For 50 years, thousands of labs around the world have
studied cells' critical internal communications, and
scientists had assumed the speakers were known. But now, in
the Dec. 17 issue of Science, Johns Hopkins
researchers report finding not just a new participant but a
brand new conversation that has implications for treating
disease and understanding biology.
Much of cells' internal communication revolves around
two very important words — "stop" and "go" —
elicited when a small bit, called phosphate, is added onto
proteins. This addition turns protein activities up or down
and fine-tunes cells' responses to what's happening outside
their borders. This communication can go awry in diseases,
including cancer, and be corrected by various drugs.
The source of these phosphate bits has been known
— a molecule called adenosine triphosphate, or ATP.
But in their new report, the Johns Hopkins scientists
describe a brand new source of phosphate that seems to work
with as many proteins as targeted by ATP but in a
completely different way.
"There are already drugs that affect particular roles
of ATP to treat cancer and other conditions, so we envision
[that] drugs that increase or decrease specific activities
of this new source of phosphate could be important in
neurologic and psychiatric illnesses, and perhaps in cancer
as well," said Solomon Snyder, professor and director of
Neuroscience, one of the departments in Johns Hopkins'
Institute for Basic Biomedical Sciences.
"Nobody in a million years would have thought there
was another way for cells to add phosphate groups to
proteins other than using ATP," he added. "Addition of
phosphates to proteins — phosphorylation — is
the most fundamental signaling mechanism in all life, and
the new source of phosphate represents a very different
kind of process than the one we've known about. It
represents a totally new form of cellular
communication."
Unlike ATP, the new phosphate source, known as
inositol pyrophosphate, or IP7, modifies proteins without
any help, just binding directly to the protein and leaving
behind one of its phosphates, the researchers report. Their
early evidence also suggests IP7 might be most important in
regulating the release of chemicals in the brain and in
controlling the cellular machinery that builds proteins.
While IP7's newly found role is likely to surprise
many, Snyder has been expecting it. In the early 1990s, he
noticed the first reports that IP7 and a related molecule
called IP8 existed, interesting to him because for 15 years
he'd been studying related inositol (pronounced
in-AH-si-tahl) phosphates.
But unlike the molecules he'd been working with, which
look like bracelets with three to six phosphate "charms,"
IP7 and IP8 had too many phosphates to fit on the bracelet.
Instead, the seventh and eighth phosphates would have to be
linked to another phosphate "charm" rather than to the
"bracelet" itself.
"ATP has a similar phosphate-phosphate connection, so
I speculated that IP7 and IP8 might also be able to give up
that extra phosphate," said Snyder, who is also a professor
of pharmacology and molecular sciences and of psychiatry.
"Proving it turned out to be very difficult
technically."
First, it took several years for then graduate student
Susan Voglmaier to isolate an enzyme that builds IP7 (from
IP6), an advance the team published in 1996. It took a few
more years for postdoctoral fellow Adolfo Saiardi to clone
the three enzymes that make IP7 and to use them to make IP7
in which the extra phosphate was radioactive.
After finally making a sufficient quantity of
radioactive IP7, Saiardi and postdoctoral fellow Rashna
Bhandari mixed it with a "puree" of mouse brain or kidney,
which produced hundreds of radioactive proteins. After
numerous experiments to rule out other possibilities, the
researchers could finally conclude that IP7 gives away its
extra phosphate to proteins.
"We think IP7 phosphorylation of proteins is as
universal as ATP phosphorylation," said Snyder, whose lab
is continuing to study IP7's protein targets and where on
proteins its phosphates are added. "The enzymes that build
IP7 are most prevalent in the brain, but they are found
everywhere. What we've learned so far is just the tip of
the iceberg."
Already, Bhandari has discovered that two of the
proteins most heavily phosphorylated by IP7 are involved in
the ribosome, the cellular machine that reads genetic
instructions and constructs proteins. IP7 also controls the
cellular "mail room" — the preparation and release of
tiny packages that contain messengers, such as
neurotransmitters in the brain that create movement, memory
and mania.
The researchers also have shown in work described in
the Journal of Biological Chemistry, online now, that
proper activity of one of the enzymes that makes IP7 is
critical in cell death — because of IP7's role in
modifying proteins.
"So drugs that activate the enzyme and stimulate
production of IP7 would increase cell death, which is what
one wants to do in cancer treatment," Snyder said. "Drugs
inhibiting the enzyme would prevent cell death, the goal in
treating stroke and neurodegenerative diseases."
The first proof of how ATP works was the impetus for
the 1992 Nobel Prize in physiology or medicine. During the
mid-1950s, the awardees, Americans Edmond Fischer and Edwin
Krebs, isolated the first example of an enzyme that takes
ATP's phosphate and gives it to a protein and showed that
the phosphate changes the protein's function.
Since then, scientists have found and studied
thousands of other ATP-controlled proteins. While IP7 also
controls protein activity, its role in cellular
communication — coordinating internal activities to
respond to external events — is likely quite distinct
from ATP's, given the differences Snyder and his colleagues
have already observed. IP7 may even add its phosphate to
phosphates already on proteins, which, if confirmed, is
completely unheard of, Snyder said.
Authors are Saiardi, Bhandari, Snyder, Adam Resnick
and Adele Snowman, all of Johns Hopkins. Saiardi is now at
University College London. The research was funded by the
National Institute of Mental Health and the National
Institute on Drug Abuse.