ROT1 encodes an essential endoplasmic-reticulum membrane protein that acts as an ER protein-folding chaperone or chaperone-associated assembly factor. Rot1 faces the ER lumen, genetically and physically cooperates with ER folding and glycosylation factors such as Kar2/BiP and Ost3, prevents aggregation in vitro, and supports maturation of secretory and membrane proteins. Its defects cause impaired ER protein folding, reduced N- and O-glycosylation of glycoproteins, cell wall defects, altered actin polarity, and bud-growth phenotypes; these downstream phenotypes are best interpreted as consequences of the core ER proteostasis/glycoprotein maturation role.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005789 endoplasmic reticulum membrane | IBA GO_REF:0000033 | ACCEPT | Summary: ROT1 is an ER membrane protein, and this phylogenetic annotation matches direct experimental localization. Reason: ER membrane localization is a core cellular component for Rot1's luminal folding and glycoprotein maturation function. Supporting Evidence: PMID:16567426 Rot1p was found to be an essential, ER-localized membrane protein facing the lumen. |
| GO:0006458 'de novo' protein folding | IBA GO_REF:0000033 | ACCEPT | Summary: Rot1 supports folding and maturation of proteins entering the secretory pathway. Reason: The core function of Rot1 is ER chaperone activity for newly synthesized secretory and membrane proteins. Supporting Evidence: PMID:18508919 Recombinant Rot1 exhibited antiaggregation activity in vitro, which was partly impaired by a temperature-sensitive rot1-2 mutation. PMID:18508919 In vivo, the rot1-2 mutation caused accelerated degradation of five proteins in the secretory pathway via ER-associated degradation, resulting in a decrease in their cellular levels. |
| GO:0007118 budding cell apical bud growth | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: Bud-growth defects are reproducible consequences of ROT1 inactivation, but they are downstream of ER proteostasis and cell-wall/glycoprotein defects. Reason: Retain as a valid phenotype/process annotation while keeping core function focused on ER folding and glycoprotein maturation. Supporting Evidence: PMID:17606994 First, it is required for the maintenance of apical growth during bud growth. |
| GO:0051082 unfolded protein binding | IBA GO_REF:0000033 | MODIFY | Summary: The evidence supports chaperone activity rather than a generic unfolded protein binding annotation. Reason: GO:0044183 protein folding chaperone better captures Rot1's ER antiaggregation and folding-support activity. Proposed replacements: protein folding chaperone Supporting Evidence: PMID:18508919 Recombinant Rot1 exhibited antiaggregation activity in vitro, which was partly impaired by a temperature-sensitive rot1-2 mutation. |
| GO:0005783 endoplasmic reticulum | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro-derived ER localization is correct but less specific than ER membrane. Reason: Rot1 functions in the ER; the ER membrane term gives the more precise core location. Supporting Evidence: UniProt:Q03691 SUBCELLULAR LOCATION: Endoplasmic reticulum membrane. |
| GO:0005789 endoplasmic reticulum membrane | IEA GO_REF:0000044 | ACCEPT | Summary: UniProtKB subcellular-location mapping to ER membrane is correct. Reason: Rot1 is an ER membrane protein with luminal orientation. Supporting Evidence: UniProt:Q03691 Endoplasmic reticulum membrane; single-pass type I membrane protein. |
| GO:0006458 'de novo' protein folding | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro-based protein-folding annotation is consistent with the primary literature. Reason: Rot1 is an ER chaperone that supports folding/maturation of newly synthesized secretory-pathway proteins. Supporting Evidence: PMID:18508919 Collectively, these results indicate that Rot1 functions as a chaperone in vivo supporting the folding of those proteins. file:yeast/ROT1/ROT1-deep-research-falcon.md In this context, an **ER chaperone** is a factor that promotes protein folding, prevents aggregation, and/or helps assemble or position protein-processing machinery at the ER translocon. |
