the structure and function of the citrate transporter SLC13A5
News Publication Date: 27-Jun-2025
Web References:
10.1126/sciadv.adx3011
References:
Wang, W.-A., Ferrada, E., Klimek, C., Osthushenrich, T., MacNamara, A., Wiedmer, T., & Superti-Furga, G. (2025). Large-scale experimental assessment of variant effects on the structure and function of the citrate transporter SLC13A5. Science Advances, 11(26), eadx3011.
Image Credits:
© CeMM / © Franzi Kreis/CeMM
Keywords: Transporter proteins, Transmembrane proteins, Biomolecules, Life sciences, Cell biology
