Yeast derlin Dfm1 employs a chaperone-like function to resolve misfolded membrane protein stress

Abstract

Protein aggregates are a common feature of diseased and aged cells. Membrane proteins comprise a quarter of the proteome, and yet, it is not well understood how aggregation of membrane proteins is regulated and what effects these aggregates can have on cellular health. We have determined in yeast that the derlin Dfm1 has a chaperone-like activity that influences misfolded membrane protein aggregation. We establish that this function of Dfm1 does not require recruitment of the ATPase Cdc48 and it is distinct from Dfm1’s previously identified function in dislocating misfolded membrane proteins from the endoplasmic reticulum (ER) to the cytosol for degradation. Additionally, we assess the cellular impacts of misfolded membrane proteins in the absence of Dfm1 and determine that misfolded membrane proteins are toxic to cells in the absence of Dfm1 and cause disruptions to proteasomal and ubiquitin homeostasis.

An ERAD‐independent role for rhomboid pseudoprotease Dfm1 in mediating sphingolipid homeostasis

“…Instead, Dfm1 interacts with Ypk1‐dependent phosphorylated Orm2, which is followed by Orm2 exit from the ER and degradation by EGAD. Our laboratory has recently identified a chaperone‐like Dfm1 function where it influences the solubility of aggregate‐prone misfolded membrane substrates along the ER membrane (preprint: Kandel et al , 2022). Similar to Dfm1's function as a mediator in sphingolipid homeostasis, its chaperone‐like role requires Dfm1's substrate‐binding and lipid‐thinning function, but not its Cdc48 recruitment function.…”

Proteotoxic stress and the ubiquitin proteasome system

“…The transcriptional program orchestrated by Rpn4 in yeast is called the p roteasome s tress r esponse (PSR) [ 131 ], and Nrf1 activates a similar program in mammalian cells called the proteasome recovery pathway [ 130 ]. Our group (the Neal lab at University of California, San Diego) and others have identified Rpn4 as a key mediator in adapting cells to misfolded membrane protein accumulation at the ER [ 132 , 133 ], as well as being crucial in adaptation to stressors that induce protein aggregation and certain forms of proteasome inhibition stress [ 131 ]. Rpn4 is a cytoplasmic protein that is normally rapidly degraded by the proteasome, both in a ubiquitin-dependent and -independent manner [ 134 , 135 ].…”

“…The absence of Ubp6 also increases the sensitivity of cells to treatment with amino acid analogs that cause disruptions to proteostasis [ 88 ]. Our lab has also observed growth stress in Ubp6 KO yeast expressing a misfolded membrane protein, as well as in cells lacking other DUBs, indicating a broad role for DUBs in maintaining Ub homeostasis and in adapting yeast cells to proteotoxic stress [ 132 ].…”

“…The mammalian rhomboid protease RHBDL4 can cleave aggregation-prone proteins and direct the fragments to degradation by the proteasome [ 190 ]. The Neal Lab has recently established the ability of rhomboid pseudoproteases to similarly act on aggregation-prone membrane proteins, with yeast rhomboid pseudoprotease Dfm1 preventing misfolded membrane protein aggregation [ 132 ]. When membrane proteins aggregate in the absence of Dfm1, it is toxic to the cells.…”

“…When membrane proteins aggregate in the absence of Dfm1, it is toxic to the cells. This toxicity appears to be primarily driven by ubiquitin stress, as the free Ub pool is reduced by roughly half and the cells grow normally when Ub expression is increased [ 132 ]. Remarkably, in cells lacking Dfm1, expressing the mammalian homologs of Dfm1, Derlin-1 or Derlin-2, restores the ability of yeast cells to prevent membrane protein aggregation and prevents ubiquitin stress [ 132 ].…”