This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. 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 b-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 N&B analysis method to monitor ConA aggregation in live cells. The N&B analysis shows a rapid and progressive formation of ConA oligomers on cell membrane, even at very low protein concentration;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-b structures.
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