First Light Shed On Gene Repair By Nanoscale Microscope

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First Light Shed On Gene Repair By Nanoscale Microscope

http://www.medicalnewstoday.com/medicalnews.php?newsid=56587

Proteins called H2AX act as " first aid " to DNA, among other roles.

For the first time, scientists using the world's most powerful light

microscope (the only one of its kind in the Americas) have seen how

H2AX is distributed in the cell nucleus: in clusters, directing the

first aid/repair after DNA injuries to the region where it is really

needed.

Many biological processes lie out of the visual reach of scientists.

The benefits of high-resolution electron microscopy are often offset

by disruptive sample preparation requirements. Light microscopy

allows easier sample prep and observations of living cells, but it

has limited resolution. By manipulating how light waves behave,

however, biophysicists are expanding the limits of light microscopy,

and one of the latest advances--the 4Pi microscope--provides never-

before-seen views of cellular components, including structures

within the nucleus.

In a paper published in the Proceedings of the National Academy of

Sciences, Joerg Bewersdorf of the Institute for Molecular Biophysics

at The Laboratory, with collaborators of the

UMass Medical School and Leica Microsystems and Kendall Knight of

the UMass Medical School, used the 4Pi microscope to examine the

cellular response to a type of severe damage to the genetic

material, DNA double-strand breaks. Such breaks provoke a rapid and

highly coordinated series of events to identify and repair the

damage. The response is critical, and there is an increased risk for

cancer, developmental abnormalities and immunological problems when

components of the repair processes are defective.

Traditional microbiological and genetics techniques can shed light

on the molecular pathways of repair, but they don't address the

astonishingly complex three-dimensional structure of the genetic

material in the nucleus. 4Pi microscopy allows researchers to

actually see the response in three dimensions, at resolutions down

to 100 nm. Therefore, the role of the physical structures in various

processes within the nucleus can now be visualized.

" The general application of these methods will provide unprecedented

insights into cellular molecular events, " said Bewersdorf. " This

study represents a significant advance in our ability to visualize

and quantify nuclear proteins in 3D. "

Bewersdorf, and Knight examined a protein called H2AX, a

kind of histone. Histones are structural proteins that act as spools

around which DNA is wound, and they can also play roles in gene

regulation and gene repair. H2AX is an early responder to DNA

damage, and its change to what is known as gamma-H2AX is important

for the coordination of signaling and repair activities. But it had

not been known how H2AX is distributed throughout the nucleus or why

its conversion to gamma-H2AX is limited to within a short distance

of a break site. By using selective staining techniques and 4Pi

visualization, Bewersdorf and colleagues determined that H2AX is

distributed in distinct clusters uniformly throughout the nucleus

and that the structure of these clusters may determine the

boundaries of where gamma-H2AX spreads in response to a break.

" The clusters may provide a platform for the immediate and robust

response observed following DNA damage, " said Bewersdorf. " Moving

forward, we will analyze the localization of the H2AX clusters

relative to other nuclear components. "

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The Laboratory is an independent, nonprofit research

institution based in Bar Harbor, Maine, with facilities in West

Sacramento, Calif. Its research staff of more than 450 investigates

the genetic basis of cancers, heart disease, osteoporosis,

Alzheimer's disease, glaucoma, diabetes, and many other human

diseases and disorders, as well as normal mammalian development and

bioinformatics. The Laboratory is also the world's source for nearly

3,000 strains of genetically defined mice, home of the Mouse

GenGenome Database and many other publicly available information

resources, and an international hub for scientific courses,

conferences, training and education.