The aim of this proposal is to understand the molecular basis for Huntington disease (HD). The recently-isolated HD gene is predicted to encode a very large protein of over 3,000 amino acids, and the sequence of the encoded protein shows virtually no homology to known proteins. The dominant mutation that apparently causes the neurodegenerative phenotype is a doubling (or more) in the length of a 5'-CAG-3' trinucleotide repeat from about twenty copies in unaffected individuals to forty or more copies in affected people. This CAG-repeat resides near the 5' end of the messenger RNA, and based on its DNA sequence context, is predicted to encode a stretch of glutamine residues seventeen amino acids from the amino terminus of the protein. The goals of this research are to determine the normal role that the HD protein plays in the cell, as well as the reason the expanded number of CAG-repeats causes the disease phenotype. Antibody and expression cloning reagents will be generated to determine in which cells the HD protein is localized, whether the CAG-repeat and the seventeen amino acids predicted to be located amino terminal to it are indeed expressed in the protein, and whether the HD protein exists as a monomer or an oligomeric species in solution. Specific hypotheses, suggested by the potential polyglutamine region, about whether the HD protein is involved in transcription control or is a substrate for a class of enzymes called transglutaminases, will be tested. A genetic screening system in yeast will be used to identify cellular proteins that interact with the HD protein. Finally, experiments will be done to determine whether the primary structure, potential oligomeric structure, any activities, or any protein:protein interactions associated with the HD protein are different in the normal versus the mutant versions of the protein. The basic information about the HD protein gained from these studies will provide insights into the normal and pathological features of neurodegeneration and the mechanisms by which an expanded number of trinucleotide repeats, which has now been found in five neurological disorders, cause the symptoms of disease. Furthermore, such knowledge is likely to be an important prerequisite for developing treatments for such diseases.
