Browse
Submit Data
Databases
API
Help

Dataset Information

28 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Beta-Subunit appendages promote 20S proteasome assembly by overcoming an Ump1-dependent checkpoint.

ABSTRACT: Proteasomes are responsible for most intracellular protein degradation in eukaryotes. The 20S proteasome comprises a dyad-symmetric stack of four heptameric rings made from 14 distinct subunits. How it assembles is not understood. Most subunits in the central pair of beta-subunit rings are synthesized in precursor form. Normally, the beta5 (Doa3) propeptide is essential for yeast proteasome biogenesis, but overproduction of beta7 (Pre4) bypasses this requirement. Bypass depends on a unique beta7 extension, which contacts the opposing beta ring. The resulting proteasomes appear normal but assemble inefficiently, facilitating identification of assembly intermediates. Assembly occurs stepwise into precursor dimers, and intermediates contain the Ump1 assembly factor and a novel complex, Pba1-Pba2. beta7 incorporation occurs late and is closely linked to the association of two half-proteasomes. We propose that dimerization is normally driven by the beta5 propeptide, an intramolecular chaperone, but beta7 addition overcomes an Ump1-dependent assembly checkpoint and stabilizes the precursor dimer.

SUBMITTER: Li X

PROVIDER: S-EPMC1864979 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Dissecting beta-ring assembly pathway of the mammalian 20S proteasome.

Project description:The 20S proteasome is the catalytic core of the 26S proteasome. It comprises four stacked rings of seven subunits each, alpha(1-7)beta(1-7)beta(1-7)alpha(1-7). Recent studies indicated that proteasome-specific chaperones and beta-subunit appendages assist in the formation of alpha-rings and dimerization of half-proteasomes, but the process involved in the assembly of beta-rings is poorly understood. Here, we clarify the mechanism of beta-ring formation on alpha-rings by characterizing assembly intermediates accumulated in cells depleted of each beta-subunit. Starting from beta2, incorporation of beta-subunits occurs in an orderly manner dependent on the propeptides of beta2 and beta5, and the C-terminal tail of beta2. Unexpectedly, hUmp1, a chaperone functioning at the final assembly step, is incorporated as early as beta2 and is required for the structural integrity of early assembly intermediates. We propose a model in which beta-ring formation is assisted by both intramolecular and extrinsic chaperones, whose roles are partially different between yeast and mammals.

| S-EPMC2519102 | biostudies-literature

Eukaryotic 20S proteasome catalytic subunit propeptides prevent active site inactivation by N-terminal acetylation and promote particle assembly.

Project description:Proteins targeted for degradation by the ubiquitin-proteasome system are degraded by the 26S proteasome. The core of this large protease is the 20S proteasome, a barrel-shaped structure made of a stack of four heptameric rings. Of the 14 different subunits that make up the yeast 20S proteasome, three have proteolytic active sites: Doa3/beta5, Pup1/beta2 and Pre3/beta1. Each of these subunits is synthesized with an N-terminal propeptide that is autocatalytically cleaved during particle assembly. We show here that the propeptides have both common and distinct functions in proteasome biogenesis. Unlike the Doa3 propeptide, which is crucial for proteasome assembly, the Pre3 and Pup1 propeptides are dispensable for cell viability and proteasome formation. However, mutants lacking these propeptide-encoding elements are defective for specific peptidase activities, are more sensitive to environmental stresses and have subtle defects in proteasome assembly. Unexpectedly, a critical function of the propeptide is the protection of the N-terminal catalytic threonine residue against Nalpha-acetylation. For all three propeptide-deleted subunits, activity of the affected catalytic center is fully restored when the Nat1-Ard1 Nalpha-acetyltransferase is mutated. In addition to delineating a novel function for proteasome propeptides, these data provide the first biochemical evidence for the postulated participation of the alpha-amino group in the proteasome catalytic mechanism.

| S-EPMC1171436 | biostudies-other

Plasticity in eucaryotic 20S proteasome ring assembly revealed by a subunit deletion in yeast.

