SIM2s coordinates programmed mitophagy with respiratory chain supercomplex remodeling during mammary epithelial differentiation.

Lactation requires mammary epithelial cells (MECs) to rapidly expand mitochondrial function while remodeling the mitochondrial population that supports milk synthesis and secretion. Programmed mitophagy is required for MEC differentiation, yet why mitochondrial turnover is necessary during this developmental transition remains poorly understood. Using Mito-QC reporter mice, we identified developmentally regulated changes in mitolysosome burden across the transition from late pregnancy to lactation that were altered by mammary-specific gain or loss of the bhlh/PAS protein, SIM2s (single-minded 2 s). In differentiating HC11 cells, mitochondrial turnover was accompanied by increased assembly and activity of respiratory supercomplexes containing complexes I, III, and IV. Depletion of PRKN prevented acquisition of this differentiation-associated respiratory profile and impaired lactogenic differentiation. SIM2s co-migrated with higher-order respiratory assemblies, and loss of SIM2s reduced supercomplex assembly and activity in HC11 cells and mammary tissue. SIM2s also localized in close proximity to complex III in differentiated mammary epithelium, whereas loss of SIM2s reduced proximity between complexes III and IV. Together, these findings support a model in which SIM2s coordinates PRKN-dependent mitochondrial turnover with respiratory-chain remodeling during MEC differentiation. Our results suggest that programmed mitophagy does more than remove mitochondria during development; it contributes to establishment of a mitochondrial population with a respiratory-chain architecture suited to the emerging differentiated state.