Abstracts of the 29th Annual Symposium of The Protein Society

Spinal bulbar muscular atrophy (SBMA) is a member of the polyglutamine (polyQ) expansion diseases, like Huntington disease, and it is caused by a genetic expansion of the polyCAG tract in exon 1 of androgen receptor (AR) that codes for the polyQ region. SBMA is a late onset disease, which involves a progressive degeneration of the motor neurons and consequent muscular atrophy. There is still no treatment available for this disease. AR is a nuclear receptor that responds to testosterone and that regulates the expression of the masculine phenotype. It is composed of an intrinsically disordered Nterminal domain (NTD) that bears the polyQ tract, a DNA binding domain and a ligand binding domain. Aggregates of AR protein with an extended polyQ are observed in the motor neurons of SBMA patients. In vitro studies showed that aggregation of Androgen Receptor takes place only in presence of testosterone1 and that the cleavage of the protein by caspase 3 is a crucial event for cytotoxicity. However, there is no clear knowledge of the mechanism of aggregation, for this protein. An increasing body of evidence supports the hypothesis that the aggregation of these proteins is controlled by regions flanking the polyQ tract, by regulating the rate of aggregation depending on their secondary structure. We have applied nuclear magnetic resonance (NMR) and circular dichroism for generating information on the secondary structure of the N-terminal cleavage product of AR by caspase 3 and we have studied its aggregation with a set of biophysical methods, like dynamic light scattering, an HPLC sedimentation assay and transmission electron microscopy. We have found that the polyQ tract of AR presents a high degree of helicity. We attribute this conformation to the N-terminal flanking region, characterized by high helicity and we have tested this hypothesis by performing mutations. We have also observed that the rate of the first step of oligomerization is not dependent on the number of glutamine repeats, but instead is due to self interactions of a region N-terminal to and far from the polyQ. Its progression to fibril is dependent to the number of glutamines in the tract. We have therefore identified two steps in the aggregation process of AR, where a motif far from the polyQ at its N-terminal drives the early oligomerization, followed by the interaction of the polyQ chains that stabilize it and determine the progression to fibrils. These findings shed a light for possible interventions on the AR oligomerization process, thus suggesting a different strategy to study the onset of the disease in SBMA patients.