Ubiquitinated protein levels remain high during ageing inC. elegans

ABSTRACT A wealth of literature in different model organisms have shown that proteostasis declines with age, which results in the accumulation of aggregated proteins that generally carry polyubiquitin chains. In Koyuncu et al. (Nature, 2021), the authors use a relatively mild homogenization approach to analyze total polyubiquitinated protein levels longitudinally in C. elegans and found that the overall level of polyubiquitinated proteins decreases in wild-type nematodes. Specifically, the substantial presence of polyubiquitinated proteins in early adulthood markedly decreases at day 10 and 15 of adulthood. This finding represents the central basis of their study where they subsequently suggest that age-dependent decrease in protein polyubiquitination in wild-type animals is due to enhanced deubiquitination. Here, we find that the mild homogenization of C. elegans used in Koyuncu et al . fails to properly extract all C. elegans proteins, because it largely omits relatively insoluble proteins in the pellet. When we perform complete homogenization of wild-type nematode proteins using SDS, sonication and heat, we unequivocally observe that polyubiquitinated protein levels do not decrease with age in C. elegans . Overall, the levels of polyubiquitinated proteins remains relatively constant post-reproduction. Therefore, our findings invalidate the main conclusion of Koyuncu et al. that the ubiquitinated proteome is rewired during ageing and demonstrate that C. elegans harbor polyubiquitinated proteins throughout their lifespan.