| GO:0005783 endoplasmic reticulum | HDA PMID:26928762 One library to make them all: streamlining the creation of y... | ACCEPT | Summary: High-throughput ER localization is consistent with direct ER-membrane evidence. Reason: ER is the correct compartment for Rot1 function, although ER membrane is more specific. Supporting Evidence: PMID:26928762 Assignments were: Ambiguous; Below threshold; Bud; Bud neck; Cell periphery; Cytosol; ER; Mitochondria; Nuclear periphery; Nucleolus; Nucleus; Punctate; Vacuole; Vacuole membrane. |
| GO:0005783 endoplasmic reticulum | HDA PMID:14562095 Global analysis of protein localization in budding yeast. | ACCEPT | Summary: High-throughput localization to ER is consistent with direct Rot1 localization studies. Reason: ER localization is core to Rot1 biology. |
| GO:0006457 protein folding | IGI PMID:16874095 Causal links between protein folding in the ER and events al... | ACCEPT | Summary: Genetic interaction evidence supports Rot1 participation in ER protein folding. Reason: The annotation reflects the core protein-folding role of Rot1 in the ER. Supporting Evidence: PMID:16874095 Rot1 is an essential ER membrane protein and may be involved in protein folding. |
| GO:0034975 protein folding in endoplasmic reticulum | IGI PMID:16874095 Causal links between protein folding in the ER and events al... | ACCEPT | Summary: This is the best biological-process term for the core Rot1 process. Reason: Rot1 acts in ER protein folding and cooperates with Kar2/BiP. Supporting Evidence: PMID:16567426 Rot1p associated with Kar2p, especially under conditions of ER stress, and maturation of a model protein, a reduced form of carboxypeptidaseY, was impaired in a kar2-1 rot1-2 double mutant. |
| GO:0005789 endoplasmic reticulum membrane | IDA PMID:16567426 Saccharomyces cerevisiae Rot1p is an ER-localized membrane p... | ACCEPT | Summary: Direct experimental evidence supports ER membrane localization. Reason: Rot1's ER membrane localization is central to its luminal folding and glycosylation-support roles. Supporting Evidence: PMID:16567426 Rot1p was found to be an essential, ER-localized membrane protein facing the lumen. |
| GO:0006458 'de novo' protein folding | IMP PMID:18508919 Saccharomyces cerevisiae Rot1 is an essential molecular chap... | ACCEPT | Summary: Mutant phenotype evidence directly supports Rot1 as an ER chaperone for newly synthesized secretory-pathway proteins. Reason: This captures Rot1's core ER protein-folding function. Supporting Evidence: PMID:18508919 The rot1-2 mutation caused accelerated degradation of five proteins in the secretory pathway via ER-associated degradation, resulting in a decrease in their cellular levels. |
| GO:0006458 'de novo' protein folding | IPI PMID:18508919 Saccharomyces cerevisiae Rot1 is an essential molecular chap... | ACCEPT | Summary: Physical interactions with client proteins support Rot1's chaperone role. Reason: The interaction evidence is directly tied to Rot1-dependent folding of secretory-pathway proteins. Supporting Evidence: PMID:18508919 we demonstrate a physical and probably transient interaction of Rot1 with four of these proteins. |
| GO:0006487 protein N-linked glycosylation | IGI PMID:22492205 The essential endoplasmic reticulum chaperone Rot1 is requir... | ACCEPT | Summary: Rot1 is required for efficient N-glycosylation in sensitized OST backgrounds, especially involving Ost3. Reason: N-glycosylation is a direct core process for Rot1's ER chaperone/assembly-factor function, particularly through the Ost3 branch of the OST machinery. This does not imply Rot1 is itself a catalytic glycosyltransferase subunit. Supporting Evidence: PMID:22492205 OST activity in vitro determined in membranes from rot1-1ost3Delta cells was found to be decreased to 45% compared with wild-type membranes. |
| GO:0006487 protein N-linked glycosylation | IPI PMID:22492205 The essential endoplasmic reticulum chaperone Rot1 is requir... | ACCEPT | Summary: Physical interaction with Ost3 supports Rot1 involvement in the Ost3-containing OST branch of N-glycosylation. Reason: The Rot1-Ost3 physical interaction supports N-glycosylation as a direct core process coupled to Rot1's ER chaperone/assembly-factor role. Supporting Evidence: PMID:22492205 By affinity chromatography, a physical interaction between Rot1 and Ost3 was demonstrated. |
