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Phospholipid transmembrane domains and lateral diffusion in fibroblasts
J M el Hage Chahine1, S Cribier, P F Devaux
1Institut de Biologie Physico-Chimique, Paris, France.
Summary
Fluorescent phospholipid diffusion in fibroblast membranes reveals distinct domains. Sphingomyelin recycling occurs rapidly, dependent on ATP and microtubules.
Area of Science:
- Cell Biology
- Membrane Biophysics
- Lipid Dynamics
Background:
- The plasma membrane's lateral organization influences cellular processes.
- Understanding lipid diffusion is key to deciphering membrane function.
- Phospholipid mobility varies based on lipid type and cellular conditions.
Purpose of the Study:
- To investigate the lateral diffusion of fluorescent phospholipids in cultured Chinese hamster lung fibroblasts.
- To differentiate the diffusion characteristics of various phospholipid classes (sphingomyelin, phosphatidylcholine, phosphatidylserine).
- To explore the role of ATP and temperature in modulating lipid mobility and membrane domains.
Main Methods:
- Modulated fringe pattern photobleaching was employed to measure lateral diffusion coefficients.
- Fluorescently labeled phospholipids, including N-[6-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl-amino)caproyl] sphingosylphosphocholine (C6-NBD-SphPCho), phosphatidylcholine (C6-NBD-PtdCho), and phosphatidylserine (C6-NBD-Ptd-Ser), were used.
- Experiments were conducted at different temperatures (7°C and higher) and in the presence/absence of ATP.
Main Results:
- Choline-containing lipids (C6-NBD-SphPCho, C6-NBD-PtdCho) exhibited a consistent lateral diffusion coefficient (D1 ≈ 0.3 x 10⁻⁹ cm²/s).
- Phosphatidylserine (C6-NBD-Ptd-Ser) showed two components: a slow D1 and a fast D2 (≈ 2 x 10⁻⁹ cm²/s) present only in ATP-containing cells, indicating asymmetric leaflet distribution.
- At higher temperatures, C6-NBD-SphPCho displayed a quasi-immobilized fraction, attributed to internalized sphingomyelin, with ~50% recycling within 30 min via an ATP- and microtubule-dependent process.
Conclusions:
- The inner and outer plasma membrane leaflets form distinct domains with differing lipid mobilities.
- ATP-dependent translocation of phosphatidylserine to the inner leaflet creates a domain with significantly higher lipid mobility.
- Sphingomyelin undergoes rapid, ATP- and microtubule-dependent recycling from the plasma membrane, suggesting dynamic membrane organization.