Detecting Protein Aggregation on Cells Surface: Concanavalin A Oligomers Formation

Abstract

A number of neurodegenerative diseases involve protein aggregation and amyloid formation. Recently evidence has emerged indicating small-transient prefibrillar oligomers as the primary pathogenic agents. Noteworthy, strict analogies exist between the behaviour of cells in culture treated with misfolded non-pathogenic proteins and in pathologic conditions, this instance together with the observation that the oligomers and fibrils are characterised by common structural features suggest that common mechanisms for cytotoxicity could exists and have to be perused in common interactions involved in aggregation.We here report an experimental study on ConcanavalinA (ConA) aggregation and its effects on cells. In vitro, close to physiological temperature, this protein readily forms fibrils involving secondary structure changes leading to β-aggregate structures.The effect of a ConA on cell cultures was tested and the formation of protein aggregates in these samples was studied by confocal fluorescence microscopy. We used the NB simultaneusly, the morphology of the cell changes indicating the progressive cell compaction and death. Cell surface probably provides nucleation sites for aggregation where high local concentration and macromolecular crowding favor aggregation. The formation of small aggregates may stimulate non-specific cellular response as a result of the exposure of reactive regions of protein structure and of the progressive formation of cross-β structures. Moreover, these oligomers could interact with the cell membrane damaging its structural organization and destroying its selective ion permeability. These results show the suitability of using ConA as a model protein and the N&B analysis as a powerful tool to measure aggregation in cells and to give new insights the relation between protein aggregation and disease. P41-RRO3155(EG).