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Molecular Genetic Approaches to Neurodegenerative Disease
Summary: Nancy Bonini is interested in mechanisms of human neurodegenerative diseases, including Huntington's, Parkinson's, ALS, and Alzheimer's diseases. She is using the powerful genetics of Drosophila to create models for human neurodegeneration in the fly, in order to provide new insight into disease mechanisms and uncover novel means of mitigating neuronal loss.
Our laboratory is applying the genetics of Drosophila melanogaster—the simple fruit fly—to the problem of human neural degeneration. Many human neurodegenerative diseases are poorly understood and untreatable, including Huntington's, ALS (amyotrophic lateral sclerosis), Parkinson's, and Alzheimer's diseases. Overall, genes in Drosophila are highly conserved to humans; thus, genes found to be critical to maintenance of the brain in flies are likely to have conserved homologs in humans. This conservation extends not only to protein sequence but also to protein and gene pathway function.
We introduce mutant human disease genes into Drosophila to re-create the respective human disease. Then, using molecular and genetic manipulations, we define mechanisms of degeneration and uncover genes and other approaches that modulate disease manifestation and progression. These studies provide the foundation to determine whether similar mechanisms function in vertebrates, with the goal to provide a basis for new approaches and techniques to interfere with disease progression in humans.
The Fly as a Model for Human Neurodegenerative Disease
We initiated these studies by introducing into Drosophila the human disease gene for spinocerebellar ataxia type 3, also called Machado-Joseph disease (SCA3/MJD). SCA3, one of a class of human neurodegenerative diseases called polyglutamine repeat diseases, is among the most prevalent dominantly inherited ataxia worldwide. Polyglutamine diseases include several additional spinocerebellar ataxias, as well as Huntington's disease. In SCA3 and other similar diseases, a CAG repeat becomes expanded within the open reading frame of the gene. This expansion leads to incorporation of an abnormally long polyglutamine run within the disease protein. A normal glutamine repeat of about 10–30 becomes expanded to 60–80 in SCA3. The expanded polyglutamine domain confers dominant toxicity to the protein, and this leads to neural dysfunction and cell loss.
Initially, we expressed in flies a normal human ataxin-3 protein (the protein encoded by the SCA3 gene) and a human disease form with an expanded polyglutamine repeat. Expression of the normal protein has no effect. Expression of the expanded polyglutamine protein causes an effect reminiscent of human disease: late-onset, progressive neural degeneration. Expanded polyglutamine proteins in flies also form intracellular inclusions—pathological protein aggregates—characteristic of the human polyglutamine diseases. These inclusions are similar to the amyloid fibrillar plaques of Alzheimer's disease, suggesting they may be akin to the aggregates of Alzheimer's and other neurodegenerative diseases. In the latter cases, the aggregated protein is cytoplasmic or extracellular, although mechanisms of protein accumulation, and subsequent neural degeneration, may be similar.
We have extended our studies to mechanisms of other human diseases, including ALS, a motor neuron degenerative disease, and Parkinson's disease, a movement disorder caused by loss of dopaminergic neurons. Rare familial forms of Parkinson's disease are associated with mutations in α-synuclein, a small protein found in neurons that accumulates in the Lewy body aggregates that characterize the human disease. Directed expression of α-synuclein in Drosophila induces adult-onset compromise of dopaminergic neurons, as in the human disorder. Another feature of Parkinson's disease is the potential impact of environmental toxins like the pesticide paraquat to cause disease. Drosophila models for parkinsonism show enhanced susceptibility to such toxins, such that the fly can be used to define the molecular basis of environmental contributions to disease. For ALS, we have revealed a new gene that is a risk factor for ALS, ataxin-2. Ataxin-2 is actually a polyglutamine disease protein, but together with our collaborators, we have shown a critical role for ataxin-2 in ALS. We are now performing genetic screens to reveal new pathways and mechanisms of disease pathogenesis. These studies emphasize the relevance of using fly genetics to approach the complexities of human disease.
Mechanisms of Degeneration and Disease Prevention
One of our goals is to reveal new methods by which to delay or prevent human neurodegenerative disease. One approach involves testing highly suspect modifier proteins, including the molecular chaperones—a set of players that modulate protein solubility. We initially used the fly to address whether altering molecular chaperone levels can affect polyglutamine-induced neurodegeneration. These studies showed remarkable rescue of degeneration by heat-shock protein 70 (Hsp70). Further, these studies revealed that, surprisingly, compromising molecular chaperone pathways may normally contribute to disease.
We extended these findings to Parkinson's disease models to show that up-regulation of molecular chaperone activity—by genes or drugs—protects against deleterious effects of α-synuclein on dopaminergic neurons. These studies also showed that compromised chaperone activity normally contributes to pathology in Parkinson's and other human neurodegenerative diseases associated with abnormal α-synuclein accumulations. Most significantly, data from others have subsequently indicated that polymorphisms that appear to compromise chaperone activity are a risk factor for Parkinson's disease in humans. These studies illustrate that model organisms such as the fly can provide important insight into human disease.
An additional power of the fly rests in the ability to perform large-scale genetic screens to define new and unsuspected mechanisms that contribute to disease. This approach does not require prior knowledge of the precise manner by which the mutation causes disease. Genetic screens can reveal a variety of players, with some screens based on loss-of-function interactions and others using gain-of-function approaches. By applying multiple experimental techniques to the problem, we are defining a range of mechanisms by which neurodegenerative disease occurs, as well as different ways to interfere in the degenerative process. Our studies have confirmed a striking role for protein-misfolding pathways and also highlighted new pathways that have not previously been implicated in neuronal integrity. These additional pathways include the RNA for the disease gene itself: that is, the RNA that encodes the mutant disease protein may contribute toxicity in polyglutamine degeneration. This indicates that there may be both RNA- and protein-level pathogenicities, underscoring an additional layer of complication in mechanism. Moreover, this raises a question regarding treatment: Should one target the protein or the RNA?
To define unifying principles of neurodegenerative disease, we continue to develop and characterize new models. How universal might some mechanisms of disease be? What are overlaps in ways to mitigate the disorders? Are the clinically distinct human diseases mechanistically distinct, or could a treatment for one be a treatment for many? With fly models, we can use the uniquely powerful genetics of Drosophila to uncover new insights into understanding and treatment of diseases.
Acute Neural Injury and Overlaps with Degenerative Situations
We are currently developing an additional line of research into acute neural injury. Damage to the nervous system is typically devastating, often leading to paralysis or extreme functional limitations. We hypothesize that there may be important mechanistic links between acute damage by injury and long-term neurological damage in degenerative conditions. Thus we are developing models for adult-stage acute neural damage, and are following the loss and recovery of neurons with time. Our goals are to first outline the biological characteristics and features of the processes, and then perform genetic screens to define mechanism. Longer term, we will address overlap with molecular pathways of degenerative disease, as well as with normal neurological decline with age. Perhaps a simple system like the fly can highlight important genetic interplay between these different processes. This would provide the foundation for common means of intervention to improve recovery upon neurological damage, to improve function in degenerative disease, and to lessen normal neurological decline with age.
Aspects of this research have received support from the David and Lucile Packard Foundation, the National Institute on Aging, and the National Institute of Neurological Disorders and Stroke.
As of November 22, 2010
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