NATURE CELL BIOLOGY VOLUME 9 | NUMBER 1 | JANUARY 2007
Presentation by: Christian Stern
Lipids and proteins are
organized in domains
(rafts)
Cell membrane
Cell membrane
Phospholipids
Sphingolipids
These compounds play important roles in signal transmission and cell recognition
Glycolipids
carbohydrate-attached lipids
Role: to provide energy and also serve as markers for cellular recognition
Cholesterol
•essential component of mammalian cell membranes, where it is required to
establish proper membrane permeability and fluidity
•Stabilizes the membrane
•Important for ex- and import of signaling molecules
•insoluble in water
•95 % intracellular
Cell membrane
Lipid raft organization scheme
A
B
Intracellular space or cytosol
Extracellular space or vesicle/Golgi apparatus lumen
1. Non-raft membrane
2. Lipid raft
3. Lipid raft associated transmembrane protein
4. Non-raft membrane protein
5. Glycosylation modifications (on glycoproteins and glycolipids)
6. GPI-anchored protein
7. Cholesterol
8. Glycolipid
Lipid raft organization scheme
Features:
•Biggest difference from normal membrane
lipid composition
•contains twice the amount of cholesterol than
normal membrane
•Cholesterol is the dynamic "glue" that holds the
raft together
•enriched in sphingolipids such as sphingomyelin
(50% more than membrane)
Lipid rafts. What’s their job?
Jobs:
Organize cellular processes by:
•Assembling of signaling molecules
•Influencing membrane fluidity
•Influencing membrane protein trafficking
•Regulating neurotransmission and receptor trafficking
History
Until 1982, it was widely accepted that phospholipids and membrane proteins were randomly
distributed in cell membranes, according to the Singer-Nicolson fluid mosaic model, published in
1972.
1982 Karnovsky et al.
“Lipids in a more ordered way” or Concept of lipid domains
1997 Simons and Ikonen
“Lipid rafts”
Problems:
Many theories and models existed:
--from lipid ‘shells’ to the idea that the membrane is a collection of contiguous rafts
with fluid inclusions—
reflects the difficulty in structurally characterizing the cell membrane
History
Until 1982, it was widely accepted that phospholipids and membrane proteins were randomly
distributed in cell membranes, according to the Singer-Nicolson fluid mosaic model, published in
1972.
1982 Karnovsky et al.
“Lipids in a more ordered way” or Concept of lipid domains
1997 Simons and Ikonen
“Lipid rafts”
2006 Keystone Symposium of Lipid rafts:
Latest Definition
Lipid Rafts:
“small (10-200nm), heterogeneous, highly dynamic,
sterol- and sphingolipid-enriched domains that
compartmentalize cellular processes.
Small rafts can sometimes be stabilized to form larger
platforms through protein-protein interactions"
Enough of history
Let’s have a closer look on Lipid Rafts!
Organization of Lipid Rafts on the cell membrane
Concentration of integral Proteins is very high:
Quinn et al., 1984: density of integral membrane proteins in the ER and Golgi:
~30,000 molecules per μm2
A membrane is a lipidprotein composite, rather
than a dilute solution of
protein in an lipid solvent
Lipid Rafts are not all the same.
They vary in composition, size, lifetime and functionality
So, how to characterise and name them?
Lipid Rafts in Caveolae
• invaginations of the plasma membrane in many vertebrate cell types
• contain clusters of glycosphingolipids, GPI-anchored proteins and a high concentration of
cholesterol
• well characterized 50–100-nm nanodomain
• detergent resistant
• can be readily identified in most cells by the marker caveolin-1
Function:
•have several functions in signal transduction.
•They are also believed to play
a role in endocytosis,
oncogenesis, and the uptake
of pathogenic bacteria and
certain viruses
Bender et al., 2002
Shells
•Smaller than 10 nm
•specific classes of plasma-membrane proteins bind to preassembled complexes of
cholesterol and sphingolipids
•The dynamics of exchange may range from the short-lived classical boundary layer lipids
(~1–10 s), in which shell lipids may rapidly interchange with non-shell lipids, to long-lived
lipids that are tightly bound through specific lipid–protein interactions
“….lateral organization probably results from preferential packing of
sphingolipids and cholesterol into moving platforms, or rafts, onto which
specific proteins attach within the bilayer.”
