Overview

The Hughes laboratory studies two related aspects of longevity research, neurodegeneration and the basic biology of aging, from the perspectives of biological mechanism and chemical biology. Our primary techniques include yeast genetics, mammalian cell culture, and high-throughput screening for chemicals and biomolecules.

Molecular and Chemical Biology of Neurodegeneration and Aging

Molecular mechanisms of neurodegeneration

Neurodegeneration is a significant cause of agerelated decline in cognitive function and quality of life. Common age-related neurodegenerative conditions include Parkinson’s disease and Alzheimer’s disease. To study the biological mechanisms that underlie neuronal dysfunction and death we have focused on Huntington’s disease (HD), a genetic condition that leads to late-onset death of neurons in the brain. Our approach to HD has been to use proteomic and chemical screening technologies to discover protein targets that modulate the severity of the disease and chemicals that can interact with these targets. A primary cause of HD can be traced to dysfunction in the solubility and molecular interaction properties of the human protein huntingtin. Perturbations in protein solubility have been implicated in a broad range of late-onset human neurological diseases, and these aspects of these disease-related phenomena can be readily modeled in simple laboratory organisms such as the yeast Saccharromyces cerevisiae.

Fig. 1 shows the behavior of normal and disease-causing forms of the human huntingtin protein when expressed in yeast cells. The micrograph in the upper panel shows the diffuse localization of normal human huntingtin when expressed in yeast. The micrograph in the lower panel shows the insoluble disease-causing form of huntingtin (bright green spots). The experimental tractability of yeast cells allows us to rapidly discover other human proteins and chemicals that can influence the solubility and disease-causing activities of this protein.

To further elucidate the role of the huntingtin protein in the pathology of HD, we have performed high-throughput genetic and biochemical screens for huntingtin and other human proteins that can interact with huntingtin. These screens, involving the use of yeast-based protein interaction discovery technologies and mass spectrometry, have allowed us to identify hundreds of human proteins that potentially play key roles in the progression of HD.

Fig. 2 shows a protein interaction network involving huntingtin (center) and human proteins that can bind to it and, in some cases, each other. The functions of these proteins and their patterns of interactions within the network provide clues about the roles played by normal and mutant huntingtin in healthy and diseased neurons. Using a technique called high-throughput siRNA screening in human cell models of HD, we are systematically evaluating the roles of these and thousands of other human proteins in HD. The purpose of these studies is to discover drug targets and genetic modifiers of HD in order to facilitate the development of therapeutic interventions. We hope that these studies will have significant implications for the understanding of other more common lateonset neurological conditions.

Discovering therapeutic compounds that prevent neurodegeneration

Our interest in neurodegeneration also extends to the discovery of compounds that can prevent death in neuronal models of HD. To date we have taken two approaches to this problem. In one study, we have screened a large chemical diversity library for compounds that can prevent the solubility defects of the disease-causing form of human huntingtin. This was accomplished by creating a yeast strain that suffers a growth defect as a result of expressing the mutant form of human huntingtin. This yeast strain was screened against 75,000 different chemicals to define those that could suppress the huntingtinmediated growth defect. The chemicals are now being tested to find those that can suppress toxicity in human neurons.

Since many diseases that include late-onset neurodegeneration are caused by pathogenic interaction between different cellular proteins, we are also developing technologies to find chemicals that can prevent these interactions. We have performed several large screens for compounds that can prevent the interaction of human proteins when expressed together in a yeast cell. These chemicals are also being evaluated and developed as potential therapeutics for the treatment of neurodegenerative disease.

Chemical biology of aging in yeast

Since late-onset neurodegeneration can be thought of as a failure of neurons to survive over long periods of time, we are interested in the possible roles of genes and chemicals that promote cellular longevity in neurologic disease. The genetic control of longevity appears to be highly conserved among the common model systems (i.e., S. cerevisiae, Drosophila, and C. elegans). We are therefore taking advantage of the tractability of yeast to perform high-throughput screens to discover chemicals that can increase longevity.

Fig. 3 shows the average loss of viability in 5,000 individual yeast cultures, each exposed to a different component of a chemical library. Over a period of 20 days, the survival of aging yeast populations decreases by more than 50%. We are interested in discovering chemical compounds that can increase the survival of aging yeast. Such compounds will be tested for their ability to increase longevity in fly and worm models of aging, and for their ability to promote survival in mammalian neurons. Thus, we are using model systems to explore the biological and chemical connections between aging and neurodegeneration, to elucidate the basic mechanisms of aging and cell survival while developing therapeutic interventions for the treatment of age-related disease.