Functional profiling and visualization of the sphingolipid metabolic network in vivo.
Journal
EMBO reports
ISSN
1469-3178
Date Issued
2025-11-10
Author(s)
Tzou, Fei-Yang
Hong, Cheng-Li
Chen, Kai-Hung
Vaughen, John P
Lin, Wan-Syuan
Hsu, Chia-Heng
Rivas-Serna, Irma Magaly
Hsu, Kai-Yi
Ho, Shuk-Man
Panganiban, Michael Raphael
Hsieh, Hsin-Ti
Li, Yi-Jhan
Hsiao, Yi
Yeh, Hsin-Chun
Yu, Cheng-Yu
Tang, Hong-Wen
Chou, Ya-Hui
Wu, Chia-Lin
Lo, Chung-Chuan
Mazurak, Vera C
Clandinin, M Thomas
Huang, Shu-Yi
Abstract
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 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.
Subjects
Brain
Cell-type Specificity
Spatial Heterogeneity
Sphingolipids
Systemic Profiling
SDGs
Type
journal article