Mitochondria-Associated Transcription Precedes Oxidative Phosphorylation in Human Embryos

Mitochondria undergo significant structural and functional changes during human pre-implantation embryogenesis, yet the transcriptional activity of both nuclear-encoded mitochondria-associated genes and mitochondrially transcribed genes across this developmental window remains poorly characterized. While mitochondria are established as the primary energy source for the early embryo, emerging evidence suggests they may also influence lineage specification through epigenetic regulation and metabolite availability. To investigate this, we applied a focused organelle-specific transcriptomic analysis framework, filtering expression data from two publicly available human single-cell RNA sequencing datasets against the MitoCarta 3.0 reference database. The first dataset spanned individual cells from the oocyte through blastocyst stage, and the second compared trophectoderm and inner cell mass cells isolated from blastocysts. Mitochondria-associated gene expression was sufficient to cluster human embryos by developmental stage. A pronounced shift in expression was identified at the 4-cell to 8-cell transition, with 115 unique differentially expressed genes across the two stages immediately following this transition compared to only 5 across the two prior stages. This transcriptional upregulation precedes the known onset of oxidative phosphorylation at approximately the 32-cell stage, suggesting mitochondrial roles in early embryogenesis beyond energy production. Mitochondrially transcribed genes were the primary drivers of clustering in earlier developmental stages, while nuclear-encoded genes drove clustering at the blastocyst stage, and mitochondrial gene expression profiles partially distinguished trophectoderm from inner cell mass lineages. These findings reframe mitochondria as active participants in early human developmental programming, with implications for lineage specification, epigenetic regulation, and the optimization of in vitro embryo culture conditions.