Simons and Ikonen, 1997
Nanodomains
• 50–200 nm in dimension in the outer leaflet of the plasma membrane
•Many proteins organized in cluster
Microdomains
•~1µm or bigger in dimension
•Visible in Light and Electron Microscopy
•MD of detergent-resistant transporters were stable in growing yeast for more than 10 min
•MD in smooth muscle cells and macrophages were stable for tens of seconds
•Exist in various cells but the stabilizing factors are largely unknown
Do they have Evidences?
Examples of lipid and protein domains in cell membranes
Single domains, enriched
in the fluorescent lipid
analogue DMPECy5
Cholesterol-rich domains
indirect
immunofluorescence
microscopy
Lipid domains with greater
relative order than the
bulk membrane, visualized
in living macrophages with
the fluorescent probe,
Laurdan.
The warmer pseudocolours represent
more ordered regions
Domains formed by the
proton–argenine
symporter
transporter (Can1p–GFP)
The scale bars represent 1 μm in a, and 5 μm in c and d.
Methods for detecting and characterizing membrane domains
Problems in Lipid Raft research:
The lipid-raft research is at a technical impasse, largely because the tools
to study biological membranes as liquids structured in space and time are
rudimentary.
Biophysical tools to study membrane domains in biological membranes
Possibility to get significant results:
When domains have a minimum size of a few protein diameters and a
minimum lifetime of ~microseconds
Problems:
Small and transient domains  Nanoscale Level
Technology with sufficient simultaneous spatial and temporal resolution is not available
Best choice:
Fluorescense microscopy  high sesitivity, easy application to single, living cells
• Secondary ion mass spectrometry (SIMS)
• Atomic force microscopy (AFM)
• Near field scanning microscopy (NSOM)
- only with quick frozen specimens
- bad time resolution
- bad time resolution
•Scattering techniques
- only used in model-membrane studies
Other challenges:
‚energy suply‘ to biological membranes
Lipids and proteins are constantly added and removed.
 changes the whole membrane organization
changes the interactions between lipids and proteins
This lipid flux is necessary to understand the structure and function of nanodomains
Open Questions
What are the effects of membrane protein levels?
What is the physiological function of lipid rafts?
What effect does flux of membrane lipids have or raft formation?
What effect do diet and drugs have on lipid rafts?
What effect do proteins located at raft boundaries have on lipid rafts?
…
Needed:
• Appropriate artificial membranes
to study domain properties such as formation, size, lifetime and morphology
Problem: complexity of bio membranes, energetics
• Computational models
 simulations of the large assemblies of molecules found in domains
Take home message:
A lipid raft is a cholesterol and sphingolipid-enriched domain or platform found in cell
membranes. These specialized membrane microdomains compartmentalize cellular
processes by serving as organizing centers for the assembly of signaling molecules,
influencing membrane fluidity and membrane protein trafficking, and regulating
neurotransmission and receptor trafficking. Lipid rafts are more ordered and tightly packed
than the surrounding bilayer, but float freely in the membrane bilayer.
Keystone symposium 2006: “small (10-200nm), heterogeneous, highly dynamic, sterol- and
sphingolipid-enriched domains that compartmentalize cellular processes. Small rafts can
sometimes be stabilized to form larger platforms through protein-protein interactions”
Organized in Shells (1-10 nm), Nanodomains (10-100 nm) and Microdomains (bigger than
100 nm)
Caveolae: well characterized 50–100-nm invaginations of the plasma membrane. Contains
clusters of glycosphingolipids, GPI-anchored proteins and a high concentration of
cholesterol. Can be identified via the marker caveolin-1. Cavveolaes have several functions
in signal transduction. They are also believed to play a role in endocytosis,
oncogenesis, and the uptake of pathogenic bacteria and certain viruses.
Thanks for your attention! 