mRNA translational dynamics mediates totipotent-like reprogramming.
Embryonic stem cells (ESCs) are characterized by their dual capacity for self-renewal and differentiation into all cell types of the embryonic lineage. A subpopulation known as 2-cell-like cells (2CLCs), which recapitulate key molecular and metabolic features of totipotent 2-cell blastomeres, has been identified within cultured mouse ESC populations. While transcriptional regulation, epigenetic modifications, and chromatin reorganization are known to be critical for the reprogramming of pluripotent ESCs into a totipotent-like state, the role of translational control in this process remains poorly understood. Using an inducible 2CLC model, we performed transcriptome-wide profiling of mRNA translation and found that global translation efficiency dynamically decreases during the early phase of totipotent-like reprogramming, correlating with reduced TORC1 signaling and translation initiation. In the later phase, although overall mitochondrial mass declines, mitochondrial translation is selectively upregulated and exhibits high translational efficiency. Importantly, pharmacological inhibition of mitochondrial translation suppressed the expression of canonical 2-cell transcripts and impaired the transition from ESCs to 2CLCs. Together, these results demonstrate that coordinated regulation of both cytosolic and mitochondrial translation during totipotent-like reprogramming, offering a new perspective for understanding cell fate determination.