The Erv41–Erv46 Complex Serves as a Retrograde Receptor to Retrieve Escaped ER Proteins

Signal-dependent sorting of proteins in the early secretory pathway is required for dynamic retention of endoplasmic reticulum (ER) and Golgi components. In this study, we identify the Erv41–Erv46 complex as a new retrograde receptor for retrieval of non–HDEL-bearing ER resident proteins. In cells lacking Erv41–Erv46 function, the ER enzyme glucosidase I (Gls1) was mislocalized and degraded in the vacuole. Biochemical experiments demonstrated that the luminal domain of Gls1 bound to the Erv41–Erv46 complex in a pH-dependent manner. Moreover, in vivo disturbance of the pH gradient across membranes by bafilomycin A1 treatment caused Gls1 mislocalization. Whole cell proteomic analyses of deletion strains using stable isotope labeling by amino acids in culture identified other ER resident proteins that depended on the Erv41–Erv46 complex for efficient localization. Our results support a model in which pH-dependent receptor binding of specific cargo by the Erv41–Erv46 complex in Golgi compartments identifies escaped ER resident proteins for retrieval to the ER in coat protein complex I–formed transport carriers

A role for eisosomes in maintenance of plasma membrane phosphoinositide levels

The plasma membrane delineates the cell and mediates its communication and material exchange with the environment. Many processes of the plasma membrane occur through interactions of proteins with phosphatidylinositol(4,5)-bisphosphate (PI(4,5)P2), which is highly enriched in this membrane and is a key determinant of its identity. Eisosomes function in lateral organization of the plasma membrane, but the molecular function of their major protein subunits, the BAR domain–containing proteins Pil1 and Lsp1, is poorly understood. Here we show that eisosomes interact with the PI(4,5)P2 phosphatase Inp51/Sjl1, thereby recruiting it to the plasma membrane. Pil1 is essential for plasma membrane localization and function of Inp51 but not for the homologous phosphatidylinositol bisphosphate phosphatases Inp52/Sjl2 and Inp53/Sjl3. Consistent with this, absence of Pil1 increases total and available PI(4,5)P2 levels at the plasma membrane. On the basis of these findings, we propose a model in which the eisosomes function in maintaining PI(4,5)P2 levels by Inp51/Sjl1 recruitment

Triacylglycerol Synthesis Enzymes Mediate Lipid Droplet Growth by Relocalizing from the ER to Lipid Droplets

Lipid droplets (LDs) store metabolic energy and membrane lipid precursors. With excess metabolic energy, cells synthesize triacylglycerol (TG) and form LDs that grow dramatically. It is unclear how TG synthesis relates to LD formation and growth. Here, we identify two LD subpopulations: smaller LDs of relatively constant size, and LDs that grow larger. The latter population contains isoenzymes for each step of TG synthesis. Glycerol-3-phosphate acyltransferase 4 (GPAT4), which catalyzes the first and rate-limiting step, relocalizes from the endoplasmic reticulum (ER) to a subset of forming LDs, where it becomes stably associated. ER-to-LD targeting of GPAT4 and other LD-localized TG synthesis isozymes is required for LD growth. Key features of GPAT4 ER-to-LD targeting and function in LD growth are conserved between Drosophila and mammalian cells. Our results explain how TG synthesis is coupled with LD growth and identify two distinct LD subpopulations based on their capacity for localized TG synthesis

Global Proteome Turnover Analyses of the Yeasts S. cerevisiae and S. pombe

Summary: How cells maintain specific levels of each protein and whether that control is evolutionarily conserved are key questions. Here, we report proteome-wide steady-state protein turnover rate measurements for the evolutionarily distant but ecologically similar yeasts, Saccharomyces cerevisiae and Schizosaccharomyces pombe. We find that the half-life of most proteins is much longer than currently thought and determined to a large degree by protein synthesis and dilution due to cell division. However, we detect a significant subset of proteins (∼15%) in both yeasts that are turned over rapidly. In addition, the relative abundances of orthologous proteins between the two yeasts are highly conserved across the 400 million years of evolution. In contrast, their respective turnover rates differ considerably. Our data provide a high-confidence resource for studying protein degradation in common yeast model systems. : Christiano et al. report protein turnover rates for S. cerevisiae and S. pombe. Overall, protein half-life is far less conserved than protein abundance. Protein turnover profiling of hrd1Δ, an endoplasmic-reticulum-associated degradation (ERAD)-defective mutant, identifies candidate substrates for this pathway in budding yeast

Dataset for: Acyl-CoA Binding Domain-Containing Protein 3 Functions as a Scaffold to Organize the Golgi Stacking Proteins and a Rab33b-GAP

Golgin45 plays important roles in Golgi stack assembly and is known to bind both the Golgi stacking protein GRASP55 and Rab2 in the medial-Golgi cisternae. In this study, we sought to further characterize the cisternal adhesion complex using a proteomics approach. We report here that Acyl-CoA Binding Domain Containing 3 (ACBD3) is likely to be a novel binding partner of Golgin45. ACBD3 interacts with Golgin45 via its GOLD domain, and its co-expression significantly increases Golgin45 targeting to the Golgi. Further, ACBD3 recruits TBC1D22, a Rab33b GTPase Activating Protein (GAP), to a large multi-protein complex, containing Golgin45 and GRASP55. These results suggest that ACBD3 may provide a scaffolding to organize the Golgi stacking proteins and a Rab33b-GAP at the medial-Golgi

The Erv41–Erv46 complex serves as a retrograde receptor to retrieve escaped ER proteins

Signal-dependent sorting of proteins in the early secretory pathway is required for dynamic retention of endoplasmic reticulum (ER) and Golgi components. In this study, we identify the Erv41–Erv46 complex as a new retrograde receptor for retrieval of non–HDEL-bearing ER resident proteins. In cells lacking Erv41–Erv46 function, the ER enzyme glucosidase I (Gls1) was mislocalized and degraded in the vacuole. Biochemical experiments demonstrated that the luminal domain of Gls1 bound to the Erv41–Erv46 complex in a pH-dependent manner. Moreover, in vivo disturbance of the pH gradient across membranes by bafilomycin A1 treatment caused Gls1 mislocalization. Whole cell proteomic analyses of deletion strains using stable isotope labeling by amino acids in culture identified other ER resident proteins that depended on the Erv41–Erv46 complex for efficient localization. Our results support a model in which pH-dependent receptor binding of specific cargo by the Erv41–Erv46 complex in Golgi compartments identifies escaped ER resident proteins for retrieval to the ER in coat protein complex I–formed transport carriers