Embargoed until 5 pm EST
NSF PR 02-27
- April 18, 2002
New Post-Genomic Technique Chronicles Protein Life
Real-time, multi-scale look
at life's building blocks
Scientists have developed a new molecular-tagging technique
to chronicle the development, movement and interactions
of proteins as they do their work in living cells.
The results are published in the April 19 issue of
Science by University of California, San Diego
(UCSD) School of Medicine and National Center for
Microscopy and Imaging Research (NCMIR) researchers
at the UCSD campus in La Jolla. The research is funded
in part by the National Science Foundation (NSF).
"Now that the human genome has been sequenced, the
next big push is to determine the properties of proteins
associated with those genes," said Mark Ellisman,
neuroscientist and bioengineer, director of NCMIR,
and co-author of the study.
Like Lego pieces that are used to build objects and
structures, proteins are the building blocks for cellular
activity and the development of tissues and organisms.
Proteins are constantly added to and removed from
the cellular building.
"If we want to follow this frenetic activity as it
takes place, we need comparably dynamic experimental
approaches," Ellisman said. "Furthermore, we need
techniques that allow us to view both the single protein
and the final structure while they are being produced,
assembled, modified and, finally, degraded."
To date, scientists have used marking techniques involving
intrinsically fluorescent structures to tag the protein
of interest. While extremely useful to monitor the
distribution of the protein in living specimens and
to witness some of its interactions with other cellular
components, these techniques don't allow for discrimination
between the different, time-separated stages of development
and degradation of the protein. In addition, the fluorescent
proteins are often larger than many of the proteins
they are attached to. And, they don't allow researchers
to explore the dynamics of the protein at different
resolution levels, from the larger cellular building
down to the macromolecular level of the individual
The UCSD team combined advanced microscopic capabilities
and molecular biology from the NCMIR with chemistry
and biochemistry to create a powerful, integrated
and innovative multiscale molecular tagging technology
that lets researchers genetically tag a protein with
a small binding area called a domain (six to 20 amino-acids
long), that then interacts with a variety of other
An important advantage of the new technique is its
application for electron microscopy. Most molecular
tagging techniques currently used for monitoring protein
distribution and fate in living cells are applicable
only to light microscopy, which doesn't provide enough
power to allow the exploration of fine structural
details of macromolecular structures. These older
techniques are not transferable to electron microscopic
evaluation, which is 1,000 to 10,000 times more powerful
than the light microscope, and is able to locate the
precise position of individual protein complexes.
Using the new tagging technology, the research team
was able to elucidate some aspects of gap junction
assembly and turnover in the living cell. For example,
they showed that newly synthesized connexins were
transported to the plasma membrane in small, 100 to
150 nanometer vesicles and incorporated at the periphery
of pre-existing gap junction plaques. Older connexins
were instead removed from the center of the plaque
and transported into degradative vesicles of various
Although the findings on gap junction refurbishing
were of great interest, the researchers were most
excited about the possibility of generalizing their
technique to study the life cycle of virtually any
protein system and being able to visualize these proteins
in the cell.
"This is a great example of how multidisciplinary teams,
composed of talented individuals who each bring their
own special expertise into the mix, can address modern
biological problems in ways that none alone could
do," said Eve Barak, program director in NSF's division
of molecular and cellular biosciences.
The Canadian Institutes of Health Research, the U.S.
National Institutes of Health, and the Howard Hughes
Medical Institutes also provided funding for the study.