SPRIZ (CCDC127) protein conserved in deuterostome animals is a rigid OMM-IMM tether that locally sets mitochondrial...

The double-membrane nature of mitochondria generates an intermembrane volume through which ions, reactive oxygen species, and metabolites must flux to reach the matrix or cytosol, but whether this volume is a genetically specified structure remains unknown. We identify SPRIZ (spatial regulator of the intermembrane zone), the product of Ccdc127, a gene conserved across deuterostome animals. SPRIZ assembles into an ∼18-nanometer homodimeric tether anchored in the outer mitochondrial membrane (OMM) and inserted into the inner mitochondrial membrane (IMM) through vertebrate-specific hydrophobic loops, physically spacing the two membranes and setting local intermembrane volume. Cryo-electron tomography of native mitochondria revealed rod-like densities spanning the intermembrane space (IMS), consistent with SPRIZ's predicted rigid coiled-coil architecture and membrane-spanning topology. In mouse liver, two-dimensional morphometry measured ∼3-fold expansion of IMS area with SPRIZ and ∼5.5-fold with lengthened mitochondrial artificial increaser of the IMS (MARTINI); 3D electron tomography confirmed MARTINI's greater OMM-IMM separation and normalized IMS volume. This expansion was functionally validated by increased ¹⁴C-mannitol accessibility to the IMS and by impaired mitochondrial Ca²⁺ uptake. Constitutive Ccdc127/SPRIZ knockout produced viable mice and, in liver mitochondria, preserved overall ultrastructure but caused modest IMS expansion, consistent with loss of local OMM-IMM constraints that stabilize membrane geometry at discrete sites. We therefore identify IMS volume as a genetically encoded and tunable architectural parameter of vertebrate mitochondria. In this model, SPRIZ is a molecular spacing system that locally sets OMM-IMM separation, shaping the environment through which metabolites, ions, and signals diffuse and establishing local gradients within the IMS.