Mitochondria-specific photoactivation to monitor local sphingosine metabolism and function.

Suihan Feng,Howard Riezman,Fabrice Pa David,Sylvie Montessuit,Takeshi Harayama,Nicolas Winssinger,Jean-Claude Martinou

doi:10.7554/elife.34555

Abstract

Photoactivation ('uncaging') is a powerful approach for releasing bioactive small-molecules in living cells. Current uncaging methods are limited by the random distribution of caged molecules within cells. We have developed a mitochondria-specific photoactivation method, which permitted us to release free sphingosine inside mitochondria and thereafter monitor local sphingosine metabolism by lipidomics. Our results indicate that sphingosine was quickly phosphorylated into sphingosine 1-phosphate (S1P) driven by sphingosine kinases. In time-course studies, the mitochondria-specific uncaged sphingosine demonstrated distinct metabolic patterns compared to globally-released sphingosine, and did not induce calcium spikes. Our data provide direct evidence that sphingolipid metabolism and signaling are highly dependent on the subcellular location and opens up new possibilities to study the effects of lipid localization on signaling and metabolic fate.

Highlights

Sphingolipids are one of the major lipid species in cellular membranes of all eukaryotic cells
Using stable isotope-labeled caged sphingosine precursors, we investigated the conversion of sphingosine into ceramides and sphingomyelins after uncaging, and found that sphingosine metabolism was highly dependent on its subcellular localization
Our results clearly show that the rapid sphingosine 1-phosphate (S1P) accumulation after uncaging was catalyzed by sphingosine kinases

Summary

Introduction

Sphingolipids are one of the major lipid species in cellular membranes of all eukaryotic cells. Apart from maintaining structural properties of membranes, at least four sphingolipid metabolites, sphingosine, ceramide, sphingosine 1-phosphate (S1P) and ceramide 1-phosphate (C1P), are signaling messengers that regulate fundamental cellular processes (Aguilera-Romero et al, 2014; Hannun and Obeid, 2008; Maceyka and Spiegel, 2014; Atilla-Gokcumen et al, 2014). Due to their essential roles, aberrant sphingolipid levels are linked to a broad range of diseases, including cancers, diabetes, inflammation and neurodegeneration (Maceyka and Spiegel, 2014; Platt, 2014; Guri et al, 2017). Cellular localization is likely to be an important regulator of the metabolism and functions of sphingolipids, there is no direct evidence to demonstrate this

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