Working with mouse, fly and human cells and tissue, researchers report new evidence that disruptions in the movement of cellular materials in and out of a cell’s control center — the nucleus — appear to be a direct cause of brain cell death in Huntington’s disease, an inherited adult neurodegenerative disorder. Moreover, they suggest, laboratory experiments with drugs designed to clear up these cellular “traffic jams” restored normal transport in and out of the nucleus and saved the cells.
In the study, published in Neuron, the researchers also conclude that potential treatments targeting the transport disruptions they identified in Huntington’s disease neurons may also work for other neurodegenerative diseases, such as ALS and forms of dementia.
In an earlier study, the team found out how a mutation in a gene — implicated in 40 percent of inherited ALS cases and 25 percent of inherited frontotemporal dementia cases — gums up transport in and out of the nucleus in neurons, ultimately shutting the cell down and leading to its death. The mutant gene makes RNA molecules that stick to a transport protein, RanGAP1. RanGAP1 in turn helps move molecules through nuclear pores that serve as passageways in the nucleus, letting proteins and genetic material flow in and out of it. They learned that this same mutation is also the most common cause of another disorder in which patients have Huntington’s-like symptoms without having the causative Huntington’s disease mutation. Additionally, they realized that other researchers previously showed that mutations in the nuclear pore protein NUP62 caused Huntington’s disease-like pathology. Because of such clues from others’ research, the scientists took on the task of investigating whether problems with nuclear transport and the nuclear pores also happened in neurons with Huntington’s disease.
The researchers used two mouse models of Huntington’s disease: one with a human version of the mutant Huntingtin protein and another with an aggressive form of the disease that contains only the first portion of the mouse Huntingtin protein. By using antibodies with glowing markers that bind to specific proteins and viewing the neurons under the microscope, they saw that the mutant Huntingtin protein clumped up in the same location of the cell as abnormal clumps of RanGAP1, the nuclear transport protein. It also clumped up in the same location as abnormal clumps of nuclear pore proteins NUP88 and NUP62.
They also observed this same clumping of Huntingtin protein with RanGAP1 and nuclear pore proteins to the wrong place in the cell in brain tissue and cultured brain cells derived from deceased patients with Huntington’s disease.
To further explore nuclear transport’s role in Huntington’s disease, the team took lab-grown mouse neurons and used chemical switches to a) turn on both an additional healthy copy of the RanGAP1 gene and a mutant version of Huntingtin; b) just turn on the mutant Huntingtin; or c) just turn on a healthy version of Huntingtin.
They then measured cell death and found that neurons with the healthy version of Huntingtin had about 17 percent of the neurons die off. Neurons with only the mutant version of Huntingtin were more likely to die, with about 33 percent dying off, but in neurons with both the mutant Huntingtin and the RanGAP1, only 24 percent of the neurons died off. The researchers think that some of the extra healthy RanGAP1 they introduced into diseased cells wasn’t bound up to the mutant Huntingtin and resumed normal nuclear transport.
Next, they looked at cell death in cultured neurons with a healthy or a mutant form of Huntingtin, or with a mutant form of Huntingtin that was treated with small amounts of an experimental drug called KPT-350, one that prevents a nuclear export protein, Exportin-1, from shuttling proteins and RNA out of the nucleus. Neurons with the healthy version of Huntingtin had about 18 percent die off, and neurons with the mutant version of Huntingtin had about 38 percent die off. Those treated with the nuclear export blocking drug had improved survival, with only about 22 percent of the cells die off. Blocking nuclear export seemed to prevent cells from dying and counteracted the defects in neurons with mutant Huntingtin.
According to the researchers, there is an average of 2000 nuclear pores per cell and each individual nuclear pore consists of multiple copies of more than 30 different proteins that each serve different functions. It may be that nuclear pores on neurons and other types of brain cells like glia are constructed of different combinations of these proteins, some of which may be more or less critical in various neurodegenerative diseases.
The team of researchers is currently working on answering this question using a new mouse model that will allow them to isolate these nuclear pore proteins from different cell types in the mouse brain to identify whether these nuclear pore components are in fact different based on brain cell types and brain locations.
Paper: “Mutant Huntingtin Disrupts the Nuclear Pore Complex”
Reprinted from materials provided by Johns Hopkins Medicine.