Genetic Mutations

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Emory University Scientists Demonstrate

New Pathway For Genetic Mutations In

Everyday Cell Life

Spontaneous chemical changes occurring within the DNA

of non-dividing cells

may result in the development of mutant proteins with

potentially serious

consequences for the development of degenerative

neurological diseases or

cancer. Findings by Emory University scientist

Doetsch, Ph.D., reported

in the April 2, 1999 issue of Science, provide a new

explanation for why and

how terminally differentiated cells -- those that are

no longer dividing and

replicating -- express unrepaired genetic damage.

Co-authors of the study

were graduate student Anand Viswanathan and Ho Jin

You, M.D., Ph.D.

Most of the cells within the adult human body are no

longer replicating, but are

responsible for manufacturing the proteins necessary

to carry out everyday

bodily processes. Yet most research on cellular DNA

damage and repair

mechanisms has focused on the cell replication

process, where damaged and

unrepaired DNA can result in errors when DNA is

copied before cells divide.

Dr. Doetsch and his colleagues concentrated on

non-dividing cells, which go

through a process called transcription in order to

manufacture proteins. During

transcription, the cell first makes an RNA copy of

the DNA molecule by using

an RNA polymerase -- a specialized protein that reads

the DNA genetic code

imprinted on one of the two DNA strands. The

polymerase then turns the

genetic code into an RNA genetic code. The base

sequence code on the

RNA, in turn, serves as a blueprint that " codes " for

a particular protein.

The Emory investigators studied a particular type of

spontaneous damage

occurring in cytosine, one of the four amino-acid

bases (A, T, G, and C) that

are combined in different sequences within the DNA to

make genes. In a

common chemical change, cytosine may lose one of its

components and

change to uracil, a base that is normally found only

in RNA. Since uracil (U)

acts more like the T base than it does the C base, it

causes genetic miscoding

that can lead to protein mutations.

" Scientists have known for awhile that cytosine

changing to uracil is a very

important type of genetic damage in the process of

cell division and DNA

synthesis, " Dr. Doetsch reports. " Because DNA serves

as the master

molecule that specifies coding information, if the

uracil damage is not repaired,

genetic mutations will result during replication. We

asked the same questions

about uracil damage from the point of view of

transcription, " he explains,

" which is a very different way of thinking about how

mutant proteins arise. "

Scientists already know that some genetic damages

that occur during

transcription may block the action of polymerases,

which is a signal for the

DNA repair machinery to move in and correct the

mistake. When

polymerases are not blocked, however, non-dividing

cells can continue the

work of transcription, all the while reading an

erroneous coding message.

" This base substitution error has very important

implications for the biological

consequences of genetic damage in non-dividing

cells, " Dr. Doetsch points

out. " You can imagine situations where this miscoding

could cause the cell to

manufacture a mutant protein, and if that protein is

the one controlling the fact

that the cell is not dividing, it could take the cell

from a non-growth state to a

growth state, possibly contributing to malignant

transformation in the case of

mammalian cells. "

" The findings also have implications for a number of

other things, including

age-related cell death in cells that do not normally

divide -- like neurons, for

example, which could lead to neurodegenerative

diseases. "

" This experiment was the 'proof of principle' that

this kind of pathway could

occur in live cells, " explains Dr. Doetsch. " The

extent to which this would

occur for different varieties of genetic damages will

probably depend on the

cells' ability to repair genetic damage and on what

part of the growth state or

non-growth state the cells were in. This research may

allow us to devise

explanations for how physiological changes might

occur in non-dividing cells

that are exposed to environmental agents that damage

the genome or even the

cells' own processes that spontaneously change the

DNA. "

In order to demonstrate that the uracil substitution

could occur in vivo (within

live organisms) as well as in vitro (in a test tube),

the Emory scientists used the

protein luciferase -- which emits the light that

makes fireflies glow -- as a

" reporter " to measure the effects of DNA damage on

cells.

They inserted the luciferase gene into two different

types of E. coli cells along

with a special chemical that kept the cells from

dividing. They then altered the

luciferase gene by substituting uracil for cytosine.

One kind of E. coli cell was

able to repair the uracil lesions, but the other one

lacked the appropriate

repair enzyme and was not able to correct the

mistake. As long as the damage

was repaired, no glowing luciferase protein could be

detected, but when the

cells were unable to repair the DNA damage,

luciferase was manufactured

and could be detected.

Note: This story has been adapted from a news release

issued by Emory University

Health Sciences Center for journalists and other

members of the public. If you wish to

quote from any part of this story, please credit

Emory University Health Sciences

Center as the original source. You may also wish to

include the following link in any

citation:

http://www.sciencedaily.com/releases/1999/04/990406043745.htm