Qualitative and quantitative multiplexed proteomic analysis of complex yeast protein fractions that modulate the assembly of the yeast prion Sup35p.

ABSTRACT

BACKGROUND: The aggregation of the baker's yeast prion Sup35p is at the origin of the transmissible [PSI(+)] trait. We and others have shown that molecular chaperones modulate Sup35p aggregation. However, other protein classes might be involved in [PSI(+)] formation. RESULTS: We designed a functional proteomic study that combines two techniques to identify modulators of Sup35p aggregation and describe the changes associated to [PSI(+)] formation. The first allows measuring the effect of fractionated Saccharomyces cerevisiae cytosolic extracts from [PSI(+)] and [psi(-)] yeast cells on Sup35p assembly. The second is a multiplex qualitative and quantitative comparison of protein composition of active and inactive fractions using a gel-free and label-free LC-MS approach. We identify changes in proteins involved in translation, folding, degradation, oxido-reduction and metabolic processes. CONCLUSION: Our functional proteomic study provides the first inventory list of over 300 proteins that directly or indirectly affect Sup35p aggregation and [PSI(+)] formation. Our results highlight the complexity of the cellular changes accompanying [PSI(+)] formation and pave the way for in vitro studies aimed to document the effect of individual and/or combinations of proteins identified here, susceptible of affecting Sup35p assembly.