Differential interactome and innate immune response activation of two structurally distinct misfolded protein oligomers

The formation of misfolded protein oligomers during early stages of amyloid aggregation and the activation of the inflammatory response are two characteristic events associated with the onset and progression of a wide range of neurodegenerative diseases. Although it has been established that misfolded oligomers are involved in the neuroinflammatory process, the links between their structural and physicochemical features and their functional effects on the immune response remain largely unknown. To explore such links we have taken advantage of two structurally distinct and well-characterized types of soluble oligomers of the protein HypF-N, denoted type A and B, which have been shown to have different hydrophobicity and cytotoxicity. We analysed the microglial inflammatory response triggered by these oligomers and found that, even though they both stimulate the release of pro-inflammatory cytokines, type B oligomers induced a substantially greater response than type A oligomers. By using a combination of confocal microscopy, protein pull-down assays and high-throughput mass spectrometric-based proteomic analysis, we found that the two types of oligomers interact differentially with microglial cells. In particular, despite the fact that both types of oligomers were found to bind to a common pool of microglial proteins, type B relative to type A oligomers showed enhanced protein binding, a finding that correlated with the observed inflammatory response. In addition, the receptors and signalling molecules in the interactome associated with inflammatory-mediated neurodegeneration revealed a number of previously unidentified proteins likely to be involved in the oligomer-elicited innate immune response.