Ruprecht-Karls-Universität Heidelberg

Sebastian Schuck
ZMBH Research Group Leader

CellNetworks Junior
Group Leader

ZMBH
Im Neuenheimer Feld 282
69120 Heidelberg, Germany
Tel.: + 49-6221 54 6745 (office)
Tel.: + 49-6221 54 6823 (lab)
Fax: +49-6221 54 5894
s.schuck@zmbh.uni-heidelberg.de


Organelle Homeostasis

Biogenesis and Degradation of the Endoplasmic Reticulum


Eukaryotic cells show striking differences in the abundance and architecture of their organelles. Moreover, cells rapidly adjust the size, shape and composition of their organelles to changing physiological demands. This remarkable capacity for adaptation enables cells to maintain homeostasis during differentiation, stress and disease. The underlying molecular mechanisms are fundamental for proper cell function and uncovering them is a fascinating challenge.

We want to elucidate how cells ensure homeostasis of their endoplasmic reticulum (ER). We are asking questions such as: How do cells know how much ER they need? How do they balance biogenesis and degradation to reach optimal ER size? How do they shape ER architecture according to their functional needs? How do they recognize damaged or redundant ER components and selectively eliminate them? To answer these questions, we investigate three related processes: (1) ER membrane biogenesis, which enables organelle expansion and remodelling, (2) ER-phagy, which mediates autophagic organelle degradation, and (3) SHRED, which regulates proteasomal degradation of misfolded cytosolic and ER membrane proteins. We explore these processes in budding yeast but also in mammalian cells.


ER biogenesis

The ER has a complex morphology and many vital functions, for example in protein folding and lipid synthesis. When the ER is unable to fold its load of newly synthesized polypeptides, misfolded proteins accumulate and cause ER stress. Misfolded proteins activate the unfolded protein response (UPR), which increases the protein folding capacity of the ER and induces ER-associated degradation (ERAD). In this way, the UPR promotes the removal of misfolded proteins. Related mechanisms cooperate with the UPR to clear troublesome proteins, such as enhanced proteasome biogenesis (Schmidt et al., 2019). Furthermore, the UPR triggers massive ER membrane expansion by activating lipid synthesis (Figure 1; Schuck et al., 2009). We recently identified genes required for ER expansion and determined how they work together to regulate lipid metabolism and ER membrane biogenesis (Bircham et al., 2020). Professional secretory cells, such as antibody-secreting plasma cells and pancreatic beta cells, need to expand the ER membrane during differentiation. Finding out how cells adjust ER size will therefore not only help us to understand how cells respond to stress but also how they differentiate.




Figure 1. ER expansion. Yeast expressing Sec63-GFP to highlight the peripheral ER (pER) and the nuclear envelope (NE). Yeast exposed to ER stress have a vastly expanded peripheral ER.


ER-phagy

Another response to ER stress is autophagy (cellular self-eating), which mediates degradation of cytoplasmic components in lysosomes. ER stress turns on selective autophagy of ER, which can occur by macroautophagy and microautophagy. Our focus is micro-ER-phagy, which entails a spectacular restructuring of ER into multilamellar whorls and their microautophagic uptake into the yeast lysosomes, called the vacuole (Figure 2; Schuck et al., 2014). We have recently identified the first components of the molecular machinery for micro-ER-phagy and have begun to unravel how it works (Schäfer et al., 2020). Nonetheless, our understanding of micro-ER-phagy is still, well, a litte crude. Through micro-ER-phagy, cells may sacrifice parts of their ER to destroy protein aggregates and damaged organelle membrane. Moreover, when stress has been resolved, micro-ER-phagy may downsize the ER and reverse organelle expansion. Thus, the UPR and ER-phagy work together to refold or degrade damaged proteins and to expand or shrink the ER as needed. Hence, ER-phagy helps maintain ER homeostasis and may be relevant for diseases impinging on ER function, such as cancer and diabetes. We are keen to learn more about the molecular events during autophagy of ER whorls and to define the physiological roles of micro-ER-phagy in both yeast and mammals.




