Reviewer #2 (Public Review): Impairment of lipid homoeostasis causes accumulation of protein aggregates in the lysosome

Abstract

Protein aggregation increases during aging and is a pathological hallmark of many age-related diseases. Protein homeostasis (proteostasis) depends on a core network of factors directly influencing protein production, folding, trafficking, and degradation. Cellular proteostasis also depends on the overall composition of the proteome and numerous environmental variables. Modulating this cellular proteostasis state can influence the stability of multiple endogenous proteins, yet the factors contributing to this state remain incompletely characterized. Here, we perform genome-wide CRISPRi screens to elucidate the modulators of proteostasis state in mammalian cells using a fluorescent dye to monitor endogenous protein aggregation. These screens recovered components of the known proteostasis network, and uncovered a link between protein and lipid homeostasis. We subsequently showed that increased lipid uptake and/or disrupted lipid metabolism led to increased lysosomal protein aggregation and, concomitantly, accumulation of sphingolipids and cholesterol esters. Surprisingly, lysosomal proteostasis impairment by lipid dysregulation is independent of lipid peroxidation or changes in lysosomal stability, nor is it caused by effects on many other aspects of lysosomal or proteasomal function. These results suggest that lipid dysregulation may have primary effects on the stability of endogenous proteins, potentially through direct biophysical mechanisms.