silencing the CMT gene too, perhaps!

[Guest guest]

New gene-silencing technique News-Medical.Net 5-Aug-2004

A new gene-silencing technique that takes place in the nucleus of human

cells, has been demonstrated by researchers at the University of

California, San Diego (UCSD) School of Medicine and the VA San Diego

Healthcare System.

The technique, called transcriptional gene silencing (TGS), provides a

new research tool to study gene function and, if continuing studies

prove the concept, it could potentially become a method for therapeutic

modification of the expression of disease-producing genes.

Selected for speedy publication in the August 5, 2004 edition of Science

Express, the study describes, for the first time, the ability to shut

down a gene literally before it is born in the nucleus of a cell. The

benefit over previous gene-silencing techniques is that the nuclear

version may have the potential to last considerably longer than current

methods that act in the cytoplasm, the cellular area outside the

nucleus.

The new technique, and older gene-silencing methods that have given rise

in recent years to a multi-million dollar pharmaceutical industry,

utilizes ribonucleic acid

(RNA), the cousin of DNA. Specifically, researchers use synthetic, short

pieces of RNA called short interfering RNA (siRNA), to shut down genes.

The synthetic versions are patterned after naturally occurring siRNA in

the body that may act as a defense against gene sequences that come from

viruses or other genetic parasites.

The study’s senior author, J. Looney, M.D., associate professor of

medicine at UCSD and the VA San Diego Healthcare System, said the new

technique provides a new tool for research investigation aimed at

elucidating the effects of different genes, and has the potential to

modify gene expression in disease, such as knocking out expression of

genes required for tumor growth. He cautioned, however, that further

studies are needed to prove the general applicability of this concept.

An understanding of siRNA begins with a look at theway by which genes

work. First, a “promoter” region within the gene must be active in order

to allow the genetic information encoded in the DNA to be copied

(transcribed) into a single strand of RNA called messenger RNA (mRNA).

During normal transcription, the mRNA leaves the nucleus and travels to

the cytoplasm of the cell, where it works with another cellular

component called the ribosome to make proteins.

Technology developed about four years ago introduced synthetic siRNA

into the cytoplasm of cells to silence specific genes. This technique

was called post-transcriptional gene silencing (PTGS). However, PTGS is

transient, with siRNA lasting only a few days in the cytoplasm. Although

this is enough time for short-term research projects, the use of siRNA

for therapeutic applications, such as treatment for viral infections

like HIV, probably require multiple siRNA treatments or the use of a

gene therapy approach.

UCSD researchers used either lentiviral vectors (molecular ferries) to

open up the nuclear membrane, or special transfection reagents which

direct the transfected synthetic siRNA to the nucleus. This allowed

siRNA access to the promoter, where it stopped the first part of the

gene-making process called transcription, before it began. Previous

research with siRNA used in the nucleus of plants has indicated that

this effect can be long lasting, giving rise to the hope that it will be

similarly long lasting in humans. Until now, however, scientists have

been unable to detect activity of siRNA directed against gene promoters

in the nucleus of human cells.

V. , Ph.D., the study’s first author and a post-doctoral

fellow in Looney’s lab, noted that “theoretically, one could envision

targeting virtually any gene at the level of the promoter and silencing

that gene. This has implications in most biological processes in which

one would want to down regulate the expression of a gene, such as those

genes involved in virus infections such as HIV, as well as human cancers

and certain genetic disorders.”

In continuing studies, the Looney lab and others in the country will

investigate this new method’s persistence within the human-cell nucleus,

its successful targeting of human promoters, and whether it is feasible

to use this technique to inhibit HIV or other viruses.

In addition to Looney and , the authors were Simon W.-L. Chan,

Ph.D., UCLA Department of Molecular, Cell and Developmental Biology; and

E. sen, Ph.D., UCLA Department of Molecular, Cell and

Developmental Biology, and the UCLA Molecular Biology Institute.

http://health.ucsd.edu

[Guest guest]

Here was a related article on gene silencing and epigenetics in

general. If anyone is interested, I'll go back and find the

background article for this.

July 23, 2004

SCIENCE JOURNAL

By SHARON BEGLEY

How a Second, Secret Genetic Code Turns Genes On and Off

With some identical twins, a slightly different hairline or tilt of

the eyebrows reveals who's who. But for this pair of brothers, the

distinguishing trait is more obvious -- and more tragic: One has had

schizophrenia since he was 22. His identical twin is healthy.

