Biology Reference
In-Depth Information
are chromatic aberrations that cause multiple wavelengths to never perfectly
overlap. In contrast, NSOM guarantees a perfect overlay between multiple
excitation wavelengths, an essential requirement to resolve the true nanoscale
landscape of cell membranes. Already in 1997, Enderle
used for the
irst time dual-colour NSOM to directly measure the association of a host
protein (protein4.1) and parasite proteins (MESA and PfHRP1) in malaria
(
et al.
)-infected dried erythrocytes.
63
As the parasitic
proteins interact with the host proteins, 100 nm sized knob-like topographical
features appear on the membrane of the host cell. To investigate the direct
interaction of host and parasite proteins, the proteins were speciically
labelled and subsequently imaged with NSOM. As expected, the luorescence
from the two labelled parasitic proteins and the labelled host protein were
found on the knob-like structures. The high-resolution of NSOM however
demonstrated that host and parasite proteins did not physically colocalized
in the same compartments.
63
The increased co-localization of individual components on the cell
membrane has been actually demonstrated on two members of the interleukin
family by combining dual-colour excitation and single-molecule detection
NSOM on dried T cells.
64
IL2R and IL15R did not interact if their organization
was monomeric. However, in their clustered form, both receptors were found
to co-localize signiicantly, suggesting that clustering of both receptors takes
place in the same nanocompartments.
Plasmodium falciparum
Interestingly, IL2R and IL15R clusters
were found to have a constant packing density albeit forming domains of
different sizes.
64
Although the receptors were found to pack at different
densities, the linear increase in the number of receptors with domain size
suggested a general
64
building block
type of assembly for these receptors
64
as
opposed to the heterogeneous packing exhibited by DC-SIGN.
60
Ianoul
et al.
have also
used dual-colour NSOM to investigate the association
β
-adrenergic receptors (
β
AR) and caveolae of the surface of dried cardiac
myocytes.
of
15-20%
β
2
ARs colocalize in caveolae. The
lack of complete colocalization of
β
2
AR with the caveolae suggested that the
diverse functional properties of the
β
2
AR could arise from its association with
multiprotein complexes of different compositions that may not be caveolar
in nature. Interestingly, the fraction of
β
2
ARs not colocalizing with caveolae
appeared proximal to it, indicating
β
2
AR complexes are pre-assembled in, or
near caveolae.
65
The study showed that
~
More conventionally used techniques such as luorescence
resonance energy transfer are unable to report on such a proximity effect at
spatial scales
65
10 nm. On the other extreme, diffraction limited techniques
such as confocal microscopy will not be able to reveal a lack of co-localization
>
Search WWH ::
Custom Search