| 2004 |
Mammalian Ecm29 is present exclusively on 26S proteasomes in HeLa cells and localizes to the centrosome and a subset of secretory compartments including endosomes, the ER, and the ERGIC, as determined by confocal immunofluorescence microscopy and glycerol-gradient fractionation. Ecm29 is up-regulated 2–3-fold in toxin-resistant CHO cells with increased ER-associated degradation, suggesting it couples 26S proteasomes to secretory compartments engaged in quality control. |
Antibody generation, glycerol-gradient fractionation, confocal immunofluorescence microscopy, CHO mutant cell line analysis |
The Journal of biological chemistry |
Medium |
15496406
|
| 2010 |
Ecm29 controls the integrity of RP-CP (regulatory particle–core particle) assemblies in yeast, recognizing RP-CP species in which CP maturation is stalled due to the lack of distinct beta subunits. Reconstitution assays demonstrated that Ecm29 functions as a scaffold protein during remodeling of incompletely matured RP-CP assemblies into regular enzymes, and is degraded upon completion of CP maturation. |
Yeast genetics, proteasome reconstitution assays, biochemical fractionation |
Molecular cell |
High |
20620957
|
| 2010 |
The C-terminal half of human Ecm29 binds myosins and kinesins (molecular motors), while the N-terminal region binds endocytic proteins Vps11, Rab11-FIP4, and rabaptin. A central fragment of Ecm29 is sufficient for 26S proteasome binding. Ecm29-26S proteasomes are present on flotillin-positive endosomes but absent from caveolin- and clathrin-decorated endosomes. Expression of the central fragment reduces proteasome association with flotillin-positive endosomes. |
Genome-wide two-hybrid screens, mass spectrometry, confocal microscopy, glycerol-gradient fractionation, domain mapping with truncation constructs |
The Journal of biological chemistry |
High |
20682791
|
| 2011 |
Not4 E3 ligase associates with the regulatory particle (RP) of the proteasome holoenzyme and interacts with Ecm29. In the absence of Not4, Ecm29 interacts less well with the proteasome, becomes ubiquitinated and degraded. This indicates that appropriate Ecm29-proteasome interaction requires Not4, establishing Ecm29 as a proteasome chaperone whose stability depends on Not4-mediated regulation. |
Yeast genetics (deletion strains), co-immunoprecipitation, ubiquitination assay, proteasome purification and activity assays |
Molecular and cellular biology |
Medium |
21321079
|
| 2011 |
Ecm29 is enriched on proteasomes purified from Saccharomyces cerevisiae rpt5-Δ3 strains (faulty proteasomes), and these Ecm29-containing proteasomes have reduced suc-LLVY-AMC hydrolytic activity. Deletion of ECM29 rescues phenotypes of rpt5-Δ3 and nas2Δ in an hsm3Δ background, establishing Ecm29 as a negative regulator/inhibitor that recognizes and inhibits aberrant proteasomes. |
Yeast genetics (deletion strains, epistasis analysis), proteasome purification, peptidase activity assay |
The Journal of biological chemistry |
Medium |
21878651
|
| 2013 |
Ecm29 inhibits proteasomal ATPase activity and induces a closed conformation of the substrate entry channel of the 20S core particle. Besides inhibiting peptide substrate cleavage in vitro, Ecm29 inhibits degradation of ubiquitin-dependent and -independent substrates in vivo. Chemical cross-linking identified that Ecm29 binds to or in close proximity to proteasomal ATPase subunit Rpt5. Ecm29 preferentially associates with mutant and nucleotide-depleted proteasomes. |
In vitro peptidase and ATPase activity assays, in vivo ubiquitin-dependent substrate degradation assays, chemical cross-linking mass spectrometry, electron microscopy (gate conformation) |
The Journal of biological chemistry |
High |
23995839
|
| 2013 |
Depletion of Ecm29 increases abundance of TLR3 and increases levels of LC3β and p62 (autophagy mediators). Loss of Ecm29 enhances TLR3 signaling, characterized by increased TRAF3 levels, increased phosphorylation of effector kinases, increased nuclear localization of IRF3, and accumulation of signaling molecules at juxtanuclear recycling endosomes. This establishes that Ecm29-associated proteasomes play a role in mediating autophagy and attenuation of TLR3-dependent signaling. |
siRNA depletion of Ecm29 in HEK-293 and HeLa cells, immunoblotting, immunofluorescence microscopy |
Science signaling |
Medium |
24084648
|
| 2016 |
Yeast Ecm29 requires the phosphorylated C-terminal tail of the proteasome α7 subunit for its association with the proteasome. This represents the first example of phosphorylation-dependent binding of a proteasome regulatory factor. The ability of Ecm29 to bind mutant proteasomes requires both the α7-tail phosphorylation-dependent binding site and a previously identified Rpt5 binding site; the need for two binding sites on different subcomplexes explains the specificity of Ecm29 for proteasome holoenzymes. |
Yeast genetics (α7 tail phosphorylation mutants), co-immunoprecipitation/pulldown, proteasome purification |
Scientific reports |
Medium |
27302526
