Aggregation State Of Huntingtin Regulates Its Interaction With Lipid Membranes

Huntingtin disease (HD) is a neurodegenerative disease caused by an expansion of a polyglutamine (polyQ) tract within the huntingtin (htt) protein. There is a critical threshold for the polyQ length, which once exceeded, causes HD. PolyQ expansion promotes the self-assembly of htt into a variety of aggregate species ranging from small oligomers to large fibrils that eventually incorporate into large inclusion bodies. The first 17 amino acids of htt (Nt17), which directly precede the polyQ tract, play a role in initiating aggregation by facilitating the formation of oligomeric intermediates that lead to fibril formation. Nt17 also functions as a lipid membrane binding domain. To characterize the ability of different aggregate forms of htt to bind lipid membranes, htt was pre-aggregated, and the ability of different aggregate states to bind lipid membranes was determined by a polydiacetylene lipid binding assay. The aggregation state of htt was verified by atomic force microscopy (AFM). While monomeric htt readily bound lipid vesicles, the interaction was enhanced by the presence of htt oligomers. To determine if the ability of oligomers to continue to aggregate on the lipid surface played a role, htt oligomers were stabilized via a lysine cross-linker, eliminating its ability to further aggregate. Cross-linked oligomers were assessed and characterized by ThT assays, filter traps, AFM, and dynamic light scattering. The ability of the cross-linked oligomers to interact with lipid membranes was then assessed using PDA assays and AFM.