Qiu, J.; Sturmey, R. G.

Glucose uptake and metabolism increase markedly during the transition from morula to blastocyst. However, the functional roles of glucose during this process in bovine embryos remain incompletely understood. Here, we demonstrate that glucose specifically modulates trophectoderm (TE) differentiation without affecting the inner cell mass (ICM), acting through Hippo signalling via the hexosamine biosynthetic pathway (HBP) and the pentose phosphate pathway (PPP) rather than glycolysis. Consistently, scRNA-seq analysis reveals that glycolysis-related genes show only stage-specific changes, whereas HBP- and PPP-related genes exhibit both stage- and lineagespecific differences between TE and ICM. Furthermore, we found that glucose deprivation does not impair pyruvate uptake or intracellular pyruvate levels in blastocysts, yet it significantly depletes the terminal metabolites of the HBP. Together, these findings reveal that glucose-dependent regulation of TE differentiation is conserved in bovine embryos.