Abstract

Each intracellular organelle critically depends on maintaining its specific lipid composition that in turn contributes to the biophysical properties of the membrane. With our knowledge increasing about the organization of membranes with defined microdomains of different lipid compositions, questions arise regarding the molecular mechanisms that underlie the targeting to/segregation from microdomains of a given protein. In addition to specific lipid-transmembrane segment interactions as a basis for partitioning, the presence in a given microdomain may alter the conformation of proteins and, thus, the activity and availability for regulatory modifications. However, for most proteins, the specific lipid environment of transmembrane segments as well as its relevance to protein function and overall membrane organization are largely unknown. To help fill this gap, we have synthesized a novel photoactive sphingolipid precursor that, together with a precursor for phosphoglycerolipids and with photo-cholesterol, was investigated in vivo with regard to specific protein transmembrane span-lipid interactions. As a proof of principle, we show specific labeling of the ceramide transporter with the sphingolipid probe and describe specific in vivo interactions of lipids with caveolin-1, phosphatidylinositol transfer protein beta, and the mature form of nicastrin. This novel photolabile sphingolipid probe allows the detection of protein-sphingolipid interactions within the membrane bilayer of living cells.

Highlights

  • Each intracellular organelle critically depends on maintaining its specific lipid composition that in turn contributes to the biophysical properties of the membrane

  • As a proof of principle, we demonstrate that addition to cells of [3H]D-erythro-photoSph results in UV-dependent labeling of the ceramide transporter (CERT)

  • Exploring novel specific protein-sphingolipid interactions in vivo, we find a direct interaction of sphingolipids with caveolin-1 and phosphatidylinositol transfer protein (PI-TP) b

Read more

Summary

Introduction

Each intracellular organelle critically depends on maintaining its specific lipid composition that in turn contributes to the biophysical properties of the membrane. We report the synthesis of a novel radioactively labeled photoactivatable sphingosine derivative, [3H]D-erythro-photo-sphingosine (photoSph), which allows the specific study of protein-sphingolipid interactions in vivo. Metabolism of this lipid precursor generates radioactive photolabile sphingolipids with a complete range of molecular species and allows the analysis of protein-lipid interactions within the lipid bilayer.

Results
Conclusion

Talk to us

Join us for a 30 min session where you can share your feedback and ask us any queries you have

Schedule a call

Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.