Functional profiling and visualization of the sphingolipid metabolic network in vivo
Sphingolipids govern diverse cellular processes; their dysregulation underlies numerous diseases. Despite extensive characterizations, understanding the orchestration of the sphingolipid network within living organisms remains challenging. We established a versatile genetic platform of CRISPR-engineered reporters of 52 sphingolipid regulators, recapitulating endogenous gene activity and protein distribution. This platform further allows conditional protein degradation for functional characterization. In addition, we developed the biosensor OlyA<sup>w</sup> to detect ceramide phosphoethanolamine and visualize membrane raft dynamics in vivo. Using this platform, we established comprehensive profiles of the sphingolipid metabolic network in the brain at the transcriptional and translational levels. The highly heterogeneous patterns indicate extensive coordination between distinct cell types and regions, suggesting the brain functions as a coherent unit to execute specific steps of sphingolipid metabolism. As a proof-of-concept application, we showed cell type-specific requirements of sphingomyelinases, including CG6962/dSMPD4 and CG3376/aSMase, degrading distinct subcellular pools of ceramide phosphoethanolamine to maintain brain function. These findings establish a foundation for future studies on brain sphingolipid metabolism and showcase the utilization of this genetic platform in elucidating in vivo mechanisms of sphingolipid metabolism.
Authors
- John P. Vaughen (ORCID: https://orcid.org/0000-0002-7141-1857)
- Chia‐Lin Wu
- Wan-Syuan Lin
- Huei‐Ming Yeh (ORCID: https://orcid.org/0000-0003-0077-3284)
- Chung‐Chuan Lo (ORCID: https://orcid.org/0000-0001-7737-7250)
- Fei‐Yang Tzou (ORCID: https://orcid.org/0000-0003-3463-1436)
- Kai-Yi Hsu
- Hong-Wen Tang (ORCID: https://orcid.org/0000-0002-8347-9891)
- Chih‐Chiang Chan (ORCID: https://orcid.org/0000-0003-2626-3805)
- Ya-Hui Chou (ORCID: https://orcid.org/0000-0001-6552-8728)
- Michael Raphael Panganiban (ORCID: https://orcid.org/0000-0001-8012-1061)
- Hsin-Ling Hsieh (ORCID: https://orcid.org/0000-0002-5604-2918)
- Shirun Ho (ORCID: https://orcid.org/0000-0002-5560-1722)
- Irma Magaly Rivas‐Serna
- Vera C. Mazurak (ORCID: https://orcid.org/0000-0002-7724-6354)
- Shu‐Yi Huang (ORCID: https://orcid.org/0000-0002-3522-0500)
- C.L. Yu (ORCID: https://orcid.org/0009-0008-2235-9352)
- Christopher W. Hsu
- Cheng-Li Hong (ORCID: https://orcid.org/0009-0008-5679-5013)
- Yi Hsiao (ORCID: https://orcid.org/0009-0000-2528-0844)
- Kaile Chen
- Yi‐Jhan Li (ORCID: https://orcid.org/0009-0004-6321-6244)
- M. Thomas Clandinin (ORCID: https://orcid.org/0009-0002-1203-0752)
Institutions
- University of Alberta (CA)
- National Taiwan University (TW)
- Chang Gung University (TW)
- University of California, San Francisco (US)
- National Tsing Hua University (TW)
- Taipei Municipal YangMing Hospital (TW)
- Duke-NUS Medical School (SG)
- National Taiwan University Hospital (TW)
- Institute of Cellular and Organismic Biology, Academia Sinica (TW)
- National Cancer Centre Singapore (SG)
Publication Details
- Journal
- EMBO Reports
- Published
- 2025-11-10
- DOI
- https://doi.org/10.1038/s44319-025-00632-0
- Citations
- 1
- Primary Topic
- Sphingolipid Metabolism and Signaling
- Type
- article
- Field-Weighted Citation Impact
- 0.81