July 6, 1999 DURHAM, N.C. -- Researchers have reported discovering the first elements of what is apparently a molecular signaling pathway important for regulating how genetic information leaves the nucleus to begin its working life as a blueprint for the cell.
According to the scientists, their finding represents an intriguing new role for a signaling molecule, derived from inositol, in the cell.
In an article in the July 2 Science, Duke University Medical Center pharmacologist John York and his colleagues reported studies in yeast that revealed the presence of enzymes called kinases ? enzymes that add phosphates to inositol to make them into signals that trigger the export of messenger RNA from the cell nucleus.
Messenger RNA (mRNA) is the molecule that, when copied from DNA genes in the nucleus, carries genetic information into the cell where it is used to build proteins.
Co-authors on the article are Audrey Odom, also of Duke, and Robert Murphy, Eric Ives and Susan Wente of Washington University School of Medicine. York is an assistant professor of pharmacology and cancer biology, and of biochemistry. Odom is a graduate student in the Medical Scientist Training Program. The research was sponsored by the Burroughs Wellcome Fund, the National Institutes of Health and the American Cancer Society.
While York emphasizes that the discovery is very basic in nature, and was made using yeast, such fundamental knowledge invariably contributes to understanding and treatment of human disease, he said. Particularly important, he emphasized, is that inositol signaling is critical to the machinery of living cells, and many of its components are conserved all the way from yeast to humans.
Inositols had already been suspected of being major signaling molecules in the cell, because some 20 different inositols had been discovered with varying numbers and arrangements of phosphates attached, like a multitude of keys with slightly different shapes. However, researchers still do not understand where and when the cell uses these molecular keys, and which locks they open.
The most well-known inositol, IP3, has three phosphates attached, and is produced in the cell in response to stimuli, via the action of a key enzyme called phospholipase C. IP3 is known to be critical for triggering the release of calcium, which in turn triggers other cell processes.
However, York and his colleagues to their surprise had found that IP3 apparently has yet unknown extended signaling roles, produced when additional phosphates are added to produce IP4, IP5 and IP6 in the cell. Production of these IPs also depends on phospholipase C activity in cells.
The latest research began when York contacted Wente, after learning through colleague Jeremy Thorner of the University of California-Berkeley that Wente had discovered a link between a phospholipase C and an essential factor GLe-1 that is essential for transporting mRNA out of the nucleus. Of particular interest was that Wente had also discovered that two other genes of unknown function also linked to GLe-1.
York theorized that the other two genes might be the blueprints for two new kinases responsible for adding phosphates to IP3 on the way to producing IP6.
"It was easy to test this hypothesis," said York. "We just introduced radio-labeled inositol into yeast strains mutated to lack the genes for one or the other of the suspected kinases. Depending on the kinase blocked, we expected to see buildup of either IP3 or IP5, with no IP6 in any case." Sure enough, said York, the experiments indicated that the mysterious genes coded for kinases, or at least their regulators, transformed IP3 to IP4, and IP5 to IP6.
According to York, still to be determined is whether IP6 directly or indirectly affects the transport of mRNA from the nucleus. IP6 may merely regulate the "gatekeeper" -- called the Nuclear Pore Complex -- a huge protein machine that controls the passage of mRNA and other substances out of the nucleus. Or, IP6 may directly affect the protein "package" that carries mRNA through the Nuclear Pore Complex.
Especially intriguing, said York, is that inositol signals are not absolutely necessary for mRNA to leave the nucleus, except at high temperatures.
"One idea is that this process is involved only in transport of specific genes that are needed to help cells adapt to stress, such as heating," he said.
Future studies will aim at identifying the protein that is the target of the IP6 signal and how it affects that protein to act as a switch. More broadly, the discovery of the two kinases has helped open up an important new research pathway, emphasized York.
"This discovery really challenges the idea that IP3 was the sole signal coming from this pathway," he said. "In fact, we know now it's not. Now we know that while this pathway begins by affecting calcium, it ends with the export of mRNA from the nucleus as mediated by IP6. We now want to find out what are the signaling roles of each molecules along this pathway, and in fact whether there's a signal after IP6."
Even broader, said York, the research offers even more hints that inositols have extremely complex signaling roles, including specialized roles in different compartments of the cell and at different times.
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