Figure 2. Micro-ER-phagy. ER stress triggers formation of ER whorls in the cytosol (left). These whorls make contact with the membrane of the vacuole, the yeast lysosome (middle). Finally, they are taken up into the lumen of the vacuole (right). Cyto, cytoplasm; VM, vacuole membrane; V, vacuole.


SHRED

Protein folding is error-prone, especially during stress. Cells possess elaborate quality control machinery, including numerous chaperones and ubiquitin ligases, to promote proper folding and degrade folding failures. Stress responses like the UPR tune quality control to current demand. We have recently uncovered a novel stress response pathway termed SHRED, for stress-induced homeostatically regulated protein degradation (Figure 3; Szoradi et al., 2018). SHRED is activated when stress stimulates transcription of the ROQ1 gene. The Roq1 protein is cleaved by the protease Ynm3. Truncated Roq1∆21 binds to the N-end rule ubiquitin ligase Ubr1 as a pseudosubstrate, reprograms its substrate specificity and directs Ubr1 towards misfolded cytosolic and ER membrane proteins. The resulting more stringent quality control enhances stress resistance. Deteriorating protein quality control during aging is a key factor for the onset of neurodegenerative diseases such as Alzheimer’s. Moreover, cancer cells suffer from chronic folding stress and depend on heightened quality control for survival. A deeper understanding of how quality control is regulated may therefore inspire new therapeutic approaches. We aim to understand the precise mechanism and structural basis of Ubr1 reprograming by Roq1 through in vitro reconstitution and to determine if SHRED exists in mammals.




Figure 3. SHRED. Under non-stress conditions, the ubiquitin ligase Ubr1 degrades proteins with positively charged N-terminal residues as part of the N-end rule pathway (left). Under stress conditions, Roq1 is produced, cleaved by Ynm3 and binds to Ubr1 as a pseudosubstrate. This reprograms Ubr1 and stimulates the degradation of misfolded proteins (right).



Selected publications

Bircham PW, Papagiannidis D, Lchterborg C, Ruffini G, Brgger B and Schuck S (2020). Control of endoplasmic reticulum membrane biogenesis by regulators of lipid metabolism. biorxiv (abstract) (PDF)

Schäfer JA, Schessner JP, Bircham PW, Tsuji T, Funaya C, Pajonk O, Schaeff K, Ruffini G, Papagiannidis D, Knop M, Fujimoto T and Schuck S (2020). ESCRT machinery mediates selective microautophagy of endoplasmic reticulum in yeast. EMBO Journal (abstract) (PDF)

Schmidt RM, Schessner JP, Borner GH and Schuck S (2019). The proteasome biogenesis regulator Rpn4 cooperates with the unfolded protein response to promote ER stress resistance. eLife (abstract) (PDF).

Szoradi T, Schaeff K, Garcia-Rivera EM, Itzhak DN, Schmidt RM, Bircham PW, Leiss K, Diaz-Miyar J, Chen VK, Muzzey D, Borner GH and Schuck S (2018). SHRED is a regulatory cascade that reprograms Ubr1 substrate specificity for enhanced protein quality control during stress. Molecular Cell (abstract) (PDF).

Schuck S, Gallagher CM and Walter P (2014). ER-phagy mediates selective degradation of endoplasmic reticulum independently of the core autophagy machinery. Journal of Cell Science (abstract) (PDF).

Schuck S, Prinz WA, Thorn KS, Voss C and Walter P (2009). Membrane expansion alleviates endoplasmic reticulum stress independently of the unfolded protein response. Journal of Cell Biology (abstract) (PDF).





ER homeostasis. (A) The unstressed, happy ER. The ER forms the nuclear envelope and extends outward as the peripheral ER. (B) The ER experiences protein folding stress. The unfolded protein response kicks in and activates genes for chaperones and lipid biosynthesis. The ER expands to make more space for protein folding. (C) The stressed ER. Chaperones and ER-associated degradation do what they can to deal with misfolded proteins. (D) The ER prepares for ER-phagy. Parts of the ER are being rearranged into ER whorls. (E) ER whorls detach from the rest of the ER, reducing ER size and perhaps taking damaged ER with them. (F) The stress is resolved and the ER is back to normal.






















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