Spatiotemporal Activation of Protein O‑GlcNAcylation in Living Cells

O-linked N-acetylglucosamine
(O-GlcNAc) is a prevalent
protein modification that plays fundamental roles in both cell physiology
and pathology. O-GlcNAc is catalyzed solely by O-GlcNAc transferase
(OGT). The study of protein O-GlcNAc function is limited by the lack
of tools to control OGT activity with spatiotemporal resolution in
cells. Here, we report light control of OGT activity in cells by replacing
a catalytically essential lysine residue with a genetically encoded
photocaged lysine. This enables the expression of a transiently inactivated
form of OGT, which can be rapidly reactivated by photo-decaging. We
demonstrate the activation of OGT activity by monitoring the time-dependent
increase of cellular O-GlcNAc and profile glycoproteins using mass-spectrometry-based
quantitative proteomics. We further apply this activation strategy
to control the morphological contraction of fibroblasts. Furthermore,
we achieved spatial activation of OGT activity predominantly in the
cytosol. Thus, our approach provides a valuable chemical tool to control
cellular O-GlcNAc with much needed spatiotemporal precision, which
aids in a better understanding of O-GlcNAc function.