| GO:0007118 budding cell apical bud growth | IMP PMID:17606994 Rot1 plays an antagonistic role to Clb2 in actin cytoskeleto... | KEEP AS NON CORE | Summary: Rot1 inactivation disrupts maintenance of apical bud growth. Reason: This is a valid cellular phenotype/process but not the primary molecular function of Rot1. Supporting Evidence: PMID:17606994 Rot1 affects the actin cytoskeleton during the cell cycle at two levels. First, it is required for the maintenance of apical growth during bud growth. |
| GO:0007118 budding cell apical bud growth | IGI PMID:17606994 Rot1 plays an antagonistic role to Clb2 in actin cytoskeleto... | KEEP AS NON CORE | Summary: Genetic interaction evidence supports the bud-growth phenotype. Reason: The process is downstream of ER proteostasis/cell-wall consequences and is therefore non-core. |
| GO:0009272 fungal-type cell wall biogenesis | IMP PMID:9545237 Cell wall integrity modulates RHO1 activity via the exchange... | KEEP AS NON CORE | Summary: rot1 mutants have cell-wall defects that affect Rho1/Tor2 pathway phenotypes. Reason: Cell wall biogenesis is a supported downstream process, likely mediated by defective maturation/glycosylation of cell-wall proteins. Supporting Evidence: PMID:9545237 rot1, rot2, big1, cwh41, gas1 and fks1 mutations all confer cell wall defects and suppress tor2(ts). |
| GO:0030950 establishment or maintenance of actin cytoskeleton polarity | IMP PMID:17606994 Rot1 plays an antagonistic role to Clb2 in actin cytoskeleto... | KEEP AS NON CORE | Summary: Actin-polarity defects are supported by phenotype analysis of rot1 cells. Reason: This is a downstream morphogenesis consequence rather than the core ER molecular function. Supporting Evidence: PMID:17606994 Rot1 is necessary to polarize actin cytoskeleton to the neck region at the end of mitosis. |
| GO:0035269 protein O-linked glycosylation via mannose | IGI PMID:22492205 The essential endoplasmic reticulum chaperone Rot1 is requir... | KEEP AS NON CORE | Summary: Rot1 perturbation affects O-mannosylation of selected glycoproteins. Reason: This is supported glycosylation biology but is best interpreted as a consequence of Rot1's ER chaperone/assembly role. Supporting Evidence: PMID:22492205 Rot1 was found to be involved also in the O-mannosylation process, as the glycosylation of distinct glycoproteins of this type were affected. |
| GO:0051082 unfolded protein binding | IDA PMID:18508919 Saccharomyces cerevisiae Rot1 is an essential molecular chap... | MODIFY | Summary: Direct antiaggregation data support chaperone activity, but the binding term is too generic. Reason: Replace with GO:0044183 protein folding chaperone to capture the functional chaperone activity. Proposed replacements: protein folding chaperone Supporting Evidence: PMID:18508919 Recombinant Rot1 exhibited antiaggregation activity in vitro, which was partly impaired by a temperature-sensitive rot1-2 mutation. |
| GO:0051082 unfolded protein binding | IMP PMID:18508919 Saccharomyces cerevisiae Rot1 is an essential molecular chap... | MODIFY | Summary: Mutant evidence supports impaired protein-folding support, not merely passive unfolded-protein binding. Reason: GO:0044183 is the better functional term for Rot1's chaperone role. Proposed replacements: protein folding chaperone Supporting Evidence: PMID:18508919 Rot1 functions as a chaperone in vivo supporting the folding of those proteins. |
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Download this section (compressed HTML)Q: Is Rot1 primarily a client-binding ER chaperone, an OST/PMT assembly factor, or a factor that couples both activities during glycoprotein maturation?
Q: Which Rot1-dependent clients directly cause the cell wall and actin-polarity phenotypes when they fail to mature correctly?
Experiment: Compare ER-associated degradation substrates, glycosylation occupancy, and Rot1 proximity-labeling partners in wild-type, rot1 temperature-sensitive, and complemented strains under permissive and restrictive conditions.
Hypothesis: Rot1 has a restricted client set enriched for secretory-pathway and cell-wall glycoproteins.
Type: client proteomics
Experiment: Reconstitute Ost3-containing and Ost6-containing OST complexes with or without purified Rot1 in microsomal or proteoliposome assays and measure substrate-specific N-glycosylation efficiency.
Hypothesis: Rot1 directly improves Ost3-containing OST function through physical association with Ost3 rather than only indirectly through global ER stress.
Type: glycosylation complex reconstitution
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