Pulsed-SILAC in single mouse embryos reveals early embryonic protein synthesis dynamics and phosphosite regulation

Schmidt, L. M.; Lechner, M.; Martin-Gonzalez, J.; Halliwell, J. A.; Mikicic, I.; Lowndes, M. P.; Hendriks, I. A.; Stuber, K. A.; Lerdrup, M.; Hoffmann, E.; Olsen, J. V.

doi:10.64898/2026.09.24.754052

bioRxiv · 10.64898/2026.09.24.754052

Pulsed-SILAC in single mouse embryos reveals early embryonic protein synthesis dynamics and phosphosite regulation

Schmidt, L. M.·Lechner, M.·Martin-Gonzalez, J.·Halliwell, J. A.·Mikicic, I.·Lowndes, M. P.·Hendriks, I. A.·Stuber, K. A.·Lerdrup, M.·Hoffmann, E.·Olsen, J. V.

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Abstract

Early embryogenesis relies extensively on maternally deposited products until zygotic genome activation, yet the dynamics for the synthesis of new proteins in mammalian embryos remains poorly characterized. To address this, we applied pulsed stable isotope labelling by amino acids in cell culture (pSILAC) combined with narrow-window data-independent acquisition mass spectrometry to single mouse oocytes and embryos to resolve de novo protein synthesis during early embryogenesis. This revealed that the maternal proteome is not a static reservoir, with components of the subcortical maternal complex and amino acid transporters SLC7A1/2 being actively synthesized during the earliest developmental stages. Furthermore, phosphoproteomic analysis identified hundreds of previously unreported phosphosites and extensive regulation during the oocyte-to-embryo transition. Notably, phosphorylation of the PRC2-interacting KLP motif of EZHIP emerged as a potential regulatory mechanism, with modification of this region reducing EZHIP-PRC2 interaction and coinciding with H3K27me3 remodelling. Together, single embryo pSILAC revealed a maternal proteome that is continuously synthesized, recycled, and post-translationally regulated during early embryogenesis.

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Schmidt, L. M., Lechner, M., Martin-Gonzalez, J., Halliwell, J. A., Mikicic, I., Lowndes, M. P., Hendriks, I. A., Stuber, K. A., Lerdrup, M., Hoffmann, E., Olsen, J. V.. 2026-09-25. Pulsed-SILAC in single mouse embryos reveals early embryonic protein synthesis dynamics and phosphosite regulation. https://doi.org/10.64898/2026.09.24.754052

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