11q Gene, ATM, Linked to Ability to Repair Damaged DNA

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National Institute of Arthritis

and Musculoskeletal and Skin Diseases

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FOR IMMEDIATE RELEASE

Wednesday, June 27, 2007

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Scientists Discover Role of Enzyme in DNA Repair

Scientists from the National Institute of Arthritis and

Musculoskeletal and Skin Diseases (NIAMS), National Cancer Institute

(NCI), and Integrative Bioinformatics Inc. have made an important

discovery about the role of an enzyme called ataxia telangiectasia

mutated protein (ATM) in the body's ability to repair damaged DNA.

NIAMS and NCI are part of the National Institutes of Health (NIH).

When DNA within a cell is damaged, the cell's protective mechanism

must do one of two things: repair the defect or " commit suicide, "

says Casellas, Ph.D., an investigator in NIAMS' Molecular

Immunology and Inflammation Branch and leading author of a new paper

describing the discovery. But the way in which the cell performs

these protective functions has been largely a mystery, says

Casellas, whose research is beginning to unravel this mystery.

Casellas' research focuses largely on certain genes that are

deliberately broken and repaired as part of the immune response.

Through a tightly controlled process of breaking and rejoining DNA

segments, immune system cells called B lymphocytes are able to

produce tens of millions of different types of antibodies to fight

almost limitless types of invaders. This process of genetic

recombination requires the activity of repair enzymes, which must be

able to recognize and repair breaks in tightly wrapped and

inaccessible DNA. During immunoglobulin gene recombination, DNA is

rendered accessible by the process of transcription, which unzips

double-stranded DNA as part of the conversion of genetic information

into functional proteins.

While transcription ensures accessibility to DNA lesions, Casellas

wondered how it was possible for repair enzymes to do their job if

transcription continued once DNA had been damaged. " Imagine a piece

of DNA as a zipper, " he says. " The head of the zipper (the

transcription complex) will repeatedly go through the two

interlocked sides, coming to the broken part, and eventually falling

off. One could imagine that this unzipping activity might interfere

with the mechanism that is trying to repair the damaged DNA. "

Casellas hypothesized that once DNA lesions were generated, a

regulatory activity would shut down transcription until repair

enzymes corrected the damage. But because B lymphocyte cells are

relatively scarce, Casellas and his colleagues chose to focus their

investigation on a more abundant family of genes, known as ribosomal

genes, as a substitute. They attached a green fluorescent protein to

Polymerase I, a key component in the machinery that transcribes

these genes, and were able to visualize the activity of this enzyme

using microscopy. They then used a particular laser attached to the

microscope to introduce DNA breaks at sites where the polymerase was

active. This microscopy approach was developed by NCI's

Kruhlak, Ph.D., first author in the report. Using the ProcessDB

software developed by Integrative Bioinformatics Inc, Phair,

Ph.D. developed a computer model that allowed the authors to test

their hypothesis and show that while transcription continued in the

cells with uninjured DNA, it came to a halt within 5 minutes at

sites where the DNA had been damaged.

While it was possible that the DNA lesions themselves physically

interfered with transcription, the authors hypothesized that repair

enzymes recruited by the damage could shut down the transcription

machinery polymerase. To test this hypothesis, they repeated the

experiment in cells that were deficient in a variety of repair

proteins. Most deficiencies did not appear to affect the arrest;

however, in cells that were missing one of three repair proteins

factors — ATM, Nbs1 or MDC1 — transcription continued even after

damage was induced.

" What these results told us was that these proteins were responsible

for shutting down the transcription machinery near sites of DNA

damage. This activity perhaps ensures repair in an undisturbed

environment. If this is indeed the case, one could suspect that in

the absence of these factors, repair is compromised, leading to

genetic aberrations, " Casellas says. Indeed, scientists already know

that people deficient in ATM develop such genetic abnormalities,

cell transformation and tumor development. Although it's too soon to

say whether these laboratory discoveries will translate into

clinical use, Casellas is enthused about the work. " With this new

technology we can visualize for the first time the interplay between

complex mechanisms such as DNA repair and gene transcription, not in

a test tube, but in living cells and in real time. This approach

will help us unravel the inner molecular pathways of our cells in

health and disease, such as cancer. "

The National Institutes of Health (NIH) — The Nation's Medical

Research Agency — includes 27 Institutes and Centers and is a

component of the U.S. Department of Health and Human Services. It is

the primary federal agency for conducting and supporting basic,

clinical and translational medical research, and it investigates the

causes, treatments, and cures for both common and rare diseases. For

more information about NIH and its programs, visit www.nih.gov.

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Reference: Kruhlak M, et al. The ATM repair pathway inhibits RNA

polymerase I transcription in response to chromosome breaks. Nature

2007;447:730-734.