Project description:The 20S proteasome is made up of four stacked heptameric rings, which in eucaryotes assemble from 14 different but related subunits. The rules governing subunit assembly and placement are not understood. We show that a different kind of proteasome forms in yeast when the Pre9/alpha3 subunit is deleted. Purified pre9Delta proteasomes show a two-fold enrichment for the Pre6/alpha4 subunit, consistent with the presence of an extra copy of Pre6 in each outer ring. Based on disulfide engineering and structure-guided suppressor analyses, Pre6 takes the position normally occupied by Pre9, a substitution that depends on a network of intersubunit salt bridges. When Arabidopsis PAD1/alpha4 is expressed in yeast, it complements not only pre6Delta but also pre6Delta pre9Delta mutants; therefore, the plant alpha4 subunit also can occupy multiple positions in a functional yeast proteasome. Importantly, biogenesis of proteasomes is delayed at an early stage in pre9Delta cells, suggesting an advantage for Pre9 over Pre6 incorporation at the alpha3 position that facilitates correct assembly.

| S-EPMC1271798 | biostudies-literature

Alpha-ring Independent Assembly of the 20S Proteasome.

Project description:Archaeal proteasomes share many features with their eukaryotic counterparts and serve as important models for assembly. Proteasomes are also found in certain bacterial lineages yet their assembly mechanism is thought to be fundamentally different. Here we investigate α-ring formation using recombinant proteasomes from the archaeon Methanococcus maripaludis. Through an engineered disulfide cross-linking strategy, we demonstrate that double α-rings are structurally analogous to half-proteasomes and can form independently of single α-rings. More importantly, via targeted mutagenesis, we show that single α-rings are not required for the efficient assembly of 20S proteasomes. Our data support updating the currently held "α-ring first" view of assembly, initially proposed in studies of archaeal proteasomes, and present a way to reconcile the seemingly separate bacterial assembly mechanism with the rest of the proteasome realm. We suggest that a common assembly network underpins the absolutely conserved architecture of proteasomes across all domains of life.

| S-EPMC4541365 | biostudies-literature

Architecture and assembly of the archaeal Cdc48*20S proteasome.

Project description:ATP-dependent proteases maintain protein quality control and regulate diverse intracellular functions. Proteasomes are primarily responsible for these tasks in the archaeal and eukaryotic domains of life. Even the simplest of these proteases function as large complexes, consisting of the 20S peptidase, a barrel-like structure composed of four heptameric rings, and one or two AAA+ (ATPase associated with a variety of cellular activities) ring hexamers, which use cycles of ATP binding and hydrolysis to unfold and translocate substrates into the 20S proteolytic chamber. Understanding how the AAA+ and 20S components of these enzymes interact and collaborate to execute protein degradation is important, but the highly dynamic nature of prokaryotic proteasomes has hampered structural characterization. Here, we use electron microscopy to determine the architecture of an archaeal Cdc48 ? 20S proteasome, which we stabilized by site-specific cross-linking. This complex displays coaxial alignment of Cdc48 and 20S and is enzymatically active, demonstrating that AAA+ unfoldase wobbling with respect to 20S is not required for function. In the complex, the N-terminal domain of Cdc48, which regulates ATP hydrolysis and degradation, packs against the D1 ring of Cdc48 in a coplanar fashion, constraining mechanisms by which the N-terminal domain alters 20S affinity and degradation activity.

| S-EPMC4035981 | biostudies-literature

Amyloid-β peptide is a substrate of the human 20S proteasome.

Project description:Intraneuronal accumulation of ubiquitin conjugates is a pathological feature of neurodegenerative disorders such as Alzheimer's disease (AD). Previous reports propose that accumulation of ubiquitinated species in AD is a result of inhibition of proteasomal activity by amyloid-β (Aβ) peptides, which leads to blocking of ubiquitin-dependent protein degradation by the proteasome. Here, we provide additional insight into proteasomal dysfunction by Aβ peptides by revealing that aggregated forms of Aβ(1-42) peptides (especially small oligomers) are, in fact, competitive substrates for the chymotrypsin-like activity of the human 20S (h20S) proteasome. In addition to examining the kinetics of the h20S proteasome activity in the presence or absence of Aβ peptides, we use gel electrophoresis, LC-MS, and TOF-MS/MS analyses to examine the degradation of Aβ(1-42) by the h20S proteasome. The observed peptide fragments resulting from proteolytic cleavage of Aβ were consistent with predicted cleavage sites from proteasome degradation. These results support that the interaction of Aβ peptides with the proteasome may play a mechanistic role in proteasomal dysfunction in AD pathology. These results may also reveal a previously unknown natural pathway for clearance of Aβ in normal or diseased cells.

| S-EPMC2992454 | biostudies-literature

An evolutionarily distinct chaperone promotes 20S proteasome α-ring assembly in plants.

