Structural and mutational insights define ERMA as the ER Mg ²⁺ ATPase and reservoir gatekeeper

Magnesium (Mg ²⁺ ) is the most abundant divalent cation in cells, yet the mechanisms mediating its organellar transport remain poorly defined. We identify endoplasmic reticulum (ER) Mg ²⁺ adenosine triphosphatase (ATPase) (ERMA) as the transporter that drives Mg ²⁺ uptake into the ER lumen, establishing the ER as a bi-ionic intracellular reservoir. MagFRET biosensors targeted to the ER demonstrate that ERMA mediates dynamic ER Mg ²⁺ storage and robust adenosine 5′-triphosphate–dependent Mg ²⁺ uptake reaching 15 to 30 millimolar. Cryo–electron microscopy structures of human and mouse ERMA reveal a P-type ATPase fold with an unwound transmembrane 4 (TM4) that coordinates Mg ²⁺ via the unique PILP backbone and the TM5 residue Q1110, whose mutation markedly impairs ERMA-mediated Mg ²⁺ uptake. Functional reconstitution of domain mutants, ERMA-SERCA chimeras, and pathogenic variants confirm ERMA as an ER-resident Mg ²⁺ pump and gatekeeper of ER Mg ²⁺ ionic equilibrium.