The proteasome is a multicatalytic proteinase complex which is characterized by its ability to cleave peptides with Arg, Phe, Tyr, Leu, and Glu adjacent to the leaving group at neutral or slightly basic pH. The proteasome has an ATP-dependent proteolytic activity. Plays an important role in the regulation of cell proliferation or cell cycle control, transcriptional regulation, immune and stress response, cell differentiation, and apoptosis. Interacts with some important proteins involved in transcription factor regulation, cell cycle transition, viral replication and even tumor initiation and progression. Inhibits the transactivation function of HIF-1A under both normoxic and hypoxia-mimicking conditions. The interaction with EMAP2 increases the proteasome-mediated HIF-1A degradation under the hypoxic conditions. Plays a role in hepatitis C virus internal ribosome entry site-mediated translation. Mediates nuclear translocation of the androgen receptor (AR) and thereby enhances androgen-mediated transactivation. Promotes MAVS degradation and thereby negatively regulates MAVS-mediated innate immune response.
Innate immunity to viruses involves receptors such as Retinoic Acid Induced Gene-1 (RIG-I), which senses viral RNA and triggers a signaling pathway involving the outer mitochondrial membrane protein mitochondrial antiviral signaling (MAVS). Recent work has identified that NLRX1, a member of another class of innate immune receptors, sequesters MAVS away from RIG-I and thereby prevents mitochondrial antiviral immunity. In this study, we demonstrate that the proteasome PSMA7 (alpha4) subunit associates with MAVS in vivo and in vitro. Expression of PSMA7 results in a potent inhibition of RIG-1 and MAVS-mediated IFN-beta promoter activity; conversely, depletion of PSMA7 with small interference RNA enhances virus-induced type I IFN production, with consequent reduction of virus replication. Furthermore, a striking reduction in the abundance of endogenous MAVS with overexpressed PSMA7 was found and virus infection leads to transient increase in the endogenous PSMA7 protein level. Cumulatively, these results suggest that PSMA7 is a negative regulator of the MAVS-mediated innate immunity that probably serves to attenuate the establishment of an antiviral state during viral infection, highlighting the biological significance of PSMA7-MAVS association as an important cellular regulatory control.
Ribozymes are small catalytic RNA molecules that can be engineered to enzymatically cleave RNA transcripts in a sequence-specific fashion and thereby inhibit expression and function of the corresponding gene product. With their simple structures and site-specific cleavage activity, they have been exploited as potential therapeutic agents in a variety of human disorders, including hepatitis C virus (HCV) infection. We have designed a hairpin ribozyme (Rz3'X) targeting the HCV minus-strand replication intermediate at position 40 within the 3'X tail. Surprisingly, Rz3'X was found to induce ganciclovir (GCV)-resistant colonies in a bicistronic cellular reporter system with HCV internal ribosome entry site (IRES)-dependent translation of herpes simplex virus thymidine kinase (TK). Rz3'X-transduced GCV-resistant HeLa reporter cells showed substantially reduced IRES-mediated HCV core protein translation compared with control vector-transduced cells. Since these reporter systems do not contain the HCV 3'X tail sequences, the results indicate that Rz3'X probably exerted an inhibitory effect on HCV IRES activity fortuitously through another gene target. A novel technique of ribozyme cleavage-based target gene identification (cleavage-specific amplification of cDNA ends) (M. Krüger, C. Beger, P. J. Welch, J. R. Barber, and F. Wong-Staal, Nucleic Acids Res. 29:e94, 2001) revealed that human 20S proteasome alpha-subunit PSMA7 mRNA was a target RNA recognized and cleaved by Rz3'X. We then showed that additional ribozymes directed against PSMA7 RNA inhibited HCV IRES activity in two assay systems: GCV resistance in the HeLa IRES TK reporter cell system and a transient transfection assay performed with a bicistronic Renilla-HCV IRES-firefly luciferase reporter in Huh7 cells. In contrast, ribozymes were inactive against IRES of encephalomyocarditis virus and human rhinovirus. Additionally, proteasome inhibitor MG132 exerted a dose-dependent inhibitory effect on HCV IRES-mediated translation but not on cap-dependent translation. These data suggest a principal role for PSMA7 in regulating HCV IRES activity, a function essential for HCV replication.
The hypoxia-inducible factor-1alpha (HIF-1alpha) is an important transcription factor for cellular responses to oxygen tension. It is rapidly degraded under normoxic conditions by the ubiquitin-dependent proteasome pathway. Here we report a critical role of the 20S proteasome subunit PSMA7 in HIF-1alpha regulation. PSMA7 was found to interact specifically with two subdomains of HIF-1alpha. PSMA7 inhibited the transactivation function of HIF-1alpha under both normoxic and hypoxia-mimicking conditions. In addition, we show that the PSMA7-mediated regulation of HIF-1alpha activity is associated with the proteasome pathway.
