The yeast ERAD-C ubiquitin ligase Doa10 recognizes an intramembrane degron

Abstract

Figure 4. Investigation of Sbh1/Sbh2 chimeric proteins. Both Sbh2 TM and ER-luminal regions contribute to the degron. (A) Schematic representation of Sbh1, Sbh2, and Sbh1/Sbh2 chimeric proteins used in this study. Sbh1-derived regions are depicted in light blue, and Sbh2-derived regions are shown in black. The nomenclature for Sbh1/Sbh2 chimeric proteins is as follows: "Sbh-XYZ" designates a chimeric protein in which the number at position X indicates the source of the cytosolic domain ("1": Sbh1; or "2": Sbh2), the number at position Y indicates the source of the TM helix, and the number at position Z indicates the source of the ER-luminal domain, respectively. (right) Comparison of Sbh1, Sbh2, and Sbh1/Sbh2 chimera TA sequences. Residues depicted bold in red represent identical residues shared between Sbh1 and Sbh2; residues depicted in light blue are Sbh1-specific residues, and residues depicted in black indicate residues specific for Sbh2; the black horizontal line indicates the TM helix, and the asterisks indicate Sbh2 residues Ser61 and Ser68, respectively. N, N terminal; C, C terminal. (B) Degradation of HA-Sbh-122. chx chase as in Fig. 1

Membrane phospholipid alteration causes chronic ER stress through early degradation of homeostatic ER-resident proteins

“…This suggests that Sbh1 could be dissociated from the Sec61 complex under lipid bilayer stress which leads to its premature degradation. This is consistent with the finding that Sbh2, the paralogue of Sbh1, becomes destabilised and rapidly degraded when unbound to the Sec61-like complex Ssh1 50 . Similarly, Sbh1 was found to interact with proteins of the ERAD pathway under lipid bilayer stress (Fig.…”

“…4a). The temperature-sensitive growth defect of the sbh1 Δ sbh2 Δ mutant at 38 °C can be rescued with either a functional Sbh1 or Sbh2 as previously reported 50 . Sbh1-HA, HA-Sbh1, and TF-C ub -Sbh1 were all sufficient to rescue the growth defect of the sbh1 Δ sbh2 Δ mutant, suggesting that Sbh1 is functional regardless of epitope tag position.…”

Beyond the cell factory: Homeostatic regulation of and by the UPR ER

“…Therefore, proteins that are beyond repair, such as terminally misfolded proteins, are first extracted from the ER by adenosine triphosphate–driven motors and targeted for proteasomal degradation through ER-associated degradation (ERAD). In yeast, where most of the ERAD components have been originally described, transmembrane protein complex including the ubiquitin ligases Hrd1p and Doa10p recognize misfolded proteins and tag them for degradation ( 22 , 23 ). Upon poly-ubiquitylation via the ERAD machinery, the AAA+ adenosine triphosphatase (ATPase) Cdc48p (p97 or valosin-containing protein in humans) drives extraction of the proteins from the ER into the cytosol, where it is subsequently degraded by the proteasome ( 24 ).…”

gp78 functions downstream of Hrd1 to promote degradation of misfolded proteins of the endoplasmic reticulum

“…Thus exceptions to these rules may exist as more substrates are analyzed. Indeed, a recent report showed that Doa10p also recognizes a newly established ERAD substrate bearing a “degron” in the transmembrane region ( Habeck et al ., 2015 ).…”