Patterning and growth are fundamental features of embryonic development that must be tightly coordinated. To understand how metabolism impacts early mesoderm development, we used mouse embryonic stem-cell-derived gastruloids, that co-expressed glucose transporters with the mesodermal marker T/Bra. We found that the glucose mimic, 2-deoxy-D-glucose (2-DG), blocked T/Bra expression and abolished axial elongation in gastruloids. However, glucose removal did not phenocopy 2-DG treatment despite a decline in glycolytic intermediates. As 2-DG can also act as a competitive inhibitor of mannose in protein glycosylation, we added mannose together with 2-DG and found that it could rescue the mesoderm specification both in vivo and in vitro. We further showed that blocking production and intracellular recycling of mannose abrogated mesoderm specification. Proteomics analysis demonstrated that mannose reversed glycosylation of the Wnt pathway regulator, secreted frizzled receptor Frzb. Our study showed how mannose controls mesoderm specification in mouse gastruloids. Display omitted •2-Deoxy-D-Glucose affects mesoderm specification through mannose inhibition•De novo and salvage mannose synthesis pathways control mesoderm specification•Mannose controls mesoderm specification in gastruloids and gastrulating embryos•Mannose-dependent protein glycosylation is required for mesoderm specification Dingare et al. identified a critical role for mannose-dependent glycosylation during early mesoderm specification both in mouse embryonic stem-cell-derived gastruloids and in gastrulating mouse embryos. Blocking intracellular mannose production and its usage in glycosylation inhibited mesoderm specification and symmetry breaking in gastruloids.