Protein Pept. Lett. 16, 486-489 (2009)[PubMed:19442227]
Proteasome subunit alpha type 7(PSMA7) is an alpha-type subunit of the 20S proteasome core complex and participates in degrading proteins through ubiquitin-proteasome pathway (UPP) which plays an important role in the regulation of cell proliferation or cell cycle control, transcriptional regulation, immune and stress response, cell differentiation, and apoptosis. Previous studies have demonstrated that PSMA7 can be a target interacting with some important proteins involved in transcription factor regulation, cell cycle transition, viral replication and even tumor initiation and progression, suggesting that PSMA7 could be a potential target for the development of clinical diagnosis and new therapeutic drugs. Here, we review the recent studies on PSMA7 involved in many different cellular processes, ranging from the cell cycle process to antigen processing and tumorigenesis.
Upon binding to androgen, the androgen receptor (AR) can translocate into the nucleus and bind to androgen response element(s) to modulate its target genes. Here we have shown that MG132, a 26 S proteasome inhibitor, suppressed AR transactivation in an androgen-dependent manner in prostate cancer LNCaP and PC-3 cells. In contrast, MG132 showed no suppressive effect on glucocorticoid receptor transactivation. Additionally, transfection of PSMA7, a proteasome subunit, enhanced AR transactivation in a dose-dependent manner. The suppression of AR transactivation by MG132 may then result in the suppression of prostate-specific antigen, a well known marker used to monitor the progress of prostate cancer. Further mechanistic studies indicated that MG132 may suppress AR transactivation via inhibition of AR nuclear translocation and/or inhibition of interactions between AR and its coregulators, such as ARA70 or TIF2. Together, our data suggest that the proteasome system plays important roles in the regulation of AR activity in prostate cancer cells and may provide a unique target site for the development of therapeutic drugs to block androgen/AR-mediated prostate tumor growth.
The detailed mechanism of eukaryotic 20S proteasome assembly is currently unknown. In the present study, we demonstrate that the 20S proteasome subunits alpha4 and alpha7 interact with each other as well as all the alpha-subunits in vivo and in vitro. The N-terminal parts of alpha4 and alpha7 are essential for these newly discovered interactions in vitro. Glycerol gradient centrifugation of soluble extracts of HEK293 cells and Western blot analyses show that several alpha-subunits are found in non-proteasomal low-density fractions. The alpha4 and alpha7 subunits co-immunoprecipitate together from these low-density fractions. The unexpected interaction between alpha4 and alpha7 may provide a molecular basis for the formation of previously reported 13S and 16S assembly intermediates.
Interacting selectively and non-covalently with any protein or protein complex (a complex of two or more proteins that may include other nonprotein molecules).
Evidence
1:
Inferred from Physical InteractionIntAct
The proteasome, a proteolytic complex present in all eukaryotic cells, is part of the ATP-dependent ubiquitin/proteasome pathway. It plays a critical role in the regulation of many physiological processes. The 20 S proteasome, the catalytic core of the 26 S proteasome, is made of four stacked rings of seven subunits each (alpha7beta7beta7alpha7). Here we studied the human 20 S proteasome using proteomics. This led to the establishment of a fine subunit reference map and to the identification of post-translational modifications. We found that the human 20 S proteasome, purified from erythrocytes, exhibited a high degree of structural heterogeneity, characterized by the presence of multiple isoforms for most of the alpha and beta subunits, including the catalytic ones, resulting in a total of at least 32 visible spots after Coomassie Blue staining. The different isoforms of a given subunit displayed shifted pI values, suggesting that they likely resulted from post-translational modifications. We then took advantage of the efficiency of complementary mass spectrometric approaches to investigate further these protein modifications at the structural level. In particular, we focused our efforts on the alpha7 subunit and characterized its N-acetylation and its phosphorylation site localized on Ser(250).
Evidence
2:
Inferred from Physical InteractionIntAct
The detailed mechanism of eukaryotic 20S proteasome assembly is currently unknown. In the present study, we demonstrate that the 20S proteasome subunits alpha4 and alpha7 interact with each other as well as all the alpha-subunits in vivo and in vitro. The N-terminal parts of alpha4 and alpha7 are essential for these newly discovered interactions in vitro. Glycerol gradient centrifugation of soluble extracts of HEK293 cells and Western blot analyses show that several alpha-subunits are found in non-proteasomal low-density fractions. The alpha4 and alpha7 subunits co-immunoprecipitate together from these low-density fractions. The unexpected interaction between alpha4 and alpha7 may provide a molecular basis for the formation of previously reported 13S and 16S assembly intermediates.
