Naturally Occurring SGLT2 Variants Reveal Critical Determinants of MAP17-Dependent Glucose Transport by SGLT2

Background: Sodium-glucose cotransporter 2 (SGLT2) is the major mechanism of glucose reabsorption at the renal proximal tubule. Recent studies showed that transport of glucose by SGLT2 requires MAP17, a small non-glycosylated transmembrane protein. However, the mechanisms that define MAP17 regulation of SGLT2 function remain unclear. Our previous work identified the interactive residues on SGLT2-MAP17 complex predicted by cryogenic electron microscope (cryo-EM). This study aims to identify key missense SNPs in SGLT2 that affect MAP17-dependent glucose transport. Methods: SGLT2 SNPs, were obtained from dbSNP. SNPs on the last transmembrane domain, where SGLT2 interacts with MAP17, were selected. Glucose transport activities were measured in Xenopus oocytesco-expressing MAP17 with either wild-type SGLT2 (WT) or mutant proteins. Glucose transport was assessed by measuring whole cell currents generated by glucose-activated Na+ transport using two-electrode voltage clamp. Results: Mutation L667P significantly reduced SGLT2-mediated glucose transport by 74% compared to WT (p<0.001). In contrast, mutations M661T and V665M enhanced glucose transport by 43% (p=0.006) and 52% (p<0.001), respectively. N654S and N654Y did not significantly alter transport function. Immunofluorescence confirmed membrane expression for all SGLT2 mutations. Conclusion: Our findings suggest that L667 represents a critical residue for MAP17 interaction. This position is evolutionarily distinct from the corresponding conserved cysteine in SGLT1, potentially explaining SGLT2's unique MAP17-dependent activation compared to SGLT1 which does not need activation by MAP17. The enhanced glucose transport observed with M661T and V665M mutations suggests that these residues may normally impose structural constraints on the SGLT2-MAP17 complex. Interestingly, our observation that the N654S mutation maintained normal glucose transport functionality appears to contrast with a clinical report of mild glucosuria in a human heterozygous carrier, suggesting that the reported phenotype may result from additional factors. Significance: This study identified and functionally validated naturally occurring SGLT2 variants at the SGLT2-MAP17 interface, offering crucial insights into the molecular determinants of renal glucose reabsorption. Funding: Other NIH Support - NIH-U54 GM104940, Private Foundation Support