Structural and functional implications of SLC13A3 and SLC9A6 mutations: an in silico approach to understanding intellectual disability

Abstract

Background: Intellectual disability (ID) is a condition that varies widely in both its clinical presentation and its genetic underpinnings. It significantly impacts patients' learning capacities and lowers their IQ below 70. The solute carrier (SLC) family is the most abundant class of transmembrane transporters and is responsible for the translocation of various substances across cell membranes, including nutrients, ions, metabolites, and medicines. The SLC13A3 gene encodes a plasma membrane-localized Na+/dicarboxylate cotransporter 3 (NaDC3) primarily expressed in the kidney, astrocytes, and the choroid plexus. In addition to three Na + ions, it brings four to six carbon dicarboxylates into the cytosol. Recently, it was discovered that patients with acute reversible leukoencephalopathy and a-ketoglutarate accumulation (ARLIAK) carry pathogenic mutations in the SLC13A3 gene, and the X-linked neurodevelopmental condition Christianson Syndrome is caused by mutations in the SLC9A6 gene, which encodes the recycling endosomal alkali cation/proton exchanger NHE6, also called sodium-hydrogen exchanger-6. As a result, there are severe impairments in the patient's mental capacity, physical skills, and adaptive behavior.

Methods and results: Two Pakistani families (A and B) with autosomal recessive and X-linked intellectual disorders were clinically evaluated, and two novel disease-causing variants in the SLC13A3 gene (NM 022829.5) and the SLC9A6 gene (NM 001042537.2) were identified using whole exome sequencing. Family-A segregated a novel homozygous missense variant (c.1478 C > T; p. Pro493Leu) in the exon-11 of the SLC13A3 gene. At the same time, family-B segregated a novel missense variant (c.1342G > A; p.Gly448Arg) in the exon-10 of the SLC9A6 gene. By integrating computational approaches, our findings provided insights into the molecular mechanisms underlying the development of ID in individuals with SLC13A3 and SLC9A6 mutations.

Conclusion: We have utilized in-silico tools in the current study to examine the deleterious effects of the identified variants, which carry the potential to understand the genotype-phenotype relationships in neurodevelopmental disorders.

Keywords: Acute reversible leukoencephalopathy; Christianson Syndrome; Exome sequencing; Intellectual disability; Molecular dynamics simulation; SLC13A3; SLC9A6.

Fig. 1

(a) Pedigree of Family A showing the autosomal recessive pattern of ID and presenting the unaffected and affected individuals of the family (A) (b) Pedigree of Family B showing the X-linked pattern of ID and presenting the unaffected and affected individuals of the family (B) Arrows are representing the DNA samples of the individuals submitted for exome sequencing

Fig. 2

(a, b, c) Physical features presented by affected individual (V-2) of family A and (d) Computed tomography (CT) scan image of the affected individual (V-2) of family A indicative of multiple large CSF spaces within and around the brain suggestive of cerebral atrophy, (e, f) physical features presented by affected individuals (V-3 and V-4) of family B

Fig. 3

(A) Representing the location of SLC13A3 (q13.12) at chromosome 20. (B) representing typical structure of the SLC13A3 gene comprising 13 exons and also indicating the location of the missense variant (c.1478 C > T; p. Pro493Leu) in exon-11 (C) Chromatograms of homozygous unaffected son (V-1), heterozygous carrier father (IV-1) and homozygous affected son (V-2) of family A. (D) Location of SLC9A6 (q26.3) at chromosome X. (E) the typical structure of SLC9A6 gene comprising 16 exons, indicating the location of the missense variant (c.1342G > A; p. Gly448Arg) in exon-10. (F) Chromatograms of hemizygous unaffected father (IV-1), heterozygous carrier mother (IV-2) and hemizygous affected son (V-3) of family B

Fig. 4

Comparative Binding Site Representations of SLC9A6 and SLC13A3. (A) Ribbon depiction of the SLC9A6 ion-binding site in the 6-transmembrane (TM) core transport domain, superimposed on NHE9 (PDB ID: 6Z3Y). A detailed view of the predicted ion-binding site of SLC9A6, with crucial residues displayed as yellow sticks and labeled accordingly. Corresponding residues in the NHE9 structure are indicated in orange. (B) Ribbon depiction of the SLC13A3 sodium and substrate binding site, superimposed on SLC13A5 (PDB ID: 7JSK). A detailed view of the predicted binding site of SLC13A3 is shown, with crucial residues represented as yellow sticks and labeled. Corresponding residues in the SLC13A5 structure are shown in orange

Fig. 5

Structural Evolution of Wild-Type and Mutant Proteins Over 300 ns MD Simulations. The comparative progression of the wild-type and mutant protein models (SLC9A6_G448R and SLC13A3_P493L) during the 300 ns Molecular Dynamics (MD) simulation is illustrated. Snapshots taken every 30 ns offer a visual representation of changes in protein conformation and dynamics throughout the simulation period. Affected mutation sites are distinctly highlighted: for SLC9A6_G448R, the focus is on the intact ion-binding site, while for SLC13A3_P493L, it’s on the regions around the Na+- and substrate-binding sites, showcasing their dynamic fluctuations