Role of the coiled-coil structure of Huntingtin in the intercellular propagation of PolyQ aggregates

Huntington Disease (HD) originates from the expansion of a polyglutamine (PolyQ) tract in the Huntingtin protein (Htt), that can assume a Coiled coil fold (Cc). We previously found that Cc structures mediate the aggregation and toxicity of Htt and hypothesize they might be implicated in other aspects of Htt dysfunction. There is evidence that expanded and aggregated Htt can be released by sick cells and taken up by healthy ones. With a prion-like mechanism, aggregated Htt can then seed the misfolding of normal Htt spreading the disorder.We hypothesize that Cc also mediate the intercellular propagation of Htt aggregates. To test this hypothesis, we performed in vitro experiments using Htt constructs with different probability to acquire a Cc fold. We assessed their ability to be released from transfected cells and be internalized by recipient cells in condition of cell contact or contact-less, by using a combination of techniques (western blot, FACS and confocal imaging).We found that impairment of the Cc structure significantly reduced Htt release, while it did not affect the uptake from cells. Moreover, Htt is released in transfected cells medium by exosomes. When exosome release is reduced by treatment with the sphingomyelinase inhibitor GW4869, we found a significant reduction in the amount of Htt present in the medium.Together these results indicate that Htt is released in exosomes through a Cc mediated mechanism and this event can be modulated pharmacologically.