Project description:The core protease (CP) subcomplex of the 26S proteasome houses the proteolytic active sites and assumes a barrel shape comprised of four co-axially stacked heptameric rings formed by structurally related α- and β-subunits. CP biogenesis typically begins with the assembly of the α-ring, which then provides a template for β-subunit integration. In eukaryotes, α-ring assembly is partially mediated by two hetero-dimeric chaperones, termed Pba1-Pba2 (Add66) and Pba3-Pba4 (also known as Irc25-Poc4) in yeast. Pba1-Pba2 initially promotes orderly recruitment of the α-subunits through interactions between their C-terminal HbYX or HbF motifs and pockets at the α5-α6 and α6-α7 interfaces. Here, we identified PBAC5 as a fifth α-ring assembly chaperone in Arabidopsis that directly binds the Pba1 homolog PBAC1 to form a trimeric PBAC5-PBAC1-PBAC2 complex. PBAC5 harbors a HbYX motif that docks with a pocket between the α4 and α5 subunits during α-ring construction. Arabidopsis lacking PBAC5, PBAC1 and/or PBAC2 are hypersensitive to proteotoxic, salt and osmotic stresses, and display proteasome assembly defects. Remarkably, whereas PBAC5 is evolutionarily conserved among plants, sequence relatives are also dispersed within other kingdoms, including a scattered array of fungal, metazoan and oomycete species.

| S-EPMC7657472 | biostudies-literature

The dominant temperature-sensitive lethal DTS7 of Drosophila melanogaster encodes an altered 20S proteasome beta-type subunit.

Project description:Proteasomes are multicatalytic complexes that function as the major proteolytic machinery in regulated protein degradation. The eukaryotic 20S proteasome proteolytic core structure comprises 14 different subunits: 7 alpha-type and 7 beta-type. DTS7 is a dominant temperature-sensitive (DTS) lethal mutation at 29 degrees that also acts as a recessive lethal at ambient temperatures. DTS7 maps to cytological position 71AB. Molecular characterization of DTS7 reveals that this is caused by a missense mutation in a beta-type subunit gene, beta2. A previously characterized DTS mutant, l(3)73Ai1, results from a missense mutation in another beta-type subunit gene, beta6. These two mutants share a very similar phenotype, show a strong allele-specific genetic interaction, and are rescued by the same extragenic suppressor, Su(DTS)-1. We propose that these mutants might act as "poison subunits," disrupting proteasome function in a dosage-dependent manner, and suggest how they may interact on the basis of the structure of the yeast 20S proteasome.

| S-EPMC1460450 | biostudies-other

Competitive Metabolite Profiling of Natural Products Reveals Subunit Specific Inhibitors of the 20S Proteasome.

Project description:We have developed a syringolin-based chemical probe and explored its utility for the profiling of metabolite extracts as potent inhibitors of the 20S proteasome. Activity-guided fractionation by competitive labeling allowed us to isolate and identify glidobactin A and C as well as luminmycin A from a Burkholderiales strain. The natural products exhibited unique subunit specificities for the proteolytic subunits of human and mouse constitutive and immunoproteasome in the lower nanomolar range. In particular, glidobactin C displayed an unprecedented ?2/?5 coinhibition profile with single-digit nanomolar potency in combination with sufficiently high cell permeability. These properties render glidobactin C a promising live cell proteasome inhibitor with potent activity against human breast cancer cell lines and comparably low immunotoxicity.

| S-EPMC7047272 | biostudies-literature

Proteasome assembly defect due to a proteasome subunit beta type 8 (PSMB8) mutation causes the autoinflammatory disorder, Nakajo-Nishimura syndrome.

Project description:Nakajo-Nishimura syndrome (NNS) is a disorder that segregates in an autosomal recessive fashion. Symptoms include periodic fever, skin rash, partial lipomuscular atrophy, and joint contracture. Here, we report a mutation in the human proteasome subunit beta type 8 gene (PSMB8) that encodes the immunoproteasome subunit ?5i in patients with NNS. This G201V mutation disrupts the ?-sheet structure, protrudes from the loop that interfaces with the ?4 subunit, and is in close proximity to the catalytic threonine residue. The ?5i mutant is not efficiently incorporated during immunoproteasome biogenesis, resulting in reduced proteasome activity and accumulation of ubiquitinated and oxidized proteins within cells expressing immunoproteasomes. As a result, the level of interleukin (IL)-6 and IFN-? inducible protein (IP)-10 in patient sera is markedly increased. Nuclear phosphorylated p38 and the secretion of IL-6 are increased in patient cells both in vitro and in vivo, which may account for the inflammatory response and periodic fever observed in these patients. These results show that a mutation within a proteasome subunit is the direct cause of a human disease and suggest that decreased proteasome activity can cause inflammation.

| S-EPMC3169106 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data