Profiling the physiological impact of aberrant folded-state protein filamentation in cells

Abstract

The formation of large polymeric structures such as cytoskeletal and enzyme filaments is crucial for normal cellular function. However, such filaments can also form due to mutations that create self-interactions at the surface of symmetric proteins. Often, the proteins forming these structures maintain a folded state and thereby differ from aggregates and amyloids that involve misfolding. We refer to this type of assemblies as agglomerates to mark this difference. While cells have quality control mechanisms to identify, buffer, and eliminate misfolded proteins, it is unclear whether similar mechanisms exist for agglomerates, or whether agglomerates are toxic to cells. Here, we profiled the physiological impact of mutation-induced folded-state protein filamentation in yeast cells. First, we devised a simple strategy to distinguish fluorescently labeled proteins forming agglomerates versus aggregates. We then profiled exogenous protein agglomerates in terms of their recognition by known quality control mechanisms, their effects on specific cellular processes and overall fitness on S. cerevisiae cultures. We found that agglomerates do not colocalize with the proteostasis machinery and do not result in measurable fitness defects. Proteomics profiling of cells expressing the wild type protein, agglomerating or misfolded variants revealed a consistent picture, with only minor, agglomerate-size-dependent changes observed and linked to the cell-wall and plasma-membrane proteins. Overall, our findings indicate that agglomerates form mostly benign structures in cells when compared to aggregates, and thereby offer a promising route for synthetic biology applications. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=146 SRC="FIGDIR/small/612878v1_ufig1.gif" ALT="Figure 1"> View larger version (84K): org.highwire.dtl.DTLVardef@7136a4org.highwire.dtl.DTLVardef@ba62dborg.highwire.dtl.DTLVardef@b02bbaorg.highwire.dtl.DTLVardef@6cbf9_HPS_FORMAT_FIGEXP M_FIG C_FIG