The mitochondrial inner membrane organizing system (MINOS) is a conserved large 
hetero-oligomeric protein complex in the mitochondrial inner membrane, crucial 
for the maintenance of cristae morphology. MINOS has been suggested to represent 
the core of an extended protein network that controls mitochondrial function and 
structure, and has been linked to several human diseases. The spatial 
arrangement of MINOS within mitochondria is ill-defined, however. Using 
super-resolution stimulated emission depletion (STED) microscopy and immunogold 
electron microscopy, we determined the distribution of three known human MINOS 
subunits (mitofilin, MINOS1, and CHCHD3) in mammalian cells. Super-resolution 
microscopy revealed that all three subunits form similar clusters within 
mitochondria, and that MINOS is more abundant in mitochondria around the nucleus 
than in peripheral mitochondria. At the submitochondrial level, mitofilin, a 
core MINOS subunit, is preferentially localized at cristae junctions. In primary 
human fibroblasts, mitofilin labeling uncovered a regularly spaced pattern of 
clusters arranged in parallel to the cell growth surfaces. We suggest that this 
array of MINOS complexes might explain the observed phenomenon of largely 
horizontally arranged cristae junctions that connect the inner boundary membrane 
to lamellar cristae. The super-resolution images demonstrate an unexpectedly 
high level of regularity in the nanoscale distribution of the MINOS complex in 
human mitochondria, supporting an integrating role of MINOS in the structural 
organization of the organelle.