Cytochemists study membranes either by first splitting them along the plane of the two
lipid leaflets, inbetween the lipid bilayer. Or they may label transmembrane proteins and watch the
dynamics of binding, membrane fluidity, or membrane uptake. Another favorite approach to
membrane function is a study of specialized junctions
formed at key membrane sites. The first group of studies in this presentation will focus
on membrane fluidity. This is made possible by the rotation of lipids and proteins in the
bilayer. Your text reading for this section is found on pages 498-501 in Alberts et
al. Molecular Biology of the Cell, Third Edition, Garland Publishing, N.Y., 1994
Return to Menu
Membrane proteins (like lipids) rotate about an axis perpendicular to the plane of the bilayer. However, they may also move laterally. The following experiments show that not only do membrane proteins move, they can rearrange. The first experiment was done on fused cells. One group of cells came from a mouse line and the other from a human cell line. The investigators obtained antibodies to proteins in the membrane and conjugated these antibodies to fluorescein (green) or rhodamine (red) fluorescent compounds. The cells were then fused, forming a "heterocaryon" (note the two nuclei). Then, the two sets of antibodies were applied. Initially the cells were labeled half red and half green. However, after 40 min at 37 C, the labels mixed and the cells showed mixing of the two dyes. Often this mixing turns out to be white fluorescence. This figure was taken from Alberts et al. Molecular Biology of the Cell, Third Edition, Garland Publishing, N.Y., 1994, Figure 10-34.
In the next experiment, the label on the membrane is "bleached" in one area
with a laser beam. The figure to the right and the figure below shows the results.
Bleaching removes the label on the protein and results in a clear area (area of no label)
in a completely labeled cell (see following figure). Then, one can watch the recovery in
the bleached area as the the proteins + label diffuse into the irradiated patch. The
following large figure shows the bleached area as white and the recovered patch as a
lighter red. One can measure recovery in the bleached region with time after bleaching.
This is shown in the cartoon to the right. The fluorescence decreases dramatically right
after laser bleaching. Then, there is a gradual recovery with time. The figures were
taken from Alberts et al. Molecular Biology of the Cell, Third Edition, Garland
Publishing, N.Y., 1994, Figure 10-36
Return to Menu
Polarized cells have one side facing a special
region in an organ and the other side facing another region. For example, in the
intestine, the lining cells are polarized. The side responsible for absorption of
nutrients faces the lumen. The lateral surfaces are specialized with barriers to entry and
the basal surface is specialized to send nutrients and water to the blood stream. Each
specialization is conferred by proteins associated with the membrane. This figure was
taken from Alberts et al. Molecular Biology of the Cell, Third Edition, Garland
Publishing, N.Y., 1994, Figure 10-37
One can prevent mobility of proteins in a lipid bilayer either by structures in or on
the cell, or by experimental means. For example, as we will see in the next section on tight junctions, apical proteins are prevented
from entering the lateral or basal domains by a tight junction that actually fuses the
membranes in places.
Return to Menu
You can also tether proteins as shown in this diagram from your text. They could be
tethered by interactions with macromolecules outside the cell (B) (like extracellular
matrix), or inside the cell (for example, cytoskeletal filaments) (C). Or they can
interact with proteins on another cell and be tethered (D). This figure was taken from
Alberts et al. Molecular Biology of the Cell, Third Edition, Garland Publishing, N.Y.,
1994, Figure 10-39
Return to Menu
Receptors are a special class of transmembrane protein that can be detected by their
ligand. They have the reactive group projecting from the surface that binds a specific
ligand. This cartoon shows the interaction between the ligand secreted by one cell (such
as a hormone) and its receptor. The lower cartoon shows interactions between two cells,
one with a receptor for a protein on the surface of the other cell. This kind of reaction
may occur when natural killer cells recognize a foreign cell.
In a later lecture, we will learn the sequence of reactions that follow receptor binding. The following cartoon diagrams them. Usually the binding causes aggregation (patching) of the receptor-ligand complex. Then the aggregates accumulate at one pole (opposite to that of the centrioles) in a "cap" (called capping). At this point they may be internalized and either degraded or used for internal functions. All of this is possible because the membrane is fluid and allows lateral movement of the proteins.This figure was taken from Alberts et al. Molecular Biology of the Cell, Third Edition, Garland Publishing, N.Y., 1994, Figure 10-35
This figure shows how a receptor-ligand complex
can be detected at the light microscopic level. The ligand is attached to biotin and then
it binds the target cell. It is detected by an avidin peroxidase conjugate which is
visualized by a reaction for the enzyme peroxidase. This protocol is called "affinity
cytochemistry". In this figure, the biotinylated ligand is labeled black (note that
it is capping on one of the cells). The orange label defines the protein in the cell by
immunocytochemistry. For more information, consult our Cytochemistry Web page
Continue studies of membrane structure.
Return to Menu