Spatiotemporal control of subcellular O-GlcNAc signaling using Opto-OGT

Ong, Q.; Lim, L. T. R.; Goh, C.; Liao, Y.; Chan, S. E.; Lim, C. J. Y.; Kam, V.; Yap, J.; Tseng, T.; Desrouleaux, R.; Wang, L. C.; Ler, S. G.; Lim, S. L.; Kim, S.; Sobota, R. M.; Bennett, A. M.; Han, W.; Yang, X.

doi:10.1101/2024.05.12.593740

bioRxiv · 10.1101/2024.05.12.593740

Spatiotemporal control of subcellular O-GlcNAc signaling using Opto-OGT

Ong, Q.·Lim, L. T. R.·Goh, C.·Liao, Y.·Chan, S. E.·Lim, C. J. Y.·Kam, V.·Yap, J.·Tseng, T.·Desrouleaux, R.·Wang, L. C.·Ler, S. G.·Lim, S. L.·Kim, S.·Sobota, R. M.·Bennett, A. M.·Han, W.·Yang, X.

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Abstract

The posttranslational modification of intracellular proteins through O-linked {beta}-N-acetylglucosamine (O-GlcNAc) is a conserved regulatory mechanism in multicellular organisms. Catalyzed by O-GlcNAc transferase (OGT), this dynamic modification plays an essential role in signal transduction, gene expression, organelle function, and systemic physiology. Here we present Opto-OGT, an optogenetic probe that allows for precise spatiotemporal control of OGT activity through light stimulation. By fusing a photosensitive cryptochrome protein to OGT, Opto-OGT can be robustly and reversibly activated with high temporal resolution by blue light and exhibits minimal background activity without illumination. Transient activation of Opto-OGT results in mTORC activation and AMPK suppression which recapitulate nutrient-sensing signaling. Furthermore, Opto-OGT can be customized to be localized at specific subcellular sites. By targeting OGT to the plasma membrane, we demonstrate downregulation of site-specific AKT phosphorylation and signaling outputs in response to insulin stimulation. Thus, Opto-OGT is a powerful tool to define the role of O-GlcNAcylation in cell signaling and physiology.

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Ong, Q., Lim, L. T. R., Goh, C., Liao, Y., Chan, S. E., Lim, C. J. Y., Kam, V., Yap, J., Tseng, T., Desrouleaux, R., Wang, L. C., Ler, S. G., Lim, S. L., Kim, S., Sobota, R. M., Bennett, A. M., Han, W., Yang, X.. 2024-05-14. Spatiotemporal control of subcellular O-GlcNAc signaling using Opto-OGT. https://doi.org/10.1101/2024.05.12.593740

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