N-terminal acetylation and replicative age affect proteasome localization and cell fitness during aging.
Bottom Line: By using a microscopy-based genome-wide screen, we identified genetic factors involved in these processes.Both relocalization of the proteasome and PSG formation were affected by two of the three N-acetylation complexes.These N-acetylation complexes also had different effects on the longevity of cells, indicating that each N-acetylation complex has different roles in proteasome location and aging.
Affiliation: Division of Cell Biology, Netherlands Cancer Institute and Netherlands Proteomics Center, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands.Show MeSH
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Mentions: We visualized proteasomes in live yeast cells by tagging the catalytically active β1 subunit (Pre3) of the proteasome with GFP. Efficient and quantitative introduction of the β1–GFP in 20S proteasomes was confirmed by native gel electrophoresis (supplementary material Fig. S1). The GFP-labeled proteasomes had a similar distribution during starvation as reported for non-modified proteasomes previously (Laporte et al., 2008). We observed that cells in starvation show a wide heterogeneity in proteasome localization (Fig. 1A). Based on proteasome localization, we defined four localization phenotypes: (1) cells with proteasome accumulation in the nucleus (Nuclear); (2) cells displaying dots of cytoplasmic proteasome clusters (PSG); (3) cells displaying both PSGs and a nuclear accumulation of proteasomes (Nuclear + PSG); and (4) cells without any of these phenotypes, where proteasomes are approximately equally distributed between the cytoplasmic and nuclear compartments (Equal) (Fig. 1B). In a typical 5-day starvation experiment the majority of the cells are either PSG or Equal, whereas a small portion of the cells is Nuclear or Nuclear + PSG.
Affiliation: Division of Cell Biology, Netherlands Cancer Institute and Netherlands Proteomics Center, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands.