A Mitochondrial Basis for Tead4 Bioavailability at the First Mammalian Cell Fate Decision

Abstract Specification of the inner cell mass (ICM) and trophectoderm (TE) at the first mammalian cell fate decision requires the transcription factor Tead4, yet what restricts Tead4 activity to presumptive TE cells remains unknown. Tead4 localizes to mitochondria, and the ICM and TE harbor distinct mitochondrial populations, but whether Tead4 distribution varies across mitochondrial subtypes in the cleavage-stage embryo has not been examined. Here we used fluorescence-activated mitochondrial sorting (FAMS) to characterize mitochondrial subpopulations in mouse metaphase-II oocytes and 8-cell embryos with respect to size, mitochondrial membrane potential (ΔΨm), and Tead4 protein content. Mitochondria are heterogeneous in size and ΔΨm in both developmental stages, with large mitochondria exhibiting markedly higher ΔΨm than small mitochondria. Tead4 protein is concentrated in the large, high-ΔΨm mitochondrial subpopulation in 8-cell embryos, with 75% of large mitochondria containing Tead4 compared to only 3% of small mitochondria. The overall size distribution of the mitochondrial pool is maintained between oocytes and 8-cell embryos; Tead4 accumulation within the large mitochondrial fraction is therefore a developmentally regulated process initiated specifically during the early embryogenesis. These findings establish for the first time that Tead4 localizes preferentially to large, high-ΔΨm mitochondria in the cleavage-stage embryo, providing a previously unrecognized cellular basis for understanding how Tead4 bioavailability may be regulated prior to TE specification.