Published in the September 28 online edition of Nature Genetics, and in the journal's October issue, the study identified a novel change in mice in a gene called mRNA nuclear export factor 1 (Nxf1). This gene normally acts as part of the cell's machinery for ensuring that only properly functioning genes are expressed, or turned on. Genes that have been disrupted by insertions of retroviruses often fail to pass this quality checkpoint.
The UCSD team demonstrated that the novel form of Nxf1 restored function to mutated genes by increasing the amount of normal product made by the mutated gene, and alleviated abnormalities in two very different mouse mutations.
"The properties of this gene could be used to engineer a system for controlling some mutations caused by retroviruses," said the study's senior author, Bruce Hamilton, Ph.D., UCSD assistant professor of medicine. "This could be particularly useful for creating mouse models of human disorders where loss of gene expression is a target of therapeutic efforts, but the dose-responsiveness required for functional recovery is unknown, such as in Fragile X syndrome and certain cancers."
Nxf1 controls expression of genes at the level of exporting messenger RNA (mRNA) from the nucleus. mRNA is an intermediary step in the production of proteins. DNA in the nucleus is transcribed into RNA, which is processed and exported to the cytoplasm. This mRNA is then translated to make proteins. However, when a retrovirus invades a cell, it sometimes inserts a copy of itself into a gene of the host cell. As a result, the host gene may be poorly expressed and depending upon the gene and how it is affected, this can cause disease.
Hamilton noted that was what surprised his team was that a protein required for a late step in gene expression – export of mRNA from the nucleus – appears to influence earlier steps in gene expression, processing of the pre-mRNA. Although previous work by others has identified genes that modify mutations by increasing the amount of RNA being initiated, or changing the ability to express a mutated gene, the new findings are the first to identify suppression at a later stage, when the mutated gene has been activated, but before it can do damage.
In their experiments, the UCSD team began by investigating a gene called Modifier-of-vibrator-1 (Mvb1), which reduces the damaging characteristics of a neurological gene mutation called vibrator. The vibrator mutation is caused by insertion of a mouse retrovirus called IAP.
The researchers studied vibrator mice, which have severe tremors, progressive degeneration of the brain stem and spinal cord, and die early. However, the UCSD team found that vibrator mice carrying the mvb1 genes showed reduced tremor severity and survived to adulthood. The team showed the Mvb1 did this by raising the amount of normal RNA made from the mutant vibrator gene.
In further experiments, the researchers showed that Mbv1 also modifies an Eya1 mutation that is a mouse model of human branchiootorenal syndrome, which is also caused by an IAP retrovirus. Eya1 is a gene required for development of the inner ear and other structures. Mutations are characterized by hearing impairment, ear malformation, and in some cases, by kidney malfunction. Again, Mbv1 modified several of the Eya1 characteristics by increasing the expression of normal RNA made from the mutant gene.
Next, the team used sophisticated laboratory techniques called positional complementation to identify Mvb1, and to determine that it is part of the mRNA nuclear export factor Nxfl. The team further showed that the variation in Nxf1 that allows it to suppress retrovirus mutations is present in a large population of wild mice. Their findings also suggested that this variant form of the gene has evolved, under natural selection, to increase its frequency in the population.
In follow-up work, the Hamilton team is clarifying the biochemical mechanism and testing the range of mutations that can be suppressed. And, they hope to adapt Nxfl for broader use as a research tool.
This work was supported by grants from the National Institutes of Health and the Medical Research Service of the U.S. Department of Veterans Affairs.
The paper's first author was Jennifer A. Floyd, B.A., UCSD Biomedical Sciences Graduate Program. Additional authors were David A. Gold, B.A., UCSD Biomedical Sciences Graduate Program; Dorothy Concepcion, B.A., Tiffany H. Poon, B.A., Erica J. Ward, M.S., UCSD Department of Medicine; Xiaobo Wang, M.D., Elizabeth Keithley, Ph.D., UCSD Division of Otolaryngology; Dan Chen, B.A., UCSD Molecular Pathology Graduate Program; Rick A. Friedman M.D., Ph.D. Steven B. Chinn, B.A., House Ear Institute, Los Angeles, CA; Hon-Tsen Yu, Ph.D., Department of Zoology, National Taiwan University; Kazuo Moriwaki, Ph.D., RIKEN Bioresource Center, Japan; and Toshihiko Shiroishi, Ph.D., Mammalian Genetics Laboratory, National Institute of Genetics, Japan.
Journal
Nature Genetics