Like all identical twins, the brothers carry the exact same sequence

of three billion chemical letters in their DNA (this is the sequence

that the Human Genome Project famously decoded). So there was no

sense in looking for a genetic difference among these usual

suspects. But because schizophrenia is at least partly heritable,

scientists suspected that the twins' DNA had to differ somewhere.

As I explained in last week's column1, there is a second, and

largely secret, genetic code beyond the well-known one of As, Ts, Cs

and Gs that make up the human genome sequence. Called " epigenetic, "

this second code acts like the volume control on a TV remote to

silence or turn up the activity of genes. It was in these epigenetic

changes that Arturas Petronis of the Centre for Addiction and Mental

Health, Toronto, and his colleagues found the difference between the

twins.

In the healthy brother, the scientists reported in 2003, molecular

silencers sit on a gene that affects dopamine, a brain chemical. In

the twin with schizophrenia, the molecular silencers were almost

absent, so the gene was operating at full volume. In another pair of

identical twins, both of whom have schizophrenia, the silencers were

also missing.

A pattern had emerged: missing silencers are linked to schizophrenia, perhaps

because that state of DNA triggers a profusion of dopamine receptors. Measured

by this second genetic code, " the twin with schizophrenia was closer to these

unrelated men than to his own twin brother, " says Dr. Petronis.

This sort of DNA difference would never be detected with standard

genetic tests, which scan for typos -- mutations -- in DNA

sequences. But with the explosion in epigenetics, biologists are now

realizing that changes that silence and unsilence genes, but leave

the DNA sequence untouched, might explain complex diseases better

than the sequence variations that have been the holy grail for 50

years.

Take cancer. Cells harbor tumor-suppressor genes that keep them from

becoming malignant. But even when there is no mutation in tumor-

suppressor genes, a cell can become cancerous. That left scientists

scratching their heads. It turns out that tumor-suppressor genes can

be abnormally silenced, by epigenetics, even when their DNA sequence

(which genetic tests for cancer detect) is perfectly normal. So far,

scientists have identified at least 60 presumably beneficial genes

that are abnormally silenced in one or another cancer, allowing

tumors to take hold.

Conversely, an unsilencing of cancer-causing genes allows these

rogue genes to turn on, Feinberg of s Hopkins School of

Medicine, Baltimore, and colleagues found. That triggers lung and

colon cancers. " About 3% of genes seem to be abnormally silenced or

activated in cancers, " says Dr. Feinberg.

Last month, a Berlin-based biotech, Epigenomics AG, reported that

the silence/unsilence pattern of one gene strongly predicts whether

breast cancer is likely to recur. Fully 90% of the women in whom

this gene was operating at normal volume were metastasis-free 10

years after treatment, compared with 65% in whom the gene was

silenced. Presumably, the gene is involved in blocking metastasis,

so silencing it spells trouble.

" Epigenetic changes are more clearly associated with the progression

of tumors than mutations are, " says Dr. Feinberg. " Epigenetics may

be as important in certain conditions as the DNA sequence is in

other cases. "

One of the oddest discoveries in epigenetics is that genes inherited

from mom and dad are not equal. Normally, the IGF2 gene you get from

dad is active, but the copy from mom is silenced. In about 10% of

people, however, the " be quiet " tag has been lost. The unsilenced

IGF2 gene is associated with colorectal cancer, Dr. Feinberg and

colleagues reported last year. Epigenomics AG is trying to turn the

discovery into a simple blood test for colorectal cancer risk.

With age, silencers on genes seem to melt away, which might help

explain why cancers and other diseases become more common the older

you get. When one of the two parental genes for a protein called

homocysteine is not properly silenced, the body produces a double

dose of it; high levels are associated with heart disease and stroke.

It is too soon to infer dietary advice from all this, but some

scientists suspect that diets too low in methyl, the molecule that

usually silences genes, may spell trouble. Sources of methyl include

folate (from liver, lentils and fortified cereals) and vitamin B-12

(in meat and fish).

Last fall, European scientists launched a " human epigenome project. "

It will scan DNA for " silence " tags and link them to disease. " The

human epigenome needs to be mapped if we are ever going to

thoroughly understand the causes of cancer and other complex

diseases, which we can't explain by mutations in the DNA sequence, "

says Randy Jirtle of Duke University, Durham, N.C.

Let the race for this second genetic code begin.

URL for this article:

http://online.wsj.com/article/0,,SB109052785446471388,00.html

Hyperlinks in this Article:

(1) http://online.wsj.com/article/0,,SB108992270861765039,00.html

(2) http://online.wsj.com/article/0,,SB108992270861765039,00.html

(3) mailto:sciencejournal@...