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9-Misc: Scientific study challenges mantra of molecular biology

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TITLE:  Researchers in Japan, at UC San Diego Discover Novel Role for
SOURCE: AScribe Newswire, USA
DATE:   Apr 30, 2003

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Researchers in Japan, at UC San Diego Discover Novel Role for Pseudogenes

LA JOLLA, Calif., April 30 (AScribe Newswire) -- The mantra of molecular
biology - DNA makes RNA, which makes protein - has pretty much ignored
pseudogenes. Considered defective copies of DNA segments, the 20,000
pseudogenes in the human genome are thought to be non-participants in the
protein-production assembly line.

Now, scientists in Japan and at the University of California, San Diego
(UCSD) School of Medicine have discovered a novel regulatory role for one
pseudogene, showing that it stabilizes a similar protein-coding gene on
another chromosome. When the pseudogene was disabled, protein-production
was compromised, with resulting abnormal kidneys and bones in laboratory
mice. When a functioning pseudogene was re-introduced into mouse embryos,
the mice developed normally.

Published in the May 1, 2003 issue of the journal Nature, the study was
led by Shinji Hirotsune, M.D., Ph.D., Division of Neuro-Science, Research
Center for Genomic Medicine, Saitama Medical School, Japan. Hirotsune
collaborated with Anthony Wynshaw-Boris, M.D., Ph.D., UCSD associate
professor of pediatrics and medicine, in whose lab he first starting
exploring pseudogene function several years ago.

"These findings have implications for treating human disorders," said
Hirotsune. "The mice get disease if the pseudogene is interrupted, so
theoretically it's possible that a malfunctioning pseudogene may cause
human disease, as well."

The team's discovery happened by chance. The researchers were making
transgenic mice for a totally different experiment. The process included
injection of DNA into fertilized eggs, causing the DNA to randomly
integrate into the mouse genome.

"You hope that a transgenic mouse will manifest direct effects from the
gene that you inject, so you can learn something about the function of
that gene," said Wynshaw-Boris. "However, because the inserted gene
randomly integrates itself, sometimes it fortuitously interrupts another
gene that's in the genome and produces mice with characteristics you
didn't expect. In this case, we noticed that one group of mice developed
unusual and severe disease characteristics."

Almost all the mice in that group died. The few that survived had severe
kidney and bone problems, which they passed on to their offspring. The
researchers decided to pursue these unexpected findings.

"In this day and age, it's relatively straightforward to clone the sites
where transgenes are integrated," Wynshaw-Boris continued. "In cloning
this site, we found three genes near the location where the injected gene
was integrated."

With further laboratory tests, the scientists ruled out two of the genes,
and determined that the third gene - a pseudogene called makorin1-p1 -
was responsible for abnormalities in the mice.

"Since a pseudogene does not have the ability to make a protein, we
wanted to know how this pseudogene could cause the disease states in the
mice," Hirotsune said.

The researchers noted that the pseudogene, makorin1-p1, is a fragmented
gene similar to a full-fledged protein-coding gene called makorin1, which
is located on a different chromosome.

In examinations of normal mouse kidneys, the protein-coding makorin1 was
shown to be strongly expressed and highly visible. In mice with the
disabled makorin1-p1 pseudogene, the expression of the makorin1 gene
appeared weak and irregular in the diseased kidney tissue. Additional lab
tests and experiments in mice determined that makorin1-p1 played a key
role in regulating the stability of makorin1.

"We further confirmed that this regulation was occurring by correcting
the defect in the line of the initial transgenic mice," Wynshaw-Boris
said. "If the mouse embryo had a defective copy of the pseudogene, we
replaced it with a normal copy, which produced a healthy mouse."

In continuing studies, the researchers hope to show the pseudogene-gene
interaction is a general mechanism taking place in many cellular interactions.

Additional authors on the paper in Nature were Noriyuki Yoshida, Research
Center for Genomic Medicine, Saitama Medical School, Japan; Amy Chen and
Lisa Garrett, Genetic Disease Research Branch, National Human Genome
Research Institute, National Institutes of Health; Fumihiro Sugiyama,
Satoru Takahashi and Ken-ichi Yagami, Institute of Basic Medical Sciences
and Laboratory Animal Resource Center, University of Tsukuba, Japan; and
Atsushi Yoshiki, Experimental Animal Division, Department of Biological
Systems, BioResource Center, RIKIN Tsukuba Institute, Japan.

Funding for the study came from PREST, Japan Science and Technology
Corporation, and PROBRAIN.

- Located in the nucleus, double-stranded DNA carries the genetic
information of a cell. Its thousands of genes act like instruction
manuals to build proteins, which perform important tasks for cell
functions or serve as the body's building blocks. RNA is an intermediate
between DNA and protein. Parts of the DNA are "transcribed" into RNA, a
single-stranded molecule, which contains the information needed to make
the protein. This protein-coding RNA moves out of the nucleus, into the
cytoplasm of the cell, and directs the assembly of amino acids that fold
into a completed protein.