Evidence
3:
Inferred from Physical InteractionIntAct
Proteome-scale protein interaction maps are available for many organisms, ranging from bacteria, yeast, worms and flies to humans. These maps provide substantial new insights into systems biology, disease research and drug discovery. However, only a small fraction of the total number of human protein-protein interactions has been identified. In this study, we map the interactions of an unbiased selection of 5026 human liver expression proteins by yeast two-hybrid technology and establish a human liver protein interaction network (HLPN) composed of 3484 interactions among 2582 proteins. The data set has a validation rate of over 72% as determined by three independent biochemical or cellular assays. The network includes metabolic enzymes and liver-specific, liver-phenotype and liver-disease proteins that are individually critical for the maintenance of liver functions. The liver enriched proteins had significantly different topological properties and increased our understanding of the functional relationships among proteins in a liver-specific manner. Our data represent the first comprehensive description of a HLPN, which could be a valuable tool for understanding the functioning of the protein interaction network of the human liver.
Evidence
4:
Inferred from Physical InteractionUniProtKB
The 26S proteasome is a multisubunit protease responsible for regulated proteolysis in eukaryotic cells. It comprises one catalytic 20S proteasome and two axially positioned 19S regulatory complexes. The 20S proteasome is composed of 28 subunits arranged in a cylindrical particle as four heteroheptameric rings, alpha1-7beta1-7beta1-7alpha1-7 (refs 4, 5), but the mechanism responsible for the assembly of such a complex structure remains elusive. Here we report two chaperones, designated proteasome assembling chaperone-1 (PAC1) and PAC2, that are involved in the maturation of mammalian 20S proteasomes. PAC1 and PAC2 associate as heterodimers with proteasome precursors and are degraded after formation of the 20S proteasome is completed. Overexpression of PAC1 or PAC2 accelerates the formation of precursor proteasomes, whereas knockdown by short interfering RNA impairs it, resulting in poor maturation of 20S proteasomes. Furthermore, the PAC complex provides a scaffold for alpha-ring formation and keeps the alpha-rings competent for the subsequent formation of half-proteasomes. Thus, our results identify a mechanism for the correct assembly of 20S proteasomes.
Evidence
5:
Inferred from Physical InteractionIntAct
Immunoproteasomes and standard proteasomes assemble by alternative pathways that bias against the formation of certain "mixed" proteasomes. Differences between beta subunit propeptides contribute to assembly specificity and an assembly chaperone, proteassemblin, may be involved via differential propeptide interactions. We investigated possible mechanisms of biased proteasome assembly and the role of proteassemblin by identifying protein-protein interactions among human 20S proteasome subunits and proteassemblin using a yeast two-hybrid interaction assay. Forty-one interactions were detected, including five involving proteassemblin and contiguous beta subunits, which suggests that proteassemblin binds to preproteasomes via a beta subunit surface. Interaction between proteassemblin and beta5, but not beta5i, suggests that proteassemblin may be involved in the propeptide-dependent differential incorporation of these subunits. Interactions between proteassemblin and beta1, beta1i, and beta7 suggest that proteassemblin may regulate preproteasome dimerization via interactions with the C-termini of these subunits, which in the mature 20S structure extend to contact opposing beta subunit rings.
Catalysis of the hydrolysis of internal peptide bonds in a polypeptide chain by a mechanism in which the hydroxyl group of a threonine residue at the active center acts as a nucleophile.
Viral protein involved in a direct and specific interaction with a host macromolecule. Viruses interact with many cellular pathways to achieve their replication cycle. Entry into the host cell, transport to the viral replication sites or viral budding are all steps that require interaction between the host and the virus. Additionally, the evasion from the host immune response requires a lot of viral proteins to associate with and inhibit cellular proteins with antiviral functions.
Enzyme which catalyzes hydrolysis reaction, i.e. the addition of the hydrogen and hydroxyl ions of water to a molecule with its consequent splitting into two or more simpler molecules.
Proteolytic enzyme with a threonine residue (Thr) in its active site. The prototype members of this class of enzymes are the proteasome catalytic subunits.
A reference proteome is a set of protein sequences derived from a complete proteome which constitutes a defined standard for a particular user community. Reference proteomes are manually defined according to a number of criteria. They cover the proteomes of well- studied model organisms and other proteomes of interest for biomedical and biotechnological research. Reference proteomes have been selected to provide broad coverage of the tree of life, and constitute a representative cross-section of the taxonomic diversity to be found within UniProtKB.
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