|
| 2016 |
KIAA0368-deficient (Ecm29 knockout) mice show normal peptidase activity and proteasome formation under basal conditions, but fail to dissociate 26S proteasomes under oxidative stress conditions where wild-type cells dissociate 26S proteasomes. This correlates with reduced degradation of damaged proteins and increased resistance to oxidative stress in KIAA0368-deficient cells, establishing Ecm29 as required for 26S proteasome dissociation under stress. |
KIAA0368-deficient mouse generation, native gel proteasome analysis, peptidase activity assays, oxidative stress challenge |
Journal of biochemistry |
Medium |
26802743
|
| 2017 |
Human Ecm29 is the main proteasome-interacting protein responsible for stress-triggered (H2O2) disassembly of the 26S proteasome. Cross-linking mass spectrometry mapped Ecm29 interactions within itself and with proteasome subunits; integrative structural modeling proposed that Ecm29 intrudes on the interaction between the 20S core particle and the 19S regulatory particle, disrupting proteasome structure in response to oxidative stress. Stress-induced 26S disassembly is conserved from yeast to human. |
XAP (in vivo cross-linking-assisted affinity purification) with SILAC quantitative MS, disuccinimidyl sulfoxide cross-linking MS, integrative structural modeling |
The Journal of biological chemistry |
High |
28821611
|
| 2020 |
Ecm29 mediates proteasome distribution to the axon initial segment (AIS) through interaction with the AIS scaffold protein ankyrin G; both the Ecm29 N-terminal domain and an intact AIS structure are required for transport and tethering of proteasomes in the AIS region. Ecm29 knockout in neurons increases NKCC1 protein density in the AIS, delays the GABAergic response switch, and increases action potential firing frequency at early postnatal ages. Ecm29 KO neurons show accelerated AIS developmental positioning, rescued by ectopic Ecm29 expression or NKCC1 inhibition. |
Ecm29 knockout mouse, confocal and live imaging, domain deletion constructs, electrophysiology, chemical convulsive seizure assay, rescue experiments |
The Journal of cell biology |
High |
31910261
|
| 2021 |
Ecm29 regulates proteasome distribution between the centrosome and cell cortex in B cells. Silencing Ecm29 shifts the proteasome from the centrosome to the cell cortex and immunological synapse (IS), resulting in increased F-actin at the centrosome, impaired centrosome and lysosome repositioning to the IS, defective antigen extraction and presentation, decreased actin retrograde flow in lamellipodia, and enhanced spreading. This establishes that Ecm29-controlled asymmetric proteasome distribution coordinates actin dynamics at the centrosome and IS. |
siRNA silencing of Ecm29 in B cells, confocal microscopy, actin dynamics measurement, antigen extraction and presentation assays |
Frontiers in cell and developmental biology |
Medium |
34055780
|
| 2023 |
ECPAS mediates 26S proteasome disassembly into 20S and 19S subcomplexes in response to glucose starvation. Loss of ECPAS abrogates 26S dissociation and reduces degradation of 20S proteasome substrates including puromycylated polypeptides. In silico modeling suggests ECPAS conformational changes initiate the disassembly process. ECPAS is also essential for ER stress response and cell survival during glucose starvation. Elevated 20S proteasome levels are found in glucose-deprived tumors in vivo in a xenograft model. |
ECPAS KO cells, subcomplex affinity purification, quantitative mass spectrometry, in silico structural modeling, xenograft mouse model, puromycylated polypeptide degradation assay |
Cell reports |
High |
37384533
|
| 2023 |
PA200 and ECPAS work cooperatively during spermatogenesis; double knockout (dKO) mice are infertile with considerably reduced proteasome activity in testes and epididymides. In epididymal sperm, ECPAS localizes to the acrosome. Mass spectrometric analysis identified LPIN1 as a target protein for both PA200 and ECPAS (confirmed by immunoblotting and immunostaining). dKO sperm display disorganization of the mitochondrial sheath. |
Double-knockout mouse generation, mass spectrometry, immunoblotting, immunostaining, ultrastructural analysis, proteasome activity assay |
Biomolecules |
Medium |
37189334
|
| 2025 |
Loss of Ecm29 in oligodendroglia or microglia (inducible conditional KO) increases susceptibility to experimental autoimmune encephalomyelitis (EAE) and reduces regulatory T cell populations in the spinal cord. Immunopeptidome profiling identified self-antigens (including NDUFA1p) whose generation fidelity depends on Ecm29/proteasome function, and intraspinal AAV-mediated expression of NDUFA1p ameliorates EAE and expands NDUFA1p-recognizing CD103+CD8+CD122+ Treg cells, establishing that Ecm29-controlled proteasome activity governs CNS immune tolerance through neuroglial antigen generation. |
Inducible conditional Ecm29 KO mice, EAE model, immunopeptidome profiling (MS), AAV-mediated antigen expression, flow cytometry |
Cell reports |
